A novel multigene expression construct for modification of glycerol metabolism in Yarrowia lipolytica

PubMed Central

2013-01-01

Background High supply of raw, residual glycerol from biodiesel production plants promote the search for novel biotechnological methods of its utilization. In this study we attempted modification of glycerol catabolism in a nonconventional yeast species Yarrowia lipolytica through genetic engineering approach. Results To address this, we developed a novel genetic construct which allows transferring three heterologous genes, encoding glycerol dehydratase, its reactivator and a wide-spectrum alcohol oxidoreductase under the control of glycerol-induced promoter. The three genes, tandemly arrayed in an expression cassette with a marker gene ura3, regulatory and targeting sequences (G3P dh promoter and XPR-like terminator, 28S rDNA as a target locus), were transferred into Yarrowia lipolytica cells. The obtained recombinant strain NCYC3825 was characterized at the molecular level and with respect to its biotechnological potential. Our experiments indicated that the novel recombinant strain stably borne one copy of the expression cassette and efficiently expressed heterologous alcohol oxidoreductase, while glycerol dehydratase and its reactivator were expressed at lower level. Comparative shake flask cultivations in glucose- and glycerol-based media demonstrated higher biomass production by the recombinant strain when glycerol was the main carbon source. During bioreactor (5 L) fed-batch cultivation in glycerol-based medium, the recombinant strain was characterized by relatively high biomass and lipids accumulation (up to 42 gDCW L-1, and a peak value of 38%LIPIDS of DCW, respectively), and production of high titers of citric acid (59 g L-1) and 2-phenylethanol (up to 1 g L-1 in shake flask cultivation), which are industrially attractive bioproducts. Conclusions Due to heterogeneous nature of the observed alterations, we postulate that the main driving force of the modified phenotype was faster growth in glycerol-based media, triggered by modifications in the red

Fatty alcohol production in Lipomyces starkeyi and Yarrowia lipolytica

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Wei; Wei, Hui; Knoshaug, Eric

Current biological pathways to produce biofuel intermediates amenable to separations and catalytic upgrading to hydrocarbon fuels are not cost effective. Previously, oleaginous yeasts have been investigated primarily for lipid production. However, yeasts store neutral lipids intracellularly making recovery difficult and expensive. In addition, once recovered from the cells, lipids are difficult to blend directly with the existing fuels without upgrading. We have, therefore, begun to investigate secreted fatty acid-derived products which can be easily recovered and upgraded to fuels. In this study, we successfully demonstrate the production of fatty alcohols by the oleaginous yeasts, Yarrowia lipolytica and Lipomyces starkeyi, throughmore » expression of the fatty acyl-CoA reductase gene from Marinobactor aquaeolei VT8. This strategy resulted in the production of 167 and 770 mg/L of fatty alcohols in shake flask from Y. lipolytica and L starkeyi, respectively. When using a dodecane overlay during fermentation, 92 and 99% of total fatty alcohols produced by Y. lipolytica and L. starkeyi, respectively, were extracted into the dodecane phase, which compares favorably to the 3 and 50% recovered, respectively, without the dodecane layer. In both oleaginous yeasts, long chain length, saturated fatty alcohols, i.e., hexadecanol (C16:0) and octadecanol (C18:0), were predominant and accounted for more than 85% of the total fatty alcohols produced. To the best of our knowledge, this is the first report of fatty alcohol production in L. starkeyi. Furthermore, this work demonstrates that the oleaginous yeasts, Y. lipolytica and L. starkeyi, can serve as platform organisms for the production of fatty acid-derived biofuels and bioproducts.« less

Fatty alcohol production in Lipomyces starkeyi and Yarrowia lipolytica

DOE PAGES

Wang, Wei; Wei, Hui; Knoshaug, Eric; ...

2016-10-24

Current biological pathways to produce biofuel intermediates amenable to separations and catalytic upgrading to hydrocarbon fuels are not cost effective. Previously, oleaginous yeasts have been investigated primarily for lipid production. However, yeasts store neutral lipids intracellularly making recovery difficult and expensive. In addition, once recovered from the cells, lipids are difficult to blend directly with the existing fuels without upgrading. We have, therefore, begun to investigate secreted fatty acid-derived products which can be easily recovered and upgraded to fuels. In this study, we successfully demonstrate the production of fatty alcohols by the oleaginous yeasts, Yarrowia lipolytica and Lipomyces starkeyi, throughmore » expression of the fatty acyl-CoA reductase gene from Marinobactor aquaeolei VT8. This strategy resulted in the production of 167 and 770 mg/L of fatty alcohols in shake flask from Y. lipolytica and L starkeyi, respectively. When using a dodecane overlay during fermentation, 92 and 99% of total fatty alcohols produced by Y. lipolytica and L. starkeyi, respectively, were extracted into the dodecane phase, which compares favorably to the 3 and 50% recovered, respectively, without the dodecane layer. In both oleaginous yeasts, long chain length, saturated fatty alcohols, i.e., hexadecanol (C16:0) and octadecanol (C18:0), were predominant and accounted for more than 85% of the total fatty alcohols produced. To the best of our knowledge, this is the first report of fatty alcohol production in L. starkeyi. Furthermore, this work demonstrates that the oleaginous yeasts, Y. lipolytica and L. starkeyi, can serve as platform organisms for the production of fatty acid-derived biofuels and bioproducts.« less

Metabolic peculiarities of the citric acid overproduction from glucose in yeasts Yarrowia lipolytica.

PubMed

Kamzolova, Svetlana V; Morgunov, Igor G

2017-11-01

Comparative study of 43 natural yeast strains belonging to 20 species for their capability for overproduction of citric acid (CA) from glucose under nitrogen limitation of cell growth was carried out. As a result, natural strain Yarrowia lipolytica VKM Y-2373 was selected. The effect of growth limitation by biogenic macroelements (nitrogen, phosphorus, or sulfur) on the CA production by the selected strain was studied. It was shown that yeasts Y. lipolytica grown under deficiency of nitrogen, phosphorus, or sulfur were able to excrete CA in industrially sufficient amounts (80-85g/L with the product yield (Y CA ) of 0.70-0.75g/g and the process selectivity of 92.5-95.3%). Based on the obtained data on activities of enzymes involved in the initial stages of glucose oxidation, the cycle of tricarboxylic acids, and the glyoxylate cycle, the conception of the mechanism responsible for the CA overproduction from glucose in Y. lipolytica was formulated. Copyright © 2017 Elsevier Ltd. All rights reserved.

Impacts of environmental conditions on product formation and morphology of Yarrowia lipolytica.

PubMed

Timoumi, Asma; Guillouet, Stéphane E; Molina-Jouve, Carole; Fillaudeau, Luc; Gorret, Nathalie

2018-05-01

The yeast Yarrowia lipolytica is an industrially important microorganism with distinctive physiological and metabolic characteristics. A variety of external factors (e.g., pH, temperature, and nutrient availability) influences the behavior of the yeast and may act as stress conditions which the cells must withstand and adapt. In this mini review, the impacts of environmental factors on the morphology and metabolite production by Y. lipolytica are summarized. In this regard, detailed insights into the effectors involved in the dimorphic transition of Y. lipolytica, the cultivation conditions employed, as well as the methods applied for the morphological characterization are highlighted. Concerning the metabolism products, a special focus is addressed on lipid and citric acid metabolites which have attracted significant attention in recent years. The dependence of lipid and citric acid productivity on key process parameters, such as media composition and physico-chemical variables, is thoroughly discussed. This review attempts to provide a recent update on the topic and will serve as a meaningful resource for researchers working in the field.

Droplet-based microfluidic high-throughput screening of heterologous enzymes secreted by the yeast Yarrowia lipolytica.

PubMed

Beneyton, Thomas; Thomas, Stéphane; Griffiths, Andrew D; Nicaud, Jean-Marc; Drevelle, Antoine; Rossignol, Tristan

2017-01-31

Droplet-based microfluidics is becoming an increasingly attractive alternative to microtiter plate techniques for enzymatic high-throughput screening (HTS), especially for exploring large diversities with lower time and cost footprint. In this case, the assayed enzyme has to be accessible to the substrate within the water-in-oil droplet by being ideally extracellular or displayed at the cell surface. However, most of the enzymes screened to date are expressed within the cytoplasm of Escherichia coli cells, which means that a lysis step must take place inside the droplets for enzyme activity to be assayed. Here, we take advantage of the excellent secretion abilities of the yeast Yarrowia lipolytica to describe a highly efficient expression system particularly suitable for the droplet-based microfluidic HTS. Five hydrolytic genes from Aspergillus niger genome were chosen and the corresponding five Yarrowia lipolytica producing strains were constructed. Each enzyme (endo-β-1,4-xylanase B and C; 1,4-β-cellobiohydrolase A; endoglucanase A; aspartic protease) was successfully overexpressed and secreted in an active form in the crude supernatant. A droplet-based microfluidic HTS system was developed to (a) encapsulate single yeast cells; (b) grow yeast in droplets; (c) inject the relevant enzymatic substrate; (d) incubate droplets on chip; (e) detect enzymatic activity; and (f) sort droplets based on enzymatic activity. Combining this integrated microfluidic platform with gene expression in Y. lipolytica results in remarkably low variability in the enzymatic activity at the single cell level within a given monoclonal population (<5%). Xylanase, cellobiohydrolase and protease activities were successfully assayed using this system. We then used the system to screen for thermostable variants of endo-β-1,4-xylanase C in error-prone PCR libraries. Variants displaying higher thermostable xylanase activities compared to the wild-type were isolated (up to 4.7-fold improvement

Metabolic engineering of oleaginous yeast Yarrowia lipolytica for limonene overproduction.

PubMed

Cao, Xuan; Lv, Yu-Bei; Chen, Jun; Imanaka, Tadayuki; Wei, Liu-Jing; Hua, Qiang

2016-01-01

Limonene, a monocyclic monoterpene, is known for its using as an important precursor of many flavoring, pharmaceutical, and biodiesel products. Currently, d-limonene has been produced via fractionation from essential oils or as a byproduct of orange juice production, however, considering the increasing need for limonene and a certain amount of pesticides may exist in the limonene obtained from the citrus industry, some other methods should be explored to produce limonene. To construct the limonene synthetic pathway in Yarrowia lipolytica , two genes encoding neryl diphosphate synthase 1 (NDPS1) and limonene synthase (LS) were codon-optimized and heterologously expressed in Y. lipolytica . Furthermore, to maximize limonene production, several genes involved in the MVA pathway were overexpressed, either in different copies of the same gene or in combination. Finally with the optimized pyruvic acid and dodecane concentration in flask culture, a maximum limonene titer and content of 23.56 mg/L and 1.36 mg/g DCW were achieved in the final engineered strain Po1f-LN-051, showing approximately 226-fold increase compared with the initial yield 0.006 mg/g DCW. This is the first report on limonene biosynthesis in oleaginous yeast Y. lipolytica by heterologous expression of codon-optimized tLS and tNDPS1 genes. To our knowledge, the limonene production 23.56 mg/L, is the highest limonene production level reported in yeast. In short, we demonstrate that Y. lipolytica provides a compelling platform for the overproduction of limonene derivatives, and even other monoterpenes.

Optimization of odd chain fatty acid production by Yarrowia lipolytica.

PubMed

Park, Young-Kyoung; Dulermo, Thierry; Ledesma-Amaro, Rodrigo; Nicaud, Jean-Marc

2018-01-01

Odd chain fatty acids (odd FAs) have a wide range of applications in therapeutic and nutritional industries, as well as in chemical industries including biofuel. Yarrowia lipolytica is an oleaginous yeast considered a preferred microorganism for the production of lipid-derived biofuels and chemicals. However, it naturally produces negligible amounts of odd chain fatty acids. The possibility of producing odd FAs using Y. lipolytica was investigated. Y. lipolytica wild-type strain was shown able to grow on weak acids; acetate, lactate, and propionate. Maximal growth rate on propionate reached 0.24 ± 0.01 h -1 at 2 g/L, and growth inhibition occurred at concentration above 10 g/L. Wild-type strain accumulated lipids ranging from 7.39 to 8.14% (w/w DCW) depending on the carbon source composition, and odd FAs represented only 0.01-0.12 g/L. We here proved that the deletion of the PHD1 gene improved odd FAs production, which reached a ratio of 46.82% to total lipids. When this modification was transferred to an obese strain, engineered for improving lipid accumulation, further increase odd FAs production reaching a total of 0.57 g/L was shown. Finally, a fed-batch co-feeding strategy was optimized for further increase odd FAs production, which generated 0.75 g/L, the best production described so far in Y. lipolytica . A Y. lipolytica strain able to accumulate high level of odd chain fatty acids, mainly heptadecenoic acid, has been successfully developed. In addition, a fed-batch co-feeding strategy was optimized to further improve lipid accumulation and odd chain fatty acid content. These lipids enriched in odd chain fatty acid can (1) improve the properties of the biodiesel generated from Y. lipolytica lipids and (2) be used as renewable source of odd chain fatty acid for industrial applications. This work paves the way for further improvements in odd chain fatty acids and fatty acid-derived compound production.

YALI0E32769g (DGA1) and YALI0E16797g (LRO1) encode major triacylglycerol synthases of the oleaginous yeast Yarrowia lipolytica.

PubMed

Athenstaedt, Karin

2011-10-01

The oleaginous yeast Yarrowia lipolytica has an outstanding capacity to produce and store triacylglycerols resembling adipocytes of higher eukaryotes. Here, the identification of two genes YALI0E32769g (DGA1) and YALI0E16797g (LRO1) encoding major triacylglycerol synthases of Yarrowia lipolytica is reported. Heterologous expression of either DGA1 or LRO1 in a mutant of the budding yeast Saccharomyces cerevisiae defective in triacylglycerol synthesis restores the formation of this neutral lipid. Whereas Dga1p requires acyl-CoA as a substrate for acylation of diacylglycerol, Lro1p is an acyl-CoA independent triacylglycerol synthase using phospholipids as acyl-donor. Growth of Yarrowia lipolytica strains deleted of DGA1 and/or LRO1 on glucose containing medium significantly decreases triacylglycerol accumulation. Most interestingly, when oleic acid serves as the carbon source the ratio of triacylglycerol accumulation in mutants to wild-type is significantly increased in strains defective in DGA1 but not in lro1Δ. In vitro experiments revealed that under these conditions an additional acyl-CoA dependent triacylglycerol synthase contributes to triacylglycerol synthesis in the respective mutants. Taken together, evidence is provided that Yarrowia lipolytica contains at least four triacylglycerol synthases, namely Lro1p, Dga1p and two additional triacylglycerol synthases whereof one is acyl-CoA dependent and specifically induced upon growth on oleic acid. Copyright © 2011 Elsevier B.V. All rights reserved.

Yarrowia lipolytica morphological mutant enables lasting in situ immobilization in bioreactor.

PubMed

Vandermies, Marie; Kar, Tambi; Carly, Frédéric; Nicaud, Jean-Marc; Delvigne, Frank; Fickers, Patrick

2018-04-26

In the present study, we have isolated and characterized a Yarrowia lipolytica morphological mutant growing exclusively in the pseudohyphal morphology. The gene responsible for this phenotype, YALI0E06519g, was identified as homologous to the mitosis regulation gene HSL1 from Saccharomyces cerevisiae. Taking advantage of its morphology, we achieved the immobilization of the Δhsl1 mutant on the metallic structured packing of immobilized-cell bioreactors. We obtained significant cell retention and growth on the support during shake flask and bioreactor experiments without an attachment step prior to the culture. The system of medium aspersion on the packing ensured oxygen availability in the absence of agitation and minimized the potential release of cells in the culture medium. Additionally, the metallic packing proved its facility of cleaning and sterilization after fermentation. This combined use of morphological mutation and bioreactor design is a promising strategy to develop continuous processes for the production of recombinant protein and metabolites using Y. lipolytica. Graphical Abstract.

Engineering ..beta..-Oxidation in Yarrowia lipolytica for Methyl Ketone Production

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sanchez i Nogue, Violeta; Ramirez, Kelsey J; Singer, Christine

Medium- and long-chain methyl ketones are fatty acid-derived compounds that can be used as biofuel blending agents, flavors and fragrances. However, their large-scale production from sustainable feedstocks is currently limited due to the lack of robust microbial biocatalysts. The oleaginous yeast Yarrowia lipolytica is a promising biorefinery platform strain for the production of methyl ketones from renewable lignocellulosic biomass due to its natively high flux towards fatty acid biosynthesis. In this study, we report the metabolic engineering of Y. lipolytica to produce long- and very long-chain methyl ketones. Truncation of peroxisomal ..beta..-oxidation by chromosomal deletion of pot1 resulted in themore » biosynthesis of saturated, mono-, and diunsaturated methyl ketones in the C13-C23 range. Additional overexpression and peroxisomal targeting of a heterologous bacterial methyl ketone biosynthesis pathway yielded an initial titer of 151.5 mg/L of saturated methyl ketones. Dissolved oxygen concentrations in the cultures were found to substantially impact cell morphology and methyl ketone biosynthesis. Bioreactor cultivation under optimized conditions resulted in a titer of 314.8 mg/L of total methyl ketones, representing more than a 6000-fold increase over the parental strain. This work highlights the potential of Y. lipolytica to serve as chassis organism for the biosynthesis of acyl-thioester derived long- and very long-chain methyl ketones.« less

Production of Medium Chain Fatty Acids by Yarrowia lipolytica: Combining Molecular Design and TALEN to Engineer the Fatty Acid Synthase.

PubMed

Rigouin, Coraline; Gueroult, Marc; Croux, Christian; Dubois, Gwendoline; Borsenberger, Vinciane; Barbe, Sophie; Marty, Alain; Daboussi, Fayza; André, Isabelle; Bordes, Florence

2017-10-20

Yarrowia lipolytica is a promising organism for the production of lipids of biotechnological interest and particularly for biofuel. In this study, we engineered the key enzyme involved in lipid biosynthesis, the giant multifunctional fatty acid synthase (FAS), to shorten chain length of the synthesized fatty acids. Taking as starting point that the ketoacyl synthase (KS) domain of Yarrowia lipolytica FAS is directly involved in chain length specificity, we used molecular modeling to investigate molecular recognition of palmitic acid (C16 fatty acid) by the KS. This enabled to point out the key role of an isoleucine residue, I1220, from the fatty acid binding site, which could be targeted by mutagenesis. To address this challenge, TALEN (transcription activator-like effector nucleases)-based genome editing technology was applied for the first time to Yarrowia lipolytica and proved to be very efficient for inducing targeted genome modifications. Among the generated FAS mutants, those having a bulky aromatic amino acid residue in place of the native isoleucine at position 1220 led to a significant increase of myristic acid (C14) production compared to parental wild-type KS. Particularly, the best performing mutant, I1220W, accumulates C14 at a level of 11.6% total fatty acids. Overall, this work illustrates how a combination of molecular modeling and genome-editing technology can offer novel opportunities to rationally engineer complex systems for synthetic biology.

Impact of culture conditions on β-carotene encapsulation using Yarrowia lipolytica cells

NASA Astrophysics Data System (ADS)

Dang, Tran Hai; Minh, Ho Thi Thu; Van Nhi, Tran Nguyen; Ngoc, Ta Thi Minh

2017-09-01

Yeast cell was reported as an effective natural preformed material for use in encapsulation of hydrophobic compounds. The encapsulation process was normally considered as passive transfer through cellular wall and cellular membrane. Beside solubility of hydrophobic compound in phospholipid membrane or plasmolysis, membrane characteristics of yeast cell which are differed between strains and influenced by culture conditions are main factors involving the accumulation of hydrophobic compound into yeast cell. In this study, the oleaginous yeast Yarrowia lipolytica was used as micro-container shell to encapsulate a high hydrophobic compound - β-carotene. Yeast cell was cultured under different conditions and wet yeast biomass was incubated with β-carotene which was dissolved in soybean oil overnight. β-carotene accumulation was then extracted and evaluated by UV-VIS spectrometry. Optimization of culture condition was investigated using the Box-Behnken model. β-carotene encapsulation efficiency in Y. lipolytica was showed to be affected by both pH of medium and agitation conditions. The highest β-carotene encapsulation efficiency was optimized at 42.8 μg/g with Y. lipolytica cultured at pH 4.5, medium volume equal to 115 ml and agitation speed at 211 rpm.

Prospect for Developing a Consolidated Bioprocessing (CBP) Strain Using Xylan as the Substrate: the Case Study of Yarrowia lipolytica

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Wei; Wei, Hui; Alahuhta, Markus

2016-07-08

To achieve the goal of developing a direct microbial sugar conversion platform for the production of lipids and drop-in fuels from cellulosic biomass substrate, Yarrowia lipolytica was used to investigate its potential for being developed as CBP strain by expressing cellulase and xylanase enzymes. Y. lipolytica is known to accumulate lipids intracellularly and is capable of metabolizing glucose and xylose to produce lipids; however, due to the lack of the biomass degrading enzymes, it cannot directly utilize lignocellulosic substrates as carbon sources. While research is continuing on the development of a Y. lipolytica strain able to degrade cellulose, in thismore » study, we present successful expression of several xylanases in Y. lipolytica. To the best of our knowledge, this is the first study introducing heterologous hemicellulose genes into the genome of Y. lipolytica. SDS-PAGE and western blotting analysis showed that the endo-xylanase gene XynII and exo-xylosidase gene XlnD were successfully expressed and secreted, and the expressed xylanases were likely either not or sparsely glycosylated, which is advantageous for expression of heterologous proteins from any species. Enzymatic activity tests further demonstrated active expression of XynII and XlnD in Y. lipolytica. Furthermore, synergistic action on converting xylan to xylose was observed when XlnD worked in concert with XynII. XlnD was able to work on the xylo-oligomers generated by XynII, enhancing the xylan conversion to monomeric xylose. The successful expression of these xylanases in Yarrowia further advances us towards our goal to develop a direct microbial conversion process using this organism. and xylose to produce lipids; however, due to the lack of the biomass degrading enzymes, it cannot directly utilize lignocellulosic substrates as carbon sources. While research is continuing on the development of a Y. lipolytica strain able to degrade cellulose, in this study, we present successful

Engineering β-oxidation in Yarrowia lipolytica for methyl ketone production.

PubMed

Hanko, Erik K R; Denby, Charles M; Sànchez I Nogué, Violeta; Lin, Weiyin; Ramirez, Kelsey J; Singer, Christine A; Beckham, Gregg T; Keasling, Jay D

2018-05-28

Medium- and long-chain methyl ketones are fatty acid-derived compounds that can be used as biofuel blending agents, flavors and fragrances. However, their large-scale production from sustainable feedstocks is currently limited due to the lack of robust microbial biocatalysts. The oleaginous yeast Yarrowia lipolytica is a promising biorefinery platform strain for the production of methyl ketones from renewable lignocellulosic biomass due to its natively high flux towards fatty acid biosynthesis. In this study, we report the metabolic engineering of Y. lipolytica to produce long- and very long-chain methyl ketones. Truncation of peroxisomal β-oxidation by chromosomal deletion of pot1 resulted in the biosynthesis of saturated, mono-, and diunsaturated methyl ketones in the C 13 -C 23 range. Additional overexpression and peroxisomal targeting of a heterologous bacterial methyl ketone biosynthesis pathway yielded an initial titer of 151.5 mg/L of saturated methyl ketones. Dissolved oxygen concentrations in the cultures were found to substantially impact cell morphology and methyl ketone biosynthesis. Bioreactor cultivation under optimized conditions resulted in a titer of 314.8 mg/L of total methyl ketones, representing more than a 6000-fold increase over the parental strain. This work highlights the potential of Y. lipolytica to serve as chassis organism for the biosynthesis of acyl-thioester derived long- and very long-chain methyl ketones. Copyright © 2018 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

Yarrowia lipolytica: a model yeast for citric acid production.

PubMed

Cavallo, Ema; Charreau, Hernán; Cerrutti, Patricia; Foresti, María Laura

2017-12-01

Every year more than 2 million tons of citric acid (CA) are produced around the world for industrial uses. Although initially extracted from citrus, the low profitability of the process and the increasing demand soon stimulated the search for more efficient methods to produce CA. Currently, most world CA demand (99%) is satisfied by fermentations with microorganisms, especially filamentous fungi and yeasts. CA production with yeasts has certain advantages over molds (e.g. higher productivity and easier cultivation), which in the last two decades have triggered a clear increase in publications and patents devoted to the use of yeasts in this field. Yarrowia lipolytica has become a model yeast that proved to be successful in different production systems. Considering the current interest evidenced in the literature, the most significant information on CA production using Y. lipolytica is summarized. The relevance on CA yields of key factors such as strains, media formulation, environmental conditions and production regimes is thoroughly discussed, with particular focus on increasing CA productivity. Besides, the possibility of tuning the mentioned variables to reduce concomitant isocitric acid production-the biggest disadvantage of using yeasts-is analyzed. Available methods for CA purification/quantification are also discussed. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Engineering Yarrowia lipolytica for Use in Biotechnological Applications: A Review of Major Achievements and Recent Innovations.

PubMed

Madzak, Catherine

2018-06-25

Yarrowia lipolytica is an oleaginous saccharomycetous yeast with a long history of industrial use. It aroused interest several decades ago as host for heterologous protein production. Thanks to the development of numerous molecular and genetic tools, Y. lipolytica is now a recognized system for expressing heterologous genes and secreting the corresponding proteins of interest. As genomic and transcriptomic tools increased our basic knowledge on this yeast, we can now envision engineering its metabolic pathways for use as whole-cell factory in various bioconversion processes. Y. lipolytica is currently being developed as a workhorse for biotechnology, notably for single-cell oil production and upgrading of industrial wastes into valuable products. As it becomes more and more difficult to keep up with an ever-increasing literature on Y. lipolytica engineering technology, this article aims to provide basic and actualized knowledge on this research area. The most useful reviews on Y. lipolytica biology, use, and safety will be evoked, together with a resume of the engineering tools available in this yeast. This mini-review will then focus on recently developed tools and engineering strategies, with a particular emphasis on promoter tuning, metabolic pathways assembly, and genome editing technologies.

Development of a novel rDNA based plasmid for enhanced cell surface display on Yarrowia lipolytica.

PubMed

Bulani, Siyavuya Ishmael; Moleleki, Lucy; Albertyn, Jacobus; Moleleki, Ntsane

2012-05-20

In this study, a novel rDNA based plasmid was developed for display of heterologous proteins on the cell surface of Yarrowia lipolytica using the C-terminal end of the glycosylphosphatidylinositol (GPI) anchored Y. lipolytica cell wall protein 1 (YlCWP1). mCherry was used as a model protein to assess the efficiency of the constructed plasmid. Y. lipolytica transformants harbouring the expression cassettes showed a purple colour phenotype on selective YNB-casamino plates as compared to control cells indicating that mCherry was displayed on the cells. Expression of mCherry on cells of Y. lipolytica was confirmed by both fluorescent microscopy and flow cytometry. Furthermore, SDS-PAGE analysis and matrix-assisted laser desorption/ionization (MALDI)-time-of (TOF)-mass spectrometry (MS) peptide mass fingerprinting (PMF) confirmed that the protein cleaved from the yeast cells using enterokinase was mCherry. Efficient cleavage of mCherry reported in this work offers an alternative purification method for displayed heterologous proteins on Y. lipolytica cells using the plasmid constructed in this study. The developed displaying system offers great potential for industrial production and purification of heterologous proteins at low cost.

Comprehensive metabolomic, lipidomic and microscopic profiling of Yarrowia lipolytica during lipid accumulation identifies targets for increased lipogenesis

DOE PAGES

Pomraning, Kyle R.; Wei, Siwei; Karagiosis, Sue A.; ...

2015-04-23

Yarrowia lipolytica is an oleaginous ascomycete yeast that accumulates large amounts of lipids and has potential as a biofuel producing organism. Despite a growing scientific literature focused on lipid production by Y. lipolytica, there remain significant knowledge gaps regarding the key biological processes involved. We applied a combination of metabolomic and lipidomic profiling approaches as well as microscopic techniques to identify and characterize the key pathways involved in de novo lipid accumulation from glucose in batch cultured, wild-type Y. lipolytica. We found that lipids accumulated rapidly and peaked at 48 hours during the five day experiment, concurrent with a shiftmore » in amino acid metabolism. We also report that Y. lipolytica secretes disaccharides early in batch culture and reabsorbs them when extracellular glucose is depleted. Exhaustion of extracellular sugars coincided with thickening of the cell wall, suggesting that genes involved in cell wall biogenesis may be a useful target for improving the efficiency of lipid producing yeast strains.« less

Urea and urine are a viable and cost-effective nitrogen source for Yarrowia lipolytica biomass and lipid accumulation.

PubMed

Brabender, Matthew; Hussain, Murtaza Shabbir; Rodriguez, Gabriel; Blenner, Mark A

2018-03-01

Yarrowia lipolytica is an industrial yeast that has been used in the sustainable production of fatty acid-derived and lipid compounds due to its high growth capacity, genetic tractability, and oleaginous properties. This investigation examines the possibility of utilizing urea or urine as an alternative to ammonium sulfate as a nitrogen source to culture Y. lipolytica. The use of a stoichiometrically equivalent concentration of urea in lieu of ammonium sulfate significantly increased cell growth when glucose was used as the carbon source. Furthermore, Y. lipolytica growth was equally improved when grown with synthetic urine and real human urine. Equivalent or better lipid production was achieved when cells are grown on urea or urine. The successful use of urea and urine as nitrogen sources for Y. lipolytica growth highlights the potential of using cheaper media components as well as exploiting and recycling non-treated human waste streams for biotechnology processes.

Synthetic Biology Expands the Industrial Potential of Yarrowia lipolytica.

PubMed

Markham, Kelly A; Alper, Hal S

2018-06-04

The oleaginous yeast Yarrowia lipolytica is quickly emerging as the most popular non-conventional (i.e., non-model organism) yeast in the bioproduction field. With a high propensity for flux through tricarboxylic acid (TCA) cycle intermediates and biological precursors such as acetyl-CoA and malonyl-CoA, this host is especially well suited to meet our industrial chemical production needs. Recent progress in synthetic biology tool development has greatly enhanced our ability to rewire this organism, with advances in genetic component design, CRISPR technologies, and modular cloning strategies. In this review we investigate recent developments in metabolic engineering and describe how the new tools being developed help to realize the full industrial potential of this host. Finally, we conclude with our vision of the developments that will be necessary to enhance future engineering efforts. Copyright © 2018 Elsevier Ltd. All rights reserved.

Engineering Yarrowia lipolytica to express secretory invertase with strong FBA1IN promoter.

PubMed

Hong, Seung-Pyo; Seip, John; Walters-Pollak, Dana; Rupert, Ross; Jackson, Raymond; Xue, Zhixiong; Zhu, Quinn

2012-02-01

Oleaginous yeast Yarrowia lipolytica is an important host for the production of lipid-derived compounds or heterologous proteins. Selection of strong promoters and effective expression systems is critical for heterologous protein secretion. To search for a strong promoter in Y. lipolytica, activities of FBA1, TDH1 and GPM1 promoters were compared to that of TEF1 promoter by constructing GUS reporter fusions. The FBA1 promoter activity was 2.2 and 5.5 times stronger than the TDH1 and GPM1 promoters, respectively. The FBA1IN promoter (FBA1 sequence of -826 to +169) containing an intron (+64 to +165) showed five-fold higher expression than the FBA1 promoter (-831 to -1). The transcriptional enhancement by the 5'-region within the FBA1 gene was confirmed by GPM1::FBA1 chimeric promoter construction. Using the strong FBA1IN promoter, four different S. cerevisiae SUC2 expression cassettes were tested for the SUC+ phenotype in Y. lipolytica. Functional invertase secretion was facilitated by the Xpr2 prepro-region with an additional 13 amino acids of mature Xpr2, or by the native Suc2 signal sequence. However, these two secretory signals in tandem, or the mature Suc2 with no secretory signal, did not direct secretion of functional invertase. Unlike previously reported Y. lipolytica SUC+ strains, our engineered stains secreted most of invertase into the medium. Copyright © 2011 John Wiley & Sons, Ltd.

Selection of Yarrowia lipolytica strains with high protein content from yeasts isolated from different marine environments

NASA Astrophysics Data System (ADS)

Chi, Zhenming; Wang, Fang; Wang, Lin; Li, Jing; Wang, Xianghong

2007-10-01

A total of 78 Yarrowia lipolytica yeast strains from seawater, sediments, mud of salterns, the guts of marine fish, and marine algae were obtained. After the crude protein of the yeasts was estimated by the method of Kjehldahl, we found that seven strains of the marine yeasts grown in soy bean cake hydrolysate with 20 g L-1 of glucose for 48 h at 28°C contained more than 41.0 g protein per 100 g of cell dry weight and the cell dry weight was more than 4.4 g per L of the culture. Among them, strain SWJ-1b contained the highest crude protein. The results of Biolog identification and molecular methods further confirmed that they indeed belonged to Y. lipolytica.

[Genetic system for maintaining the mitochondrial human genome in yeast Yarrowia lipolytica].

PubMed

Isakova, E P; Deryabina, Yu I; Velyakova, A V; Biryukova, J K; Teplova, V V; Shevelev, A B

2016-01-01

For the first time, the possibility of maintaining an intact human mitochondrial genome in a heterologous system in the mitochondria of yeast Yarrowia lipolytica is shown. A method for introducing directional changes into the structure of the mitochondrial human genome replicating in Y. lipolytica by an artificially induced ability of yeast mitochondria for homologous recombination is proposed. A method of introducing and using phenotypic selection markers for the presence or absence of defects in genes tRNA-Lys and tRNA-Leu of the mitochondrial genome is developed. The proposed system can be used to correct harmful mutations of the human mitochondrial genome associated with mitochondrial diseases and for preparative amplification of intact mitochondrial DNA with an adjusted sequence in yeast cells. The applicability of the new system for the correction of mutations in the genes of Lys- and Leu-specific tRNAs of the human mitochondrial genome associated with serious and widespread human mitochondrial diseases such as myoclonic epilepsy with lactic acidosis (MELAS) and myoclonic epilepsy with ragged-red fibers (MERRF) is shown.

CRISPR-Cas9-Mediated Genome Editing and Transcriptional Control in Yarrowia lipolytica.

PubMed

Schwartz, Cory; Wheeldon, Ian

2018-01-01

The discovery and adaptation of RNA-guided nucleases has resulted in the rapid development of efficient, scalable, and easily accessible synthetic biology tools for targeted genome editing and transcriptional control. In these systems, for example CRISPR-Cas9 from Streptococcus pyogenes, a protein with nuclease activity is targeted to a specific nucleotide sequence by a short RNA molecule, whereupon binding it cleaves the targeted nucleotide strand. To extend this genome-editing ability to the industrially important oleaginous yeast Yarrowia lipolytica, we developed a set of easily usable and effective CRISPR-Cas9 episomal vectors. In this protocols chapter, we first present a method by which arbitrary protein-coding genes can be disrupted via indel formation after CRISPR-Cas9 targeting. A second method demonstrates how the same CRISPR-Cas9 system can be used to induce markerless gene cassette integration into the genome by inducing homologous recombination after DNA cleavage by Cas9. Finally, we describe how a catalytically inactive form of Cas9 fused to a transcriptional repressor can be used to control transcription of native genes in Y. lipolytica. The CRISPR-Cas9 tools and strategies described here greatly increase the types of genome editing and transcriptional control that can be achieved in Y. lipolytica, and promise to facilitate more advanced engineering of this important oleaginous host.

Irradiation of Yarrowia lipolytica NRRL YB-567 creating novel strains with enhanced ammonia and oil production on protein and carbohydrate substrates

USDA-ARS?s Scientific Manuscript database

Increased interest in sustainable production of renewable diesel and other valuable bioproducts is redoubling efforts to improve economic feasibility of microbial-based oil production. The yeast Yarrowia lipolytica is capable of employing a wide variety of substrates to produce oil and valuable co-p...

An ortholog of farA of Aspergillus nidulans is implicated in the transcriptional activation of genes involved in fatty acid utilization in the yeast Yarrowia lipolytica

DOE Office of Scientific and Technical Information (OSTI.GOV)

Poopanitpan, Napapol; Kobayashi, Satoshi; Fukuda, Ryouichi

2010-11-26

Research highlights: {yields} POR1 is a Yarrowia lipolytica ortholog of farA involved in fatty acid response in A. nidulans. {yields} Deletion of POR1 caused growth defects on fatty acids. {yields} {Delta}por1 strain exhibited defects in the induction of genes involved in fatty acid utilization. -- Abstract: The yeast Yarrowia lipolytica effectively utilizes hydrophobic substrates such as fatty acids and n-alkanes. To identify a gene(s) regulating fatty acid utilization in Y. lipolytica, we first studied homologous genes to OAF1 and PIP2 of Saccharomyces cerevisiae, but their disruption did not change growth on oleic acid at all. We next characterized a Y.more » lipolytica gene, POR1 (primary oleate regulator 1), an ortholog of farA encoding a transcriptional activator that regulates fatty acid utilization in Aspergillus nidulans. The deletion mutant of POR1 was defective in the growth on various fatty acids, but not on glucose, glycerol, or n-hexadecane. It exhibited slight defect on n-decane. The transcriptional induction of genes involved in {beta}-oxidation and peroxisome proliferation by oleate was distinctly diminished in the {Delta}por1 strains. These data suggest that POR1 encodes a transcriptional activator widely regulating fatty acid metabolism in Y. lipolytica.« less

New finding and optimal production of a novel extracellular alkaline lipase from Yarrowia lipolytica NRRL Y-2178.

PubMed

Lee, Geon-Ho; Bae, Jae-Han; Suh, Min-Jung; Kim, In-Hwan; Hou, Ching T; Kim, Hak-Ryul

2007-06-01

Lipases are industrially useful versatile enzymes that catalyze numerous different reactions including hydrolysis of triglycerides, transesterification, and chiral synthesis of esters under natural conditions. Although lipases from various sources have been widely used in industrial applications, such as in food, chemical, pharmaceutical, and detergent industries, there are still substantial current interests in developing new microbial lipases, specifically those functioning in abnormal conditions. We screened 17 lipase-producing yeast strains, which were prescreened for substrate specificity of lipase from more than 500 yeast strains from the Agricultural Research Service Culture Collection (Peoria, IL, U.S.A.), and selected Yarrowia lipolytica NRRL Y-2178 as a best lipase producer. This report presents new finding and optimal production of a novel extracellular alkaline lipase from Y. lipolytica NRRL Y-2178. Optimal c ulture conditions f orlipase production by Y. lipolytica NRRL Y-2178 were 72 h incubation time, 27.5 degrees C, pH 9.0. Glycerol and glucose were efficiently used as the most efficient carbon sources, and a combination of yeast extract and peptone was a good nitrogen source for lipase production by Y. lipolytica NRRL Y-2178. These results suggested that Y. lipolytica NRRL Y-2178 showsgood industrial potential as a new alkaline lipase producer.

Analysis of ATP-citrate lyase and malic enzyme mutants of Yarrowia lipolytica points out the importance of mannitol metabolism in fatty acid synthesis.

PubMed

Dulermo, Thierry; Lazar, Zbigniew; Dulermo, Rémi; Rakicka, Magdalena; Haddouche, Ramedane; Nicaud, Jean-Marc

2015-09-01

The role of the two key enzymes of fatty acid (FA) synthesis, ATP-citrate lyase (Acl) and malic enzyme (Mae), was analyzed in the oleaginous yeast Yarrowia lipolytica. In most oleaginous yeasts, Acl and Mae are proposed to provide, respectively, acetyl-CoA and NADPH for FA synthesis. Acl was mainly studied at the biochemical level but no strain depleted for this enzyme was analyzed in oleaginous microorganisms. On the other hand the role of Mae in FA synthesis in Y. lipolytica remains unclear since it was proposed to be a mitochondrial NAD(H)-dependent enzyme and not a cytosolic NADP(H)-dependent enzyme. In this study, we analyzed for the first time strains inactivated for corresponding genes. Inactivation of ACL1 decreases FA synthesis by 60 to 80%, confirming its essential role in FA synthesis in Y. lipolytica. Conversely, inactivation of MAE1 has no effects on FA synthesis, except in a FA overaccumulating strain where it improves FA synthesis by 35%. This result definitively excludes Mae as a major key enzyme for FA synthesis in Y. lipolytica. During the analysis of both mutants, we observed a negative correlation between FA and mannitol level. As mannitol and FA pathways may compete for carbon storage, we inactivated YlSDR, encoding a mannitol dehydrogenase converting fructose and NADPH into mannitol and NADP+. The FA content of the resulting mutant was improved by 60% during growth on fructose, demonstrating that mannitol metabolism may modulate FA synthesis in Y. lipolytica. Copyright © 2015. Published by Elsevier B.V.

[The Engineering of a Yarrowia lipolytica Yeast Strain Capable of Homologous Recombination of the Mitochondrial Genome].

PubMed

Isakova, E P; Epova, E Yu; Sekova, V Yu; Trubnikova, E V; Kudykina, Yu K; Zylkova, M V; Guseva, M A; Deryabina, Yu I

2015-01-01

None of the studied eukaryotic species has a natural system for homologous recombination of the mitochondrial genome. We propose an integrated genetic construct pQ-SRUS, which allows introduction of the recA gene from Bacillus subtilis into the nuclear genome of an extremophilic yeast, Yarrowia lipolytica. The targeting of recombinant RecA to the yeast mitochondria is provided by leader sequences (5'-UTR and 3'-UTR) derived from the SOD2 gene mRNA, which exhibits affinity to the outer mitochondrial membrane and thus provides cotranslational transport of RecA to the inner space of the mitochondria. The Y. lipolytica strain bearing the pQ-SRUS construct has the unique ability to integrate DNA constructs into the mitochondrial genome. This fact was confirmed using a tester construct, pQ-NIHN, intended for the introduction of the EYFP gene into the translation initiation region of the Y. lipolytica ND1 mitochondrial gene. The Y. lipolytica strain bearing pQ-SRUS makes it possible to engineer recombinant producers based on Y. lipolytica bearing transgenes in the mitochondrial genome. They are promising for the construction of a genetic system for in vivo replication and modification of the human mitochondrial genome. These strains may be used as a tool for the treatment of human mitochondrial diseases (including genetically inherited ones).

Robust signal peptides for protein secretion in Yarrowia lipolytica: identification and characterization of novel secretory tags.

PubMed

Celińska, Ewelina; Borkowska, Monika; Białas, Wojciech; Korpys, Paulina; Nicaud, Jean-Marc

2018-06-01

Upon expression of a given protein in an expression host, its secretion into the culture medium or cell-surface display is frequently advantageous in both research and industrial contexts. Hence, engineering strategies targeting folding, trafficking, and secretion of the proteins gain considerable interest. Yarrowia lipolytica has emerged as an efficient protein expression platform, repeatedly proved to be a competitive secretor of proteins. Although the key role of signal peptides (SPs) in secretory overexpression of proteins and their direct effect on the final protein titers are widely known, the number of reports on manipulation with SPs in Y. lipolytica is rather scattered. In this study, we assessed the potential of ten different SPs for secretion of two heterologous proteins in Y. lipolytica. Genomic and transcriptomic data mining allowed us to select five novel, previously undescribed SPs for recombinant protein secretion in Y. lipolytica. Their secretory potential was assessed in comparison with known, widely exploited SPs. We took advantage of Golden Gate approach, for construction of expression cassettes, and micro-volume enzymatic assays, for functional screening of large libraries of recombinant strains. Based on the adopted strategy, we identified novel secretory tags, characterized their secretory capacity, indicated the most potent SPs, and suggested a consensus sequence of a potentially robust synthetic SP to expand the molecular toolbox for engineering Y. lipolytica.

Bioconversion of R-(+)-limonene to perillic acid by the yeast Yarrowia lipolytica

PubMed Central

Ferrara, Maria Antonieta; Almeida, Débora S.; Siani, Antonio C.; Lucchetti, Leonardo; Lacerda, Paulo S.B.; Freitas, André; Tappin, Marcelo R.R.; Bon, Elba P.S.

2013-01-01

Perillyl derivatives are increasingly important due to their flavouring and antimicrobial properties as well as their potential as anticancer agents. These terpenoid species, which are present in limited amounts in plants, may be obtained via bioconversion of selected monoterpene hydrocarbons. In this study, seventeen yeast strains were screened for their ability to oxidize the exocyclic methyl group in the p-menthene moiety of limonene into perillic acid. Of the yeast tested, the highest efficiency was observed for Yarrowia lipolytica ATCC 18942. The conversion of R (+)-limonene by Y. lipolytica was evaluated by varying the pH (3 to 8) and the temperature (25 to 30 °C) in a reaction medium containing 0.5% v/v limonene and 10 g/L of stationary phase cells (dry weight). The best results, corresponding to 564 mg/L of perillic acid, were obtained in buffered medium at pH 7.1 that was incubated at 25 °C for 48 h. The stepwise addition of limonene increased the perillic acid concentration by over 50%, reaching 855 mg/L, whereas the addition of glucose or surfactant to the reaction medium did not improve the bioconversion process. The use of Y. lipolytica showed promise for ease of further downstream processing, as perillic acid was the sole oxidised product of the bioconversion reaction. Moreover, bioprocesses using safe and easy to cultivate yeast cells have been favoured in industry. PMID:24688495

Engineering Yarrowia lipolytica as a platform for synthesis of drop-in transportation fuels and oleochemicals

PubMed Central

Xu, Peng; Qiao, Kangjian; Ahn, Woo Suk; Stephanopoulos, Gregory

2016-01-01

Harnessing lipogenic pathways and rewiring acyl-CoA and acyl-ACP (acyl carrier protein) metabolism in Yarrowia lipolytica hold great potential for cost-efficient production of diesel, gasoline-like fuels, and oleochemicals. Here we assessed various pathway engineering strategies in Y. lipolytica toward developing a yeast biorefinery platform for sustainable production of fuel-like molecules and oleochemicals. Specifically, acyl-CoA/acyl-ACP processing enzymes were targeted to the cytoplasm, peroxisome, or endoplasmic reticulum to generate fatty acid ethyl esters and fatty alkanes with tailored chain length. Activation of endogenous free fatty acids and the subsequent reduction of fatty acyl-CoAs enabled the efficient synthesis of fatty alcohols. Engineering a hybrid fatty acid synthase shifted the free fatty acids to a medium chain-length scale. Manipulation of alternative cytosolic acetyl-CoA pathways partially decoupled lipogenesis from nitrogen starvation and unleashed the lipogenic potential of Y. lipolytica. Taken together, the strategies reported here represent promising steps to develop a yeast biorefinery platform that potentially upgrades low-value carbons to high-value fuels and oleochemicals in a sustainable and environmentally friendly manner. PMID:27621436

Yarrowia lipolytica possesses two plasma membrane alkali metal cation/H+ antiporters with different functions in cell physiology.

PubMed

Papouskova, Klara; Sychrova, Hana

2006-04-03

The family of Nha antiporters mediating the efflux of alkali metal cations in exchange for protons across the plasma membrane is conserved in all yeast species. Yarrowia lipolytica is a dimorphic yeast, phylogenetically very distant from the model yeast Saccharomyces cerevisiae. A search in its sequenced genome revealed two genes (designated as YlNHA1 and YlNHA2) with homology to the S. cerevisiae NHA1 gene, which encodes a plasma membrane alkali metal cation/H+ antiporter. Upon heterologous expression of both YlNHA genes in S. cerevisiae, we showed that Y. lipolytica antiporters differ not only in length and sequence, but also in their affinity for individual substrates. While the YlNha1 protein mainly increased cell tolerance to potassium, YlNha2p displayed a remarkable transport capacity for sodium. Thus, Y. lipolytica is the first example of a yeast species with two plasma membrane alkali metal cation/H+ antiporters differing in their putative functions in cell physiology; cell detoxification vs. the maintenance of stable intracellular pH, potassium content and cell volume.

Steroid biotransformations in biphasic systems with Yarrowia lipolytica expressing human liver cytochrome P450 genes

PubMed Central

2012-01-01

Background Yarrowia lipolytica efficiently metabolizes and assimilates hydrophobic compounds such as n-alkanes and fatty acids. Efficient substrate uptake is enabled by naturally secreted emulsifiers and a modified cell surface hydrophobicity and protrusions formed by this yeast. We were examining the potential of recombinant Y. lipolytica as a biocatalyst for the oxidation of hardly soluble hydrophobic steroids. Furthermore, two-liquid biphasic culture systems were evaluated to increase substrate availability. While cells, together with water soluble nutrients, are maintained in the aqueous phase, substrates and most of the products are contained in a second water-immiscible organic solvent phase. Results For the first time we have co-expressed the human cytochromes P450 2D6 and 3A4 genes in Y. lipolytica together with human cytochrome P450 reductase (hCPR) or Y. lipolytica cytochrome P450 reductase (YlCPR). These whole-cell biocatalysts were used for the conversion of poorly soluble steroids in biphasic systems. Employing a biphasic system with the organic solvent and Y. lipolytica carbon source ethyl oleate for the whole-cell bioconversion of progesterone, the initial specific hydroxylation rate in a 1.5 L stirred tank bioreactor was further increased 2-fold. Furthermore, the product formation was significantly prolonged as compared to the aqueous system. Co-expression of the human CPR gene led to a 4-10-fold higher specific activity, compared to the co-overexpression of the native Y. lipolytica CPR gene. Multicopy transformants showed a 50-70-fold increase of activity as compared to single copy strains. Conclusions Alkane-assimilating yeast Y. lipolytica, coupled with the described expression strategies, demonstrated its high potential for biotransformations of hydrophobic substrates in two-liquid biphasic systems. Especially organic solvents which can be efficiently taken up and/or metabolized by the cell might enable more efficient bioconversion as compared

Enhanced α-ketoglutaric acid production and recovery in Yarrowia lipolytica yeast by effective pH controlling.

PubMed

Morgunov, Igor G; Kamzolova, Svetlana V; Samoilenko, Vladimir A

2013-10-01

The replacement of chemical synthesis by environmentally friendly energy-efficient technologies for production of valuable metabolites is a principal strategy of developing biotechnological industry all over the world. In the present study, we develop a method for α-ketoglutaric acid (KGA) production from rapeseed oil with the use of Yarrowia lipolytica yeast. Sixty strains of Y. lipolytica yeasts were tested for their ability to produce KGA, and the strain Y. lipolytica 212 (Y. lipolytica VKM Y-2412) was selected as a promising KGA producer. Using a three-stage pH controlling, in which pH was 4.5 in the growth phase, then since 72 to 144 h, pH was maintained at 3.5 and in the later phase of acid production, the titration by KOH was switch off, selected strain produced 106.5 g l(-1) of KGA with mass yield of 0.95 g g(-1). KGA in the form of monopotassium salt was isolated from the culture broth and purified. The isolation procedure involved separation of biomass, extraction of residual triglycerides, filtrate bleaching, and acidification with mineral acid (to pH 2.8-3.4), concentration, precipitation of mineral salts, and crystallization of the product. The purity of KGA isolated from the culture filtrate reached 99.1 %.

Development of recombinant Yarrowia lipolytica producing virus-like particles of a fish nervous necrosis virus.

PubMed

Luu, Van-Trinh; Moon, Hye Yun; Hwang, Jee Youn; Kang, Bo-Kyu; Kang, Hyun Ah

2017-08-01

Nervous necrosis virus (NNV) causes viral encephalopathy and retinopathy, a devastating disease of many species of cultured marine fish worldwide. In this study, we used the dimorphic non-pathogenic yeast Yarrowia lipolytica as a host to express the capsid protein of red-spotted grouper nervous necrosis virus (RGNNV-CP) and evaluated its potential as a platform for vaccine production. An initial attempt was made to express the codon-optimized synthetic genes encoding intact and N-terminal truncated forms of RGNNV-CP under the strong constitutive TEF1 promoter using autonomously replicating sequence (ARS)-based vectors. The full-length recombinant capsid proteins expressed in Y. lipolytica were detected not only as monomers and but also as trimers, which is a basic unit for formation of NNV virus-like particles (VLPs). Oral immunization of mice with whole recombinant Y. lipolytica harboring the ARS-based plasmids was shown to efficiently induce the formation of IgG against RGNNV-CP. To increase the number of integrated copies of the RGNNV-CP expression cassette, a set of 26S ribosomal DNA-based multiple integrative vectors was constructed in combination with a series of defective Ylura3 with truncated promoters as selection markers, resulting in integrants harboring up to eight copies of the RGNNV-CP cassette. Sucrose gradient centrifugation and transmission electron microscopy of this high-copy integrant were carried out to confirm the expression of RGNNV-CPs as VLPs. This is the first report on efficient expression of viral capsid proteins as VLPs in Y. lipolytica, demonstrating high potential for the Y. lipolytica expression system as a platform for recombinant vaccine production based on VLPs.

Investigating Proteome and Transcriptome Defense Response of Apples Induced by Yarrowia lipolytica.

PubMed

Zhang, Hongyin; Chen, Liangliang; Sun, Yiwen; Zhao, Lina; Zheng, Xiangfeng; Yang, Qiya; Zhang, Xiaoyun

2017-04-01

A better understanding of the mode of action of postharvest biocontrol agents on fruit surfaces is critical for the advancement of successful implementation of postharvest biocontrol products. This is due to the increasing importance of biological control of postharvest diseases over chemical and other control methods. However, most of the mechanisms involved in biological control remain unknown and need to be explored. Yarrowia lipolytica significantly inhibited blue mold decay of apples caused by Penicillium expansum. The findings also demonstrated that Y. lipolytica stimulated the activities of polyphenoloxidase, peroxidase, chitinase, l-phenylalanine ammonia lyase involved in enhancing defense responses in apple fruit tissue. Proteomic and transcriptomic analysis revealed a total of 35 proteins identified as up- and down-regulated in response to the Y. lipolytica inducement. These proteins were related to defense, biotic stimulus, and stress responses, such as pathogenesis-related proteins and dehydrin. The analysis of the transcriptome results proved that the induced resistance was mediated by a crosstalk between salicylic acid (SA) and ethylene/jasmonate (ET/JA) pathways. Y. lipolytica treatment activated the expression of isochorismate synthase gene in the SA pathway, which up-regulates the expression of PR4 in apple. The expression of 1-aminocyclopropane-1-carboxylate oxidase gene and ET-responsive transcription factors 2 and 4, which are involved in the ET pathway, were also activated. In addition, cytochrome oxidase I, which plays an important role in JA signaling for resistance acquisition, was also activated. However, not all of the genes had a positive effect on the SA and ET/JA signal pathways. As transcriptional repressors in JA signaling, TIFY3B and TIFY11B were triggered by the yeast, but the gene expression levels were relatively low. Taken together, Y. lipolytica induced the SA and ET/JA signal mediating the defense pathways by stimulating

Optimized invertase expression and secretion cassette for improving Yarrowia lipolytica growth on sucrose for industrial applications.

PubMed

Lazar, Zbigniew; Rossignol, Tristan; Verbeke, Jonathan; Crutz-Le Coq, Anne-Marie; Nicaud, Jean-Marc; Robak, Małgorzata

2013-11-01

Yarrowia lipolytica requires the expression of a heterologous invertase to grow on a sucrose-based substrate. This work reports the construction of an optimized invertase expression cassette composed of Saccharomyces cerevisiae Suc2p secretion signal sequence followed by the SUC2 sequence and under the control of the strong Y. lipolytica pTEF promoter. This new construction allows a fast and optimal cleavage of sucrose into glucose and fructose and allows cells to reach the maximum growth rate. Contrary to pre-existing constructions, the expression of SUC2 is not sensitive to medium composition in this context. The strain JMY2593, expressing this new cassette with an optimized secretion signal sequence and a strong promoter, produces 4,519 U/l of extracellular invertase in bioreactor experiments compared to 597 U/l in a strain expressing the former invertase construction. The expression of this cassette strongly improved production of invertase and is suitable for simultaneously high production level of citric acid from sucrose-based media.

A Rac Homolog Is Required for Induction of Hyphal Growth in the Dimorphic Yeast Yarrowia lipolytica

PubMed Central

Hurtado, Cleofe A. R.; Beckerich, Jean-Marie; Gaillardin, Claude; Rachubinski, Richard A.

2000-01-01

Dimorphism in fungi is believed to constitute a mechanism of response to adverse conditions and represents an important attribute for the development of virulence by a number of pathogenic fungal species. We have isolated YlRAC1, a gene encoding a 192-amino-acid protein that is essential for hyphal growth in the dimorphic yeast Yarrowia lipolytica and which represents the first Rac homolog described for fungi. YlRAC1 is not an essential gene, and its deletion does not affect the ability to mate or impair actin polarization in Y. lipolytica. However, strains lacking functional YlRAC1 show alterations in cell morphology, suggesting that the function of YlRAC1 may be related to some aspect of the polarization of cell growth. Northern blot analysis showed that transcription of YlRAC1 increases steadily during the yeast-to-hypha transition, while Southern blot analysis of genomic DNA suggested the presence of several RAC family members in Y. lipolytica. Interestingly, strains lacking functional YlRAC1 are still able to grow as the pseudohyphal form and to invade agar, thus pointing to a function for YlRAC1 downstream of MHY1, a previously isolated gene encoding a C2H2-type zinc finger protein with the ability to bind putative stress response elements and whose activity is essential for both hyphal and pseudohyphal growth in Y. lipolytica. PMID:10762235

Unraveling fatty acid transport and activation mechanisms in Yarrowia lipolytica.

PubMed

Dulermo, Rémi; Gamboa-Meléndez, Heber; Ledesma-Amaro, Rodrigo; Thévenieau, France; Nicaud, Jean-Marc

2015-09-01

Fatty acid (FA) transport and activation have been extensively studied in the model yeast species Saccharomyces cerevisiae but have rarely been examined in oleaginous yeasts, such as Yarrowia lipolytica. Because the latter begins to be used in biodiesel production, understanding its FA transport and activation mechanisms is essential. We found that Y. lipolytica has FA transport and activation proteins similar to those of S. cerevisiae (Faa1p, Pxa1p, Pxa2p, Ant1p) but mechanism of FA peroxisomal transport and activation differs greatly with that of S. cerevisiae. While the ScPxa1p/ScPxa2p heterodimer is essential for growth on long-chain FAs, ΔYlpxa1 ΔYlpxa2 is not impaired for growth on FAs. Meanwhile, ScAnt1p and YlAnt1p are both essential for yeast growth on medium-chain FAs, suggesting they function similarly. Interestingly, we found that the ΔYlpxa1 ΔYlpxa2 ΔYlant1 mutant was unable to grow on short-, medium-, or long-chain FAs, suggesting that YlPxa1p, YlPxa2p, and YlAnt1p belong to two different FA degradation pathways. We also found that YlFaa1p is involved in FA storage in lipid bodies and that FA remobilization largely depended on YlFat1p, YlPxa1p and YlPxa2p. This study is the first to comprehensively examine FA intracellular transport and activation in oleaginous yeast. Copyright © 2015. Published by Elsevier B.V.

Biomass production by novel strains of Yarrowia lipolytica using raw glycerol, derived from biodiesel production.

PubMed

Juszczyk, Piotr; Tomaszewska, Ludwika; Kita, Agnieszka; Rymowicz, Waldemar

2013-06-01

This study demonstrated the potential applicability of the isolated strains of Yarrowia lipolytica for the valorization of glycerol waste generated during biodiesel production, throughout biomass production. Twenty-one strains were isolated from different environments and identified as Y. lipolytica. Biomass production from pure glycerol (25 g L(-1)) was performed in the shake-flasks experiment. Eight strains with the best biomass production ability were chosen for studies in bioreactor (pH 3.5). The analysis of technological process parameters and biomass chemical composition demonstrated that S6 strain was the most suitable for biomass production. Its application allowed obtaining 11.7 and 12.3 g L(-1) of the biomass with 1.30 and 1.37 g L(-1) h(-1) productivity, respectively when pure and raw glycerol (25 g L(-1)) was used. In the yeast protein amino acid profile the contents of lysine, threonine and phenylalanine/tyrosine were higher than required by FAO/WHO. According to the EAAI, the nutritional value of the biomass reached up to 72.3%. Copyright © 2013 Elsevier Ltd. All rights reserved.

Sugar versus fat: elimination of glycogen storage improves lipid accumulation in Yarrowia lipolytica.

PubMed

Bhutada, Govindprasad; Kavšcek, Martin; Ledesma-Amaro, Rodrigo; Thomas, Stéphane; Rechberger, Gerald N; Nicaud, Jean-Marc; Natter, Klaus

2017-05-01

Triacylglycerol (TAG) and glycogen are the two major metabolites for carbon storage in most eukaryotic organisms. We investigated the glycogen metabolism of the oleaginous Yarrowia lipolytica and found that this yeast accumulates up to 16% glycogen in its biomass. Assuming that elimination of glycogen synthesis would result in an improvement of lipid accumulation, we characterized and deleted the single gene coding for glycogen synthase, YlGSY1. The mutant was grown under lipogenic conditions with glucose and glycerol as substrates and we obtained up to 60% improvement in TAG accumulation compared to the wild-type strain. Additionally, YlGSY1 was deleted in a background that was already engineered for high lipid accumulation. In this obese background, TAG accumulation was also further increased. The highest lipid content of 52% was found after 3 days of cultivation in nitrogen-limited glycerol medium. Furthermore, we constructed mutants of Y. lipolytica and Saccharomyces cerevisiae that are deleted for both glycogen and TAG synthesis, demonstrating that the ability to store carbon is not essential. Overall, this work showed that glycogen synthesis is a competing pathway for TAG accumulation in oleaginous yeasts and that deletion of the glycogen synthase has beneficial effects on neutral lipid storage. © FEMS 2017.

Functional analysis of recombinant human and Yarrowia lipolytica O-GlcNAc transferases expressed in Saccharomyces cerevisiae.

PubMed

Oh, Hye Ji; Moon, Hye Yun; Cheon, Seon Ah; Hahn, Yoonsoo; Kang, Hyun Ah

2016-10-01

O-linked β-N-acetylglucosamine (O-GlcNAc) glycosylation is an important post-translational modification in many cellular processes. It is mediated by O-GlcNAc transferases (OGTs), which catalyze the addition of O-GlcNAc to serine or threonine residues of the target proteins. In this study, we expressed a putative Yarrowia lipolytica OGT (YlOGT), the only homolog identified in the subphylum Saccharomycotina through bioinformatics analysis, and the human OGT (hOGT) as recombinant proteins in Saccharomyces cerevisiae, and performed their functional characterization. Immunoblotting assays using antibody against O-GlcNAc revealed that recombinant hOGT (rhOGT), but not the recombinant YlOGT (rYlOGT), undergoes auto-O-GlcNAcylation in the heterologous host S. cerevisiae. Moreover, the rhOGT expressed in S. cerevisiae showed a catalytic activity during in vitro assays using casein kinase II substrates, whereas no such activity was obtained in rYlOGT. However, the chimeric human-Y. lipolytica OGT, carrying the human tetratricopeptide repeat (TPR) domain along with the Y. lipolytica catalytic domain (CTD), mediated the transfer of O-GlcNAc moiety during the in vitro assays. Although the overexpression of full-length OGTs inhibited the growth of S. cerevisiae, no such inhibition was obtained upon overexpression of only the CTD fragment, indicating the role of TPR domain in growth inhibition. This is the first report on the functional analysis of the fungal OGT, indicating that the Y. lipolytica OGT retains its catalytic activity, although the physiological role and substrates of YlOGT remain to be elucidated.

Multi-omics analysis reveals regulators of the response to nitrogen limitation in Yarrowia lipolytica.

PubMed

Pomraning, Kyle R; Kim, Young-Mo; Nicora, Carrie D; Chu, Rosalie K; Bredeweg, Erin L; Purvine, Samuel O; Hu, Dehong; Metz, Thomas O; Baker, Scott E

2016-02-25

Yarrowia lipolytica is an oleaginous ascomycete yeast that stores lipids in response to limitation of nitrogen. While the enzymatic pathways responsible for neutral lipid accumulation in Y. lipolytica are well characterized, regulation of these pathways has received little attention. We therefore sought to characterize the response to nitrogen limitation at system-wide levels, including the proteome, phosphoproteome and metabolome, to better understand how this organism regulates and controls lipid metabolism and to identify targets that may be manipulated to improve lipid yield. We found that ribosome structural genes are down-regulated under nitrogen limitation, during which nitrogen containing compounds (alanine, putrescine, spermidine and urea) are depleted and sugar alcohols and TCA cycle intermediates accumulate (citrate, fumarate and malate). We identified 1219 novel phosphorylation sites in Y. lipolytica, 133 of which change in their abundance during nitrogen limitation. Regulatory proteins, including kinases and DNA binding proteins, are particularly enriched for phosphorylation. Within lipid synthesis pathways, we found that ATP-citrate lyase, acetyl-CoA carboxylase and lecithin cholesterol acyl transferase are phosphorylated during nitrogen limitation while many of the proteins involved in β-oxidation are down-regulated, suggesting that storage lipid accumulation may be regulated by phosphorylation of key enzymes. Further, we identified short DNA elements that associate specific transcription factor families with up- and down-regulated genes. Integration of metabolome, proteome and phosphoproteome data identifies lipid accumulation in response to nitrogen limitation as a two-fold result of increased production of acetyl-CoA from excess citrate and decreased capacity for β-oxidation.

Study of trans-trans farnesol effect on hyphae formation by Yarrowia lipolytica.

PubMed

Nunes, Patrícia Martins Botelho; da Rocha, Silvia Maria; Amaral, Priscilla Filomena Fonseca; da Rocha-Leão, Maria Helena Miguez

2013-12-01

Dimorphism is an ability of certain fungi related to its adaptation to the environment and provides a selective advantage under stress conditions and is associated to the development of human diseases. Hyphae inducing- and inhibitory-effect of farnesol on hyphae formation by the dimorphic yeast Yarrowia lipolytica was evaluated through digital image analysis. The agitation speed of the culture was the most effective hyphae inducer in comparison to bovine calf serum and N-acetylglucosamine. In low agitation system, bovine calf serum was more effective for hyphae formation inducing 57 % of hyphae transition. Farnesol inhibited hyphae formation even in low concentration (300 μM) and this effect increased with increasing concentrations. In the presence of N-acetylglucosamine, this effect was more evident in comparison to the presence of bovine calf serum, which might have protected the cells from farnesol. Digital image analysis was an important tool to evaluate this phenomenon.

New Insights into Sulfur Metabolism in Yeasts as Revealed by Studies of Yarrowia lipolytica

PubMed Central

Hébert, Agnès; Forquin-Gomez, Marie-Pierre; Roux, Aurélie; Aubert, Julie; Junot, Christophe; Heilier, Jean-François; Landaud, Sophie; Bonnarme, Pascal

2013-01-01

Yarrowia lipolytica, located at the frontier of hemiascomycetous yeasts and fungi, is an excellent candidate for studies of metabolism evolution. This yeast, widely recognized for its technological applications, in particular produces volatile sulfur compounds (VSCs) that fully contribute to the flavor of smear cheese. We report here a relevant global vision of sulfur metabolism in Y. lipolytica based on a comparison between high- and low-sulfur source supplies (sulfate, methionine, or cystine) by combined approaches (transcriptomics, metabolite profiling, and VSC analysis). The strongest repression of the sulfate assimilation pathway was observed in the case of high methionine supply, together with a large accumulation of sulfur intermediates. A high sulfate supply seems to provoke considerable cellular stress via sulfite production, resulting in a decrease of the availability of the glutathione pathway's sulfur intermediates. The most limited effect was observed for the cystine supply, suggesting that the intracellular cysteine level is more controlled than that of methionine and sulfate. Using a combination of metabolomic profiling and genetic experiments, we revealed taurine and hypotaurine metabolism in yeast for the first time. On the basis of a phylogenetic study, we then demonstrated that this pathway was lost by some of the hemiascomycetous yeasts during evolution. PMID:23220962

The expression of the Cuphea palustris thioesterase CpFatB2 in Yarrowia lipolytica triggers oleic acid accumulation.

PubMed

Stefan, Alessandra; Hochkoeppler, Alejandro; Ugolini, Luisa; Lazzeri, Luca; Conte, Emanuele

2016-01-01

The conversion of industrial by-products into high-value added compounds is a challenging issue. Crude glycerol, a by-product of the biodiesel production chain, could represent an alternative carbon source for the cultivation of oleaginous yeasts. Here, we developed five minimal synthetic glycerol-based media, with different C/N ratios, and we analyzed the production of biomass and fatty acids by Yarrowia lipolytica Po1g strain. We identified two media at the expense of which Y. lipolytica was able to accumulate ∼5 g L(-1) of biomass and 0.8 g L(-1) of fatty acids (0.16 g of fatty acids per g of dry weight). These optimized media contained 0.5 g L(-1) of urea or ammonium sulfate and 20 g L(-1) of glycerol, and were devoid of yeast extract. Moreover, Y. lipolytica was engineered by inserting the FatB2 gene, coding for the CpFatB2 thioesterase from Cuphea palustris, in order to modify the fatty acid composition towards the accumulation of medium-chain fatty acids. Contrary to the expected, the expression of the heterologous gene increased the production of oleic acid, and concomitantly decreased the level of saturated fatty acids. © 2015 American Institute of Chemical Engineers.

Substrates and oxygen dependent citric acid production by Yarrowia lipolytica: insights through transcriptome and fluxome analyses.

PubMed

Sabra, Wael; Bommareddy, Rajesh Reddy; Maheshwari, Garima; Papanikolaou, Seraphim; Zeng, An-Ping

2017-05-08

Unlike the well-studied backer yeast where catabolite repression represents a burden for mixed substrate fermentation, Yarrowia lipolytica, an oleaginous yeast, is recognized for its potential to produce single cell oils and citric acid from different feedstocks. These versatilities of Y. lipolytica with regards to substrate utilization make it an attractive host for biorefinery application. However, to develop a commercial process for the production of citric acid by Y. lipolytica, it is necessary to better understand the primary metabolism and its regulation, especially for growth on mixed substrate. Controlling the dissolved oxygen concentration (pO 2 ) in Y. lipolytica cultures enhanced citric acid production significantly in cultures grown on glucose in mono- or dual substrate fermentations, whereas with glycerol as mono-substrate no significant effect of pO 2 was found on citrate production. Growth on mixed substrate with glucose and glycerol revealed a relative preference of glycerol utilization by Y. lipolytica. Under optimized conditions with pO 2 control, the citric acid titer on glucose in mono- or in dual substrate cultures was 55 and 50 g/L (with productivity of 0.6 g/L*h in both cultures), respectively, compared to a maximum of 18 g/L (0.2 g/L*h) with glycerol in monosubstrate culture. Additionally, in dual substrate fermentation, glycerol limitation was found to trigger citrate consumption despite the presence of enough glucose in pO 2 -limited culture. The metabolic behavior of this yeast on different substrates was investigated at transcriptomic and 13 C-based fluxomics levels. Upregulation of most of the genes of the pentose phosphate pathway was found in cultures with highest citrate production with glucose in mono- or in dual substrate fermentation with pO 2 control. The activation of the glyoxylate cycle in the oxygen limited cultures and the imbalance caused by glycerol limitation might be the reason for the re-consumption of citrate in

Citric acid production from hydrolysate of pretreated straw cellulose by Yarrowia lipolytica SWJ-1b using batch and fed-batch cultivation.

PubMed

Liu, Xiaoyan; Lv, Jinshun; Zhang, Tong; Deng, Yuanfang

2015-01-01

In this study, crude cellulase produced by Trichoderma reesei Rut-30 was used to hydrolyze pretreated straw. After the compositions of the hydrolysate of pretreated straw were optimized, the study showed that natural components of pretreated straw without addition of any other components such as (NH4)2SO4, KH2PO4, or Mg(2+) were suitable for citric acid production by Yarrowia lipolytica SWJ-1b, and the optimal ventilatory capacity was 10.0 L/min/L medium. Batch and fed-batch production of citric acid from the hydrolysate of pretreated straw by Yarrowia lipolytica SWJ-1b has been investigated. In the batch cultivation, 25.4 g/L and 26.7 g/L citric acid were yields from glucose and hydrolysate of straw cellulose, respectively, while the cultivation time was 120 hr. In the three-cycle fed-batch cultivation, citric acid (CA) production was increased to 42.4 g/L and the cultivation time was extended to 240 hr. However, iso-citric acid (ICA) yield in fed-batch cultivation (4.0 g/L) was similar to that during the batch cultivation (3.9 g/L), and only 1.6 g/L of reducing sugar was left in the medium at the end of fed-batch cultivation, suggesting that most of the added carbon was used in the cultivation.

Dual CRISPR-Cas9 Cleavage Mediated Gene Excision and Targeted Integration in Yarrowia lipolytica.

PubMed

Gao, Difeng; Smith, Spencer; Spagnuolo, Michael; Rodriguez, Gabriel; Blenner, Mark

2018-05-29

CRISPR-Cas9 technology has been successfully applied in Yarrowia lipolytica for targeted genomic editing including gene disruption and integration; however, disruptions by existing methods typically result from small frameshift mutations caused by indels within the coding region, which usually resulted in unnatural protein. In this study, a dual cleavage strategy directed by paired sgRNAs is developed for gene knockout. This method allows fast and robust gene excision, demonstrated on six genes of interest. The targeted regions for excision vary in length from 0.3 kb up to 3.5 kb and contain both non-coding and coding regions. The majority of the gene excisions are repaired by perfect nonhomologous end-joining without indel. Based on this dual cleavage system, two targeted markerless integration methods are developed by providing repair templates. While both strategies are effective, homology mediated end joining (HMEJ) based method are twice as efficient as homology recombination (HR) based method. In both cases, dual cleavage leads to similar or improved gene integration efficiencies compared to gene excision without integration. This dual cleavage strategy will be useful for not only generating more predictable and robust gene knockout, but also for efficient targeted markerless integration, and simultaneous knockout and integration in Y. lipolytica. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Omega-3 production by fermentation of Yarrowia lipolytica: From fed-batch to continuous.

PubMed

Xie, Dongming; Miller, Edward; Sharpe, Pamela; Jackson, Ethel; Zhu, Quinn

2017-04-01

The omega-3 fatty acid, cis-5,8,11,14,17-eicosapentaenoic acid (C20:5; EPA) has wide-ranging benefits in improving heart health, immune function, and mental health. A sustainable source of EPA production through fermentation of metabolically engineered Yarrowia lipolytica has been developed. In this paper, key fed-batch fermentation conditions were identified to achieve 25% EPA in the yeast biomass, which is so far the highest EPA titer reported in the literature. Dynamic models of the EPA fermentation process were established for analyzing, optimizing, and scaling up the fermentation process. In addition, model simulations were used to develop a two-stage continuous process and compare to single-stage continuous and fed- batch processes. The two stage continuous process, which is equipped with a smaller growth fermentor (Stage 1) and a larger production fermentor (Stage 2), was found to be a superior process to achieve high titer, rate, and yield of EPA. A two-stage continuous fermentation experiment with Y. lipolytica strain Z7334 was designed using the model simulation and then tested in a 2 L and 5 L fermentation system for 1,008 h. Compared with the standard 2 L fed-batch process, the two-stage continuous fermentation process improved the overall EPA productivity by 80% and EPA concentration in the fermenter by 40% while achieving comparable EPA titer in biomass and similar conversion yield from glucose. During the long-term experiment it was also found that the Y. lipolytica strain evolved to reduce byproduct and increase lipid production. This is one of the few continuous fermentation examples that demonstrated improved productivity and concentration of a final product with similar conversion yield compared with a fed-batch process. This paper suggests the two-stage continuous fermentation could be an effective process to achieve improved production of omega-3 and other fermentation products where non-growth or partially growth associated kinetics

Three alcohol dehydrogenase genes and one acetyl-CoA synthetase gene are responsible for ethanol utilization in Yarrowia lipolytica.

PubMed

Gatter, Michael; Ottlik, Stephanie; Kövesi, Zsolt; Bauer, Benjamin; Matthäus, Falk; Barth, Gerold

2016-10-01

The non-conventional yeast Yarrowia lipolytica is able to utilize a wide range of different substrates like glucose, glycerol, ethanol, acetate, proteins and various hydrophobic molecules. Although most metabolic pathways for the utilization of these substrates have been clarified by now, it was not clear whether ethanol is oxidized by alcohol dehydrogenases or by an alternative oxidation system inside the cell. In order to detect the genes that are required for ethanol utilization in Y. lipolytica, eight alcohol dehydrogenase (ADH) genes and one alcohol oxidase gene (FAO1) have been identified and respective deletion strains were tested for their ability to metabolize ethanol. As a result of this, we found that the availability of ADH1, ADH2 or ADH3 is required for ethanol utilization in Y. lipolytica. A strain with deletions in all three genes is lacking the ability to utilize ethanol as sole carbon source. Although Adh2p showed by far the highest enzyme activity in an in vitro assay, the availability of any of the three genes was sufficient to enable a decent growth. In addition to ADH1, ADH2 and ADH3, an acetyl-CoA synthetase encoding gene (ACS1) was found to be essential for ethanol utilization. As Y. lipolytica is a non-fermenting yeast, it is neither able to grow under anaerobic conditions nor to produce ethanol. To investigate whether Y. lipolytica may produce ethanol, the key genes of alcoholic fermentation in S. cerevisiae, ScADH1 and ScPDC1, were overexpressed in an ADH and an ACS1 deletion strain. However, instead of producing ethanol, the respective strains regained the ability to use ethanol as single carbon source and were still not able to grow under anaerobic conditions. Copyright © 2016 Elsevier Inc. All rights reserved.

Characterization of phosphatidic acid phosphatase activity in the oleaginous yeast Yarrowia lipolytica and its role in lipid biosynthesis.

PubMed

Hardman, Derell; McFalls, Daniel; Fakas, Stylianos

2017-02-01

Phosphatidic acid phosphatase (PAP) catalyses the committed step of triacylglycerol (TAG) biosynthesis and thus regulates the amounts of TAG produced by the cell. TAG is the target of biotechnological processes developed for the production of food lipids or biofuels. These processes are using oleaginous microorganisms like the yeast Yarrowia lipolytica as the TAG producers. Thus manipulating key enzymatic activities like PAP in Y. lipolytica could drive lipid biosynthesis towards TAG production and increase TAG yields. In this study, PAP activity in Y. lipolytica was characterized in detail and its role in lipid biosynthesis was addressed. PAP activity increased 2.5-fold with the addition of Mg 2+ (1 mm) in the assay mixture, which means that most of the PAP activity was due to Mg 2+ -dependent PAP enzymes (e.g. Pah1, App1). In contrast, N-ethylmaleimide (NEM) potently inhibited PAP activity, indicating the presence of NEM-sensitive PAP enzymes (e.g. App1, Lpp1). Localization studies revealed that the majority of PAP activity resides in the membrane fraction, while the cytosolic fraction harbours only a small amount of activity. PAP activity was regulated in a growth-dependent manner, being induced at the early exponential phase and declining thereafter. PAP activity did not correlate with TAG synthesis, which increased as cells progressed from the exponential phase to the early stationary phase. In stationary phase, TAG was mobilized with the concomitant synthesis of sterols and sterol esters. These results provide the first insights into the role of PAP in lipid biosynthesis by Y. lipolytica. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

Citric acid production in Yarrowia lipolytica SWJ-1b yeast when grown on waste cooking oil.

PubMed

Liu, Xiaoyan; Lv, Jinshun; Xu, Jiaxing; Zhang, Tong; Deng, Yuanfang; He, Jianlong

2015-03-01

In this study, citric acid was produced from waste cooking oil by Yarrowia lipolytica SWJ-1b. To get the maximal yield of citric acid, the compositions of the medium for citric acid production were optimized, and our results showed that extra nitrogen and magnesium rather than vitamin B1 and phosphate were needed for CA accumulation when using waste cooking oil. The results also indicated that the optimal initial concentration of the waste cooking oil in the medium for citric acid production was 80.0 g/l, and the ideal inoculation size was 1 × 10(7) cells/l of medium. We also reported that during 10-l fermentation, 31.7 g/l of citric acid, 6.5 g/l of isocitric acid, 5.9 g/l of biomass, and 42.1 g/100.0 g cell dry weight of lipid were attained from 80.0 g/l of waste cooking oil within 336 h. At the end of the fermentation, 94.6 % of the waste cooking oil was utilized by the cells of Y. lipolytica SWJ-1b, and the yield of citric acid was 0.4 g/g waste cooking oil, which suggested that waste cooking oil was a suitable carbon resource for citric acid production.

The RAD52 ortholog of Yarrowia lipolytica is essential for nuclear integrity and DNA repair.

PubMed

Campos-Góngora, Eduardo; Andaluz, Encarnación; Bellido, Alberto; Ruiz-Herrera, José; Larriba, German

2013-08-01

Yarrowia lipolytica (Yl) is a dimorphic fungus that has become a well-established model for a number of biological processes, including secretion of heterologous and chimerical proteins. However, little is known on the recombination machinery responsible for the integration in the genome of the exogenous DNA encoding for those proteins. We have carried out a phenotypic analysis of rad52 deletants of Y. lipolytica. YlRad52 exhibited 20-30% identity with Rad52 homologues of other eukaryotes, including Saccharomyces cerevisiae and Candida albicans. Ylrad52-Δ strains formed colonies on YPD-agar plates which were spinier and smaller than those from wild type, whereas in YPD liquid cultures they exhibited a decreased grow rate and contained cells with aberrant morphology and fragmented chromatin, supporting a role for homologous recombination (HR) in genome stability under nondamaging conditions. In addition, Ylrad52 mutants showed moderate to high sensitivity to UV light, oxidizing agents and compounds that cause single- (SSB) and double-strand breaks (DSB), indicating an important role for Rad52 in DNA repair. These findings extend to Yl previous observations indicating that RAD52 is a crucial gene for DNA repair in other fungi, including S. cerevisiae, C. albicans and Schizosaccharomyces pombe. © 2013 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

Sustainable source of omega-3 eicosapentaenoic acid from metabolically engineered Yarrowia lipolytica: from fundamental research to commercial production.

PubMed

Xie, Dongming; Jackson, Ethel N; Zhu, Quinn

2015-02-01

The omega-3 fatty acids, cis-5, 8, 11, 14, and 17-eicosapentaenoic acid (C20:5; EPA) and cis-4, 7, 10, 13, 16, and 19-docosahexaenoic acid (C22:6; DHA), have wide-ranging benefits in improving heart health, immune function, mental health, and infant cognitive development. Currently, the major source for EPA and DHA is from fish oil, and a minor source of DHA is from microalgae. With the increased demand for EPA and DHA, DuPont has developed a clean and sustainable source of the omega-3 fatty acid EPA through fermentation using metabolically engineered strains of Yarrowia lipolytica. In this mini-review, we will focus on DuPont's technology for EPA production. Specifically, EPA biosynthetic and supporting pathways have been introduced into the oleaginous yeast to synthesize and accumulate EPA under fermentation conditions. This Yarrowia platform can also produce tailored omega-3 (EPA, DHA) and/or omega-6 (ARA, GLA) fatty acid mixtures in the cellular lipid profiles. Fundamental research such as metabolic engineering for strain construction, high-throughput screening for strain selection, fermentation process development, and process scale-up were all needed to achieve the high levels of EPA titer, rate, and yield required for commercial application. Here, we summarize how we have combined the fundamental bioscience and the industrial engineering skills to achieve large-scale production of Yarrowia biomass containing high amounts of EPA, which led to two commercial products, New Harvest™ EPA oil and Verlasso® salmon.

Influence of oxygen availability on the metabolism and morphology of Yarrowia lipolytica: insights into the impact of glucose levels on dimorphism.

PubMed

Timoumi, Asma; Bideaux, Carine; Guillouet, Stéphane E; Allouche, Yohan; Molina-Jouve, Carole; Fillaudeau, Luc; Gorret, Nathalie

2017-10-01

Dynamic behavior of Yarrowia lipolytica W29 strain under conditions of fluctuating, low, and limited oxygen supply was characterized in batch and glucose-limited chemostat cultures. In batch cultures, transient oscillations between oxygen-rich and -deprived environments induced a slight citric acid accumulation (lower than 29 mg L -1 ). By contrast, no citric acid was detected in continuous fermentations for all stress conditions: full anoxia (zero pO 2 value, 100% N 2 ), limited (zero pO 2 value, 75% of cell needs), and low (pO 2 close to 2%) dissolved oxygen (DO) levels. The macroscopic behavior (kinetic parameters, yields, viability) of Y. lipolytica was not significantly affected by the exposure to DO fluctuations under both modes of culture. Nevertheless, conditions of oxygen limitation resulted in the destabilization of the glucose-limited growth during the continuous cultivations. Morphological responses of Y. lipolytica to DO oscillations were different between batch and chemostat runs. Indeed, a yeast-to-mycelium transition was induced and progressively intensified during the batch fermentations (filamentous subpopulation reaching 74% (v/v)). While, in chemostat bioreactors, the culture consisted mainly of yeast-like cells (mean diameter not exceeding 5.7 μm) with a normal size distribution. During the continuous cultures, growth at low DO concentration did not induce any changes in Y. lipolytica morphology. Dimorphism (up to 80.5% (v/v) of filaments) was only detected under conditions of oxygen limitation in the presence of a residual glucose excess (more than 0.75 g L -1 ). These data suggest an impact of glucose levels on the signaling pathways regulating dimorphic responses in Y. lipolytica.

An evolutionary metabolic engineering approach for enhancing lipogenesis in Yarrowia lipolytica.

PubMed

Liu, Leqian; Pan, Anny; Spofford, Caitlin; Zhou, Nijia; Alper, Hal S

2015-05-01

Lipogenic organisms provide an ideal platform for biodiesel and oleochemical production. Through our previous rational metabolic engineering efforts, lipogenesis titers in Yarrowia lipolytica were significantly enhanced. However, the resulting strain still suffered from decreased biomass generation rates. Here, we employ a rapid evolutionary metabolic engineering approach linked with a floating cell enrichment process to improve lipogenesis rates, titers, and yields. Through this iterative process, we were able to ultimately improve yields from our prior strain by 55% to achieve production titers of 39.1g/L with upwards of 76% of the theoretical maximum yield of conversation. Isolated cells were saturated with up to 87% lipid content. An average specific productivity of 0.56g/L/h was achieved with a maximum instantaneous specific productivity of 0.89g/L/h during the lipid production phase in fermentation. Genomic sequencing of the evolved strains revealed a link between a decrease/loss of function mutation of succinate semialdehyde dehydrogenase, uga2, suggesting the importance of gamma-aminobutyric acid assimilation in lipogenesis. This linkage was validated through gene deletion experiments. This work presents an improved host strain that can serve as a platform for efficient oleochemical production. Copyright © 2015 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

Regulation of nitrogen metabolism by GATA zinc finger transcription factors in Yarrowia lipolytica

DOE PAGES

Pomraning, Kyle R.; Bredeweg, Erin L.; Baker, Scott E.; ...

2017-02-15

Here, fungi accumulate lipids in a manner dependent on the quantity and quality of the nitrogen source on which they are growing. In the oleaginous yeast Yarrowia lipolytica, growth on a complex source of nitrogen enables rapid growth and limited accumulation of neutral lipids, while growth on a simple nitrogen source promotes lipid accumulation in large lipid droplets. Here we examined the roles of nitrogen catabolite repression and its regulation by GATA zinc finger transcription factors on lipid metabolism in Y. lipolytica. Deletion of the GATA transcription factor genes gzf3 and gzf2 resulted in nitrogen source-specific growth defects and greatermore » accumulation of lipids when the cells were growing on a simple nitrogen source. Deletion of gzf1, which is most similar to activators of genes repressed by nitrogen catabolite repression in filamentous ascomycetes, did not affect growth on the nitrogen sources tested. We examined gene expression of wild-type and GATA transcription factor mutants on simple and complex nitrogen sources and found that expression of enzymes involved in malate metabolism, beta-oxidation, and ammonia utilization are strongly upregulated on a simple nitrogen source. Deletion of gzf3 results in overexpression of genes with GATAA sites in their promoters, suggesting that it acts as a repressor, while gzf2 is required for expression of ammonia utilization genes but does not grossly affect the transcription level of genes predicted to be controlled by nitrogen catabolite repression. Both GATA transcription factor mutants exhibit decreased expression of genes controlled by carbon catabolite repression via the repressor mig1, including genes for beta-oxidation, highlighting the complex interplay between regulation of carbon, nitrogen, and lipid metabolism.« less

Regulation of nitrogen metabolism by GATA zinc finger transcription factors in Yarrowia lipolytica

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pomraning, Kyle R.; Bredeweg, Erin L.; Baker, Scott E.

Here, fungi accumulate lipids in a manner dependent on the quantity and quality of the nitrogen source on which they are growing. In the oleaginous yeast Yarrowia lipolytica, growth on a complex source of nitrogen enables rapid growth and limited accumulation of neutral lipids, while growth on a simple nitrogen source promotes lipid accumulation in large lipid droplets. Here we examined the roles of nitrogen catabolite repression and its regulation by GATA zinc finger transcription factors on lipid metabolism in Y. lipolytica. Deletion of the GATA transcription factor genes gzf3 and gzf2 resulted in nitrogen source-specific growth defects and greatermore » accumulation of lipids when the cells were growing on a simple nitrogen source. Deletion of gzf1, which is most similar to activators of genes repressed by nitrogen catabolite repression in filamentous ascomycetes, did not affect growth on the nitrogen sources tested. We examined gene expression of wild-type and GATA transcription factor mutants on simple and complex nitrogen sources and found that expression of enzymes involved in malate metabolism, beta-oxidation, and ammonia utilization are strongly upregulated on a simple nitrogen source. Deletion of gzf3 results in overexpression of genes with GATAA sites in their promoters, suggesting that it acts as a repressor, while gzf2 is required for expression of ammonia utilization genes but does not grossly affect the transcription level of genes predicted to be controlled by nitrogen catabolite repression. Both GATA transcription factor mutants exhibit decreased expression of genes controlled by carbon catabolite repression via the repressor mig1, including genes for beta-oxidation, highlighting the complex interplay between regulation of carbon, nitrogen, and lipid metabolism.« less

Production of Laccase by Recombinant Yarrowia lipolytica from Molasses: Bioprocess Development Using Statistical Modeling and Increase Productivity in Shake-Flask and Bioreactor Cultures.

PubMed

Darvishi, Farshad; Moradi, Marzieh; Madzak, Catherine; Jolivalt, Claude

2017-03-01

Laccases are used in numerous applications, from green degradation of various xenobiotic compounds, waste detoxification, textile dye bleaching, and delignification of lignocellulose materials to biofuel production. In this study, the recombinant Yarrowia lipolytica YL4 strain carrying the white-rot fungus Trametes versicolor laccase IIIb gene was used for laccase production from beet molasses as an agro-industrial residue. Response surface methodology was used to statistical optimization of the production of laccase by Y. lipolytica using an industrial medium containing molasses which allows a six times increase in laccase activity compared to primary medium contains glucose after 144 h. In bioreactor cultivation after 48 h, laccase production reached to 3.7- and 22.5-fold more than optimized and primary media in shake-flask cultures, respectively. Laccase productivity in bioreactor (0.0937 U/h) was higher than shake-flask culture (0.0084 U/h). The present study provides valuable information about statistical optimization of bioprocess development for cost-effective production of laccase and other heterologous proteins in Y. lipolytica from beet molasses as sole carbon source, thus allowing the valorization and decreasing environmental pollution of this agro-industrial waste.

Extracellular expression of YlLip11 with a native signal peptide from Yarrowia lipolytica MSR80 in three different yeast hosts.

PubMed

Kumari, Arti; Baronian, Keith; Kunze, Gotthard; Gupta, Rani

2015-06-01

Lipase YlLip11 from Yarrowia lipolytica was expressed with a signal peptide encoding sequence in Arxula adeninivorans, Saccharomyces cerevisiae and Hansenula polymorpha using the Xplor®2 transformation/expression platform and an expression module with the constitutive Arxula-derived TEF1 promoter. The YlLip11 signal peptide was functional in all of the yeast hosts with 97% of the recombinant enzyme being secreted into the culture medium. However, recombinant YlLip11 with His Tag fused at C-terminal was not active. The best recombinant YlLip11 producing A. adeninivorans G1212/YRC102-YlLip11 transformant cultivated in shake flasks produced 2654 U/L lipase, followed by S. cerevisiae SEY6210/YRC103-YlLip11 (1632U/L) and H. polymorpha RB11/YRC103-YlLip11 (1144U/L). Although the biochemical parameters of YlLip11 synthesized in different hosts were similar, their glycosylation level and thermo stability differed. The protein synthesized by the H. polymorpha transformant had the highest degree of glycosylation and with a t1/2 of 60min at 70°C, exhibited the highest thermostability. Copyright © 2015 Elsevier Inc. All rights reserved.

The Strictly Aerobic Yeast Yarrowia lipolytica Tolerates Loss of a Mitochondrial DNA-Packaging Protein

PubMed Central

Bakkaiova, Jana; Arata, Kosuke; Matsunobu, Miki; Ono, Bungo; Aoki, Tomoyo; Lajdova, Dana; Nebohacova, Martina; Nosek, Jozef; Miyakawa, Isamu

2014-01-01

Mitochondrial DNA (mtDNA) is highly compacted into DNA-protein structures termed mitochondrial nucleoids (mt-nucleoids). The key mt-nucleoid components responsible for mtDNA condensation are HMG box-containing proteins such as mammalian mitochondrial transcription factor A (TFAM) and Abf2p of the yeast Saccharomyces cerevisiae. To gain insight into the function and organization of mt-nucleoids in strictly aerobic organisms, we initiated studies of these DNA-protein structures in Yarrowia lipolytica. We identified a principal component of mt-nucleoids in this yeast and termed it YlMhb1p (Y. lipolytica mitochondrial HMG box-containing protein 1). YlMhb1p contains two putative HMG boxes contributing both to DNA binding and to its ability to compact mtDNA in vitro. Phenotypic analysis of a Δmhb1 strain lacking YlMhb1p resulted in three interesting findings. First, although the mutant exhibits clear differences in mt-nucleoids accompanied by a large decrease in the mtDNA copy number and the number of mtDNA-derived transcripts, its respiratory characteristics and growth under most of the conditions tested are indistinguishable from those of the wild-type strain. Second, our results indicate that a potential imbalance between subunits of the respiratory chain encoded separately by nuclear DNA and mtDNA is prevented at a (post)translational level. Third, we found that mtDNA in the Δmhb1 strain is more prone to mutations, indicating that mtHMG box-containing proteins protect the mitochondrial genome against mutagenic events. PMID:24972935

Modeling and optimization of lipid accumulation by Yarrowia lipolytica from glucose under nitrogen depletion conditions.

PubMed

Robles-Rodríguez, Carlos E; Muñoz-Tamayo, Rafael; Bideaux, Carine; Gorret, Nathalie; Guillouet, Stéphane E; Molina-Jouve, Carole; Roux, Gilles; Aceves-Lara, César A

2018-05-01

Oleaginous yeasts have been seen as a feasible alternative to produce the precursors of biodiesel due to their capacity to accumulate lipids as triacylglycerol having profiles with high content of unsaturated fatty acids. The yeast Yarrowia lipolytica is a promising microorganism that can produce lipids under nitrogen depletion conditions and excess of the carbon source. However, under these conditions, this yeast also produces citric acid (overflow metabolism) decreasing lipid productivity. This work presents two mathematical models for lipid production by Y. lipolytica from glucose. The first model is based on Monod and inhibition kinetics, and the second one is based on the Droop quota model approach, which is extended to yeast. The two models showed good agreements with the experimental data used for calibration and validation. The quota based model presented a better description of the dynamics of nitrogen and glucose dynamics leading to a good management of N/C ratio which makes this model interesting for control purposes. Then, quota model was used to evaluate, by means of simulation, a scenario for optimizing lipid productivity and lipid content. For that, a control strategy was designed by approximating the flow rates of glucose and nitrogen with piecewise linear functions. Simulation results achieved productivity of 0.95 g L -1  hr -1 and lipid content fraction of 0.23 g g -1 , which indicates that this strategy is a promising alternative for the optimization of lipid production. © 2017 Wiley Periodicals, Inc.

Overproduction of Fatty Acid Ethyl Esters by the Oleaginous Yeast Yarrowia lipolytica through Metabolic Engineering and Process Optimization.

PubMed

Gao, Qi; Cao, Xuan; Huang, Yu-Ying; Yang, Jing-Lin; Chen, Jun; Wei, Liu-Jing; Hua, Qiang

2018-05-18

Recent advances in the production of biofuels by microbes have attracted attention due to increasingly limited fossil fuels. Biodiesels, especially fatty acid ethyl esters (FAEEs), are considered a potentially fully sustainable fuel in the near future due to similarities with petrodiesels and compatibility with existing infrastructure. However, biosynthesis of FAEEs is limited by the supply of precursor lipids and acetyl-CoA. In the present study, we explored the production potential of an engineered biosynthetic pathway coupled to the addition of ethanol in the oleaginous yeast Yarrowia lipolytica. This type of yeast is able to supply a greater amount of precursor lipids than species typically used. To construct the FAEEs synthesis pathway, WS genes that encode wax ester synthases (WSs) from different species were codon-optimized and heterologously expressed in Y. lipolytica. The most productive engineered strain was found to express a WS gene from Marinobacter hydrocarbonoclasticus strain DSM 8798. To stepwisely increase FAEEs production, we optimized the promoter of WS overexpression, eliminated β-oxidation by deleting the PEX10 gene in our engineered strains, and redirected metabolic flux toward acetyl-CoA. The new engineered strain, coupled with an optimized ethanol concentration, led to an approximate 5.5-fold increase in extracellular FAEEs levels compared to the wild-type strain and a maximum FAEEs titer of 1.18 g/L in shake flask cultures. In summary, the present study demonstrated that an engineered Y. lipolytica strain possessed a high capacity for FAEEs production and may serve as a platform for more efficient biodiesel production in the future.

Inference and interrogation of a coregulatory network in the context of lipid accumulation in Yarrowia lipolytica.

PubMed

Trébulle, Pauline; Nicaud, Jean-Marc; Leplat, Christophe; Elati, Mohamed

2017-01-01

Complex phenotypes, such as lipid accumulation, result from cooperativity between regulators and the integration of multiscale information. However, the elucidation of such regulatory programs by experimental approaches may be challenging, particularly in context-specific conditions. In particular, we know very little about the regulators of lipid accumulation in the oleaginous yeast of industrial interest Yarrowia lipolytica . This lack of knowledge limits the development of this yeast as an industrial platform, due to the time-consuming and costly laboratory efforts required to design strains with the desired phenotypes. In this study, we aimed to identify context-specific regulators and mechanisms, to guide explorations of the regulation of lipid accumulation in Y. lipolytica . Using gene regulatory network inference, and considering the expression of 6539 genes over 26 time points from GSE35447 for biolipid production and a list of 151 transcription factors, we reconstructed a gene regulatory network comprising 111 transcription factors, 4451 target genes and 17048 regulatory interactions (YL-GRN-1) supported by evidence of protein-protein interactions. This study, based on network interrogation and wet laboratory validation (a) highlights the relevance of our proposed measure, the transcription factors influence, for identifying phases corresponding to changes in physiological state without prior knowledge (b) suggests new potential regulators and drivers of lipid accumulation and (c) experimentally validates the impact of six of the nine regulators identified on lipid accumulation, with variations in lipid content from +43.2% to -31.2% on glucose or glycerol.

A comparative study on glycerol metabolism to erythritol and citric acid in Yarrowia lipolytica yeast cells.

PubMed

Tomaszewska, Ludwika; Rakicka, Magdalena; Rymowicz, Waldemar; Rywińska, Anita

2014-09-01

Citric acid and erythritol biosynthesis from pure and crude glycerol by three acetate-negative mutants of Yarrowia lipolytica yeast was investigated in batch cultures in a wide pH range (3.0-6.5). Citric acid biosynthesis was the most effective at pH 5.0-5.5 in the case of Wratislavia 1.31 and Wratislavia AWG7. With a decreasing pH value, the direction of biosynthesis changed into erythritol synthesis accompanied by low production of citric acid. Pathways of glycerol conversion into erythritol and citric acid were investigated in Wratislavia K1 cells. Enzymatic activity was compared in cultures run at pH 3.0 and 4.5, that is, under conditions promoting the production of erythritol and citric acid, respectively. The effect of pH value (3.0 and 4.5) and NaCl presence on the extracellular production and intracellular accumulation of citric acid and erythritol was compared as well. Low pH and NaCl resulted in diminished activity of glycerol kinase, whereas such conditions stimulated the activity of glycerol-3-phosphate dehydrogenase. The presence of NaCl strongly influenced enzymes activity - the effective erythritol production was correlated with a high activity of transketolase and erythrose reductase. Therefore, presented results confirmed that transketolase and erythrose reductase are involved in the overproduction of erythritol in the cells of Y. lipolytica yeast. © 2014 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

Overlapping reading frames at the LYS5 locus in the yeast Yarrowia lipolytica.

PubMed Central

Xuan, J W; Fournier, P; Declerck, N; Chasles, M; Gaillardin, C

1990-01-01

Mutants affected at the LYS5 locus of Yarrowia lipolytica lack detectable dehydrogenase (SDH) activity. The LYS5 gene has previously been cloned, and we present here the sequence of the 2.5-kilobase-pair (kb) DNA fragment complementing the lys5 mutation. Two large antiparallel open reading frames (ORF1 and ORF2) were observed, flanked by potential transcription signals. Both ORFs appear to be transcribed, but several lines of evidence suggest that only ORF2 is translated and encodes SDH. (i) The global amino acid compositions of Saccharomyces cerevisiae SDH and of the putative ORF2 product are similar and that of ORF1 is dissimilar. (ii) An in-frame translational fusion of ORF2 with the Escherichia coli lacZ gene was introduced into yeast cells and resulted in a beta-galactosidase activity regulated similarly to SDH; no beta-galactosidase activity was obtained with an in-frame fusion of ORF1 with lacZ. (iii) The introduction of a stop codon at the beginning of ORF2 prevented SDH expression in yeast cells, whereas no phenotypic effect was observed when ORF1 translation was blocked. Images PMID:2388625

Leucine Biosynthesis Is Involved in Regulating High Lipid Accumulation in Yarrowia lipolytica

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kerkhoven, Eduard J.; Kim, Young-Mo; Wei, Siwei

ABSTRACT The yeastYarrowia lipolyticais a potent accumulator of lipids, and lipogenesis in this organism can be influenced by a variety of factors, such as genetics and environmental conditions. Using a multifactorial study, we elucidated the effects of both genetic and environmental factors on regulation of lipogenesis inY. lipolyticaand identified how two opposite regulatory states both result in lipid accumulation. This study involved comparison of a strain overexpressing diacylglycerol acyltransferase (DGA1) with a control strain grown under either nitrogen or carbon limitation conditions. A strong correlation was observed between the responses on the transcript and protein levels. Combination ofDGA1overexpression with nitrogen limitationmore » resulted in a high level of lipid accumulation accompanied by downregulation of several amino acid biosynthetic pathways, including that of leucine in particular, and these changes were further correlated with a decrease in metabolic fluxes. This downregulation was supported by the measured decrease in the level of 2-isopropylmalate, an intermediate of leucine biosynthesis. Combining the multi-omics data with putative transcription factor binding motifs uncovered a contradictory role for TORC1 in controlling lipid accumulation, likely mediated through 2-isopropylmalate and a Leu3-like transcription factor. IMPORTANCEThe ubiquitous metabolism of lipids involves refined regulation, and an enriched understanding of this regulation would have wide implications. Various factors can influence lipid metabolism, including the environment and genetics. We demonstrated, using a multi-omics and multifactorial experimental setup, that multiple factors affect lipid accumulation in the yeastYarrowia lipolytica. Using integrative analysis, we identified novel interactions between nutrient restriction and genetic factors involving regulators that are highly conserved among eukaryotes. Given that lipid metabolism is involved in many

Functional overexpression and characterization of lipogenesis-related genes in the oleaginous yeast Yarrowia lipolytica.

PubMed

Silverman, Andrew M; Qiao, Kangjian; Xu, Peng; Stephanopoulos, Gregory

2016-04-01

Single cell oil (SCO) is an attractive energy source due to scalability, utilization of low-cost renewable feedstocks, and type of product(s) made. Engineering strains capable of producing high lipid titers and yields is crucial to the economic viability of these processes. However, lipid synthesis in cells is a complex phenomenon subject to multiple layers of regulation, making gene target identification a challenging task. In this study, we aimed to identify genes in the oleaginous yeast Yarrowia lipolytica whose overexpression enhances lipid production by this organism. To this end, we examined the effect of the overexpression of a set of 44 native genes on lipid production in Y. lipolytica, including those involved in glycerolipid synthesis, fatty acid synthesis, central carbon metabolism, NADPH generation, regulation, and metabolite transport and characterized each resulting strain's ability to produce lipids growing on both glucose and acetate as a sole carbon source. Our results suggest that a diverse subset of genes was effective at individually influencing lipid production in Y. lipolytica, sometimes in a substrate-dependent manner. The most productive strain on glucose overexpressed the diacylglycerol acyltransferase DGA2 gene, increasing lipid titer, cellular content, and yield by 236, 165, and 246 %, respectively, over our control strain. On acetate, our most productive strain overexpressed the acylglycerol-phosphate acyltransferase SLC1 gene, with a lipid titer, cellular content, and yield increase of 99, 91, and 151 %, respectively, over the control strain. Aside from genes encoding enzymes that directly catalyze the reactions of lipid synthesis, other ways by which lipogenesis was increased in these cells include overexpressing the glycerol-3-phosphate dehydrogenase (GPD1) gene to increase production of glycerol head groups and overexpressing the 6-phosphogluconolactonase (SOL3) gene from the oxidative pentose phosphate pathway to increase NADPH

Physical and physiological impacts of different foam control strategies during a process involving hydrophobic substrate for the lipase production by Yarrowia lipolytica.

PubMed

Kar, Tambi; Destain, Jacqueline; Thonart, Philippe; Delvigne, Frank

2012-05-01

The potentialities for the intensification of the process of lipase production by the yeast Yarrowia lipolytica on a renewable hydrophobic substrate (methyl oleate) have to be investigated. The key factor governing the lipase yield is the intensification of the oxygen transfer rate, considering the fact that Y. lipolytica is a strict aerobe. However, considering the nature of the substrate and the capacity for protein excretion and biosurfactant production of Y. lipolytica, intensification of oxygen transfer rate is accompanied by an excessive formation of foam. Two different foam control strategies have thus been implemented: a classical chemical foam control strategy and a mechanical foam control (MFM) based on the Stirring As Foam Disruption principle. The second strategy allows foam control without any modifications of the physico-chemical properties of the broth. However, the MFM system design induced the formation of a persistent foam layer in the bioreactor. This phenomenon has led to the segregation of microbial cells between the foam phase and the liquid phase in the case of the bioreactors operated with MFM control, and induced a reduction at the level of the lipase yield. More interestingly, flow cytometry experiments have shown that the residence time of microbial cells in the foam phase tends to induce a dimorphic transition which could potentially explain the reduction of lipase excretion.

Developing cellulolytic Yarrowia lipolytica as a platform for the production of valuable products in consolidated bioprocessing of cellulose.

PubMed

Guo, Zhong-Peng; Robin, Julien; Duquesne, Sophie; O'Donohue, Michael Joseph; Marty, Alain; Bordes, Florence

2018-01-01

Both industrial biotechnology and the use of cellulosic biomass as feedstock for the manufacture of various commercial goods are prominent features of the bioeconomy. In previous work, with the aim of developing a consolidated bioprocess for cellulose bioconversion, we conferred cellulolytic activity of Yarrowia lipolytica , one of the most widely studied "nonconventional" oleaginous yeast species. However, further engineering this strain often leads to the loss of previously introduced heterologous genes due to the presence of multiple LoxP sites when using Cre -recombinase to remove previously employed selection markers. In the present study, we first optimized the strategy of expression of multiple cellulases and rescued selection makers to obtain an auxotrophic cellulolytic Y. lipolytica strain. Then we pursued the quest, exemplifying how this cellulolytic Y. lipolytica strain can be used as a CBP platform for the production of target products. Our results reveal that overexpression of SCD1 gene, encoding stearoyl-CoA desaturase, and DGA1 , encoding acyl-CoA:diacylglycerol acyltransferase, confers the obese phenotype to the cellulolytic Y. lipolytica . When grown in batch conditions and minimal medium, the resulting strain consumed 12 g/L cellulose and accumulated 14% (dry cell weight) lipids. Further enhancement of lipid production was achieved either by the addition of glucose or by enhancing cellulose consumption using a commercial cellulase cocktail. Regarding the latter option, although the addition of external cellulases is contrary to the concept of CBP, the amount of commercial cocktail used remained 50% lower than that used in a conventional process (i.e., without internalized production of cellulases). The introduction of the LIP2 gene into cellulolytic Y. lipolytica led to the production of a strain capable of producing lipase 2 while growing on cellulose. Remarkably, when the strain was grown on glucose, the expression of six cellulases did not

A survey of yeast from the Yarrowia clade for lipid production in dilute-acid pretreated lignocellulosic biomass hydrolysate

USDA-ARS?s Scientific Manuscript database

Yarrowia lipolytica is an oleaginous yeast species that has attracted attention as a model organism for synthesis of single cell oil. Among over 50 isolates of Y. lipolytica identified, only a few of the strains have been studied extensively. Furthermore, 12 other yeast species were recently assigne...

Identification of the Transcription Factor Znc1p, which Regulates the Yeast-to-Hypha Transition in the Dimorphic Yeast Yarrowia lipolytica

PubMed Central

Martinez-Vazquez, Azul; Gonzalez-Hernandez, Angelica; Domínguez, Ángel; Rachubinski, Richard; Riquelme, Meritxell; Cuellar-Mata, Patricia; Guzman, Juan Carlos Torres

2013-01-01

The dimorphic yeast Yarrowia lipolytica is used as a model to study fungal differentiation because it grows as yeast-like cells or forms hyphal cells in response to changes in environmental conditions. Here, we report the isolation and characterization of a gene, ZNC1, involved in the dimorphic transition in Y. lipolytica. The ZNC1 gene encodes a 782 amino acid protein that contains a Zn(II)2C6 fungal-type zinc finger DNA-binding domain and a leucine zipper domain. ZNC1 transcription is elevated during yeast growth and decreases during the formation of mycelium. Cells in which ZNC1 has been deleted show increased hyphal cell formation. Znc1p-GFP localizes to the nucleus, but mutations within the leucine zipper domain of Znc1p, and to a lesser extent within the Zn(II)2C6 domain, result in a mislocalization of Znc1p to the cytoplasm. Microarrays comparing gene expression between znc1::URA3 and wild-type cells during both exponential growth and the induction of the yeast-to-hypha transition revealed 1,214 genes whose expression was changed by 2-fold or more under at least one of the conditions analyzed. Our results suggest that Znc1p acts as a transcription factor repressing hyphal cell formation and functions as part of a complex network regulating mycelial growth in Y. lipolytica. PMID:23826133

Induction of a non-specific permeability transition in mitochondria from Yarrowia lipolytica and Dipodascus (Endomyces) magnusii yeasts.

PubMed

Kovaleva, Mariya V; Sukhanova, Evgeniya I; Trendeleva, Tatyana A; Zyl'kova, Marina V; Ural'skaya, Ludmila A; Popova, Kristina M; Saris, Nils-Erik L; Zvyagilskaya, Renata A

2009-06-01

In this study we used tightly-coupled mitochondria from Yarrowia lipolytica and Dipodascus (Endomyces) magnusii yeasts, possessing a respiratory chain with the usual three points of energy conservation. High-amplitude swelling and collapse of the membrane potential were used as parameters for demonstrating induction of the mitochondrial permeability transition due to opening of a pore (mPTP). Mitochondria from Y. lipolytica, lacking a natural mitochondrial Ca(2+) uptake pathway, and from D. magnusii, harboring a high-capacitive, regulated mitochondrial Ca(2+) transport system (Bazhenova et al. J Biol Chem 273:4372-4377, 1998a; Bazhenova et al. Biochim Biophys Acta 1371:96-100, 1998b; Deryabina and Zvyagilskaya Biochemistry (Moscow) 65:1352-1356, 2000; Deryabina et al. J Biol Chem 276:47801-47806, 2001) were very resistant to Ca(2+) overload. However, exposure of yeast mitochondria to 50-100 microM Ca(2+) in the presence of the Ca(2+) ionophore ETH129 induced collapse of the membrane potential, possibly due to activation of the fatty acid-dependent Ca(2+)/nH(+)-antiporter, with no classical mPTP induction. The absence of response in yeast mitochondria was not simply due to structural limitations, since large-amplitude swelling occurred in the presence of alamethicin, a hydrophobic, helical peptide, forming voltage-sensitive ion channels in lipid membranes. Ca(2+)- ETH129-induced activation of the Ca(2+)/H(+)-antiport system was inhibited and prevented by bovine serum albumin, and partially by inorganic phosphate and ATP. We subjected yeast mitochondria to other conditions known to induce the permeability transition in animal mitochondria, i.e., Ca(2+) overload (in the presence of ETH129) combined with palmitic acid (Mironova et al. J Bioenerg Biomembr 33:319-331, 2001; Sultan and Sokolove Arch Biochem Biophys 386:37-51, 2001), SH-reagents, carboxyatractyloside (an inhibitor of the ADP/ATP translocator), depletion of intramitochondrial adenine nucleotide pools

High-throughput fermentation screening for the yeast Yarrowia lipolytica with real-time monitoring of biomass and lipid production.

PubMed

Back, Alexandre; Rossignol, Tristan; Krier, François; Nicaud, Jean-Marc; Dhulster, Pascal

2016-08-23

Because the model yeast Yarrowia lipolytica can synthesize and store lipids in quantities up to 20 % of its dry weight, it is a promising microorganism for oil production at an industrial scale. Typically, optimization of the lipid production process is performed in the laboratory and later scaled up for industrial production. However, the scale-up process can be complicated by genetic modifications that are optimized for one set of growing conditions can confer a less-than-optimal phenotype in a different environment. To address this issue, small cultivation systems have been developed that mimic the conditions in benchtop bioreactors. In this work, we used one such microbioreactor system, the BioLector, to develop high-throughput fermentation procedures that optimize growth and lipid accumulation in Y. lipolytica. Using this system, we were able to monitor lipid and biomass production in real time throughout the culture duration. The BioLector can monitor the growth of Y. lipolytica in real time by evaluating scattered light; this produced accurate measurements until cultures reached an equivalent of OD600nm = 115 and a cell dry weight of 100 g L(-1). In addition, a lipid-specific fluorescent probe was applied which reliably monitored lipid production up to a concentration of 12 g L(-1). Through screening various growing conditions, we determined that a carbon/nitrogen ratio of 35 was the most efficient for lipid production. Further screening showed that ammonium chloride and glycerol were the most valuable nitrogen and carbon sources, respectively, for growth and lipid production. Moreover, a carbon concentration above 1 M appeared to impair growth and lipid accumulation. Finally, we used these optimized conditions to screen engineered strains of Y. lipolytica with high lipid-accumulation capability. The growth and lipid content of the strains cultivated in the BioLector were compared to those grown in benchtop bioreactors. To our knowledge, this is the

High performance microbiological transformation of L-tyrosine to L-dopa by Yarrowia lipolytica NRRL-143

PubMed Central

Ali, Sikander; Shultz, Jeffry L; Ikram-ul-Haq

2007-01-01

Background The 3,4-dihydroxy phenyl L-alanine (L-dopa) is a drug of choice for Parkinson's disease, controlling changes in energy metabolism enzymes of the myocardium following neurogenic injury. Aspergillus oryzae is commonly used for L-dopa production; however, potential improvements in ease of handling, growth rate and environmental impact have led to an interest in exploiting alternative yeasts. The two important elements required for L-dopa production are intracellular tyrosinases (thus pre-grown yeast cells are required for the transformation of L-tyrosine to L-dopa) and L-ascorbate, which acts as a reducing agent. Results Pre-grown cells of Yarrowia lipolytica NRRL-143 were used for the microbiological transformation of L-tyrosine to L-dopa. Different diatomite concentrations (0.5–3.0 mg/ml) were added to the acidic (pH 3.5) reaction mixture. Maximum L-dopa biosynthesis (2.96 mg/ml L-dopa from 2.68 mg/ml L-tyrosine) was obtained when 2.0 mg/ml diatomite was added 15 min after the start of the reaction. After optimizing reaction time (30 min), and yeast cell concentration (2.5 mg/ml), an overall 12.5 fold higher L-dopa production rate was observed when compared to the control. Significant enhancements in Yp/s, Qs and qs over the control were observed. Conclusion Diatomite (2.0 mg/ml) addition 15 min after reaction commencement improved microbiological transformation of L-tyrosine to L-dopa (3.48 mg/ml; p ≤ 0.05) by Y. lipolytica NRRL-143. A 35% higher substrate conversion rate was achieved when compared to the control. PMID:17705832

L-Phenylalanine catabolism and 2-phenylethanol synthesis in Yarrowia lipolytica--mapping molecular identities through whole-proteome quantitative mass spectrometry analysis.

PubMed

Celińska, Ewelina; Olkowicz, Mariola; Grajek, Włodzimierz

2015-08-01

A world-wide effort is now being pursued towards the development of flavors and fragrances (F&F) production independently from traditional sources, as well as autonomously from depleting fossil fuel supplies. Biotechnological production of F&F by microbes has emerged as a vivid solution to the current market limitations. Amongst a wide variety of fragrant chemicals, 2-PE is of significant interest to both scientific and industrial community. Although the general overview of the 2-PE synthesis pathway is commonly known, involvement of particular molecular identities in this pathway has not been elucidated in Yarrowia lipolytica to date. The aim of this study was mapping molecular identities involved in 2-PE synthesis in Y. lipolytica. To acquire a comprehensive landscape of the proteins that are directly and indirectly involved in L-Phe degradation and 2-PE synthesis, we took advantage of comprehensibility and sensitivity of high-throughput LC-MS/MS-quantitative analysis. Amongst a number of proteins involved in amino acid turnover and the central carbon metabolism, enzymes involved in L-Phe conversion to 2-PE have been identified. Results on yeast-to-hyphae transition in relation to the character of the provided nitrogen source have been presented. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Regulation of Nitrogen Metabolism by GATA Zinc Finger Transcription Factors in Yarrowia lipolytica

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pomraning, Kyle R.; Bredeweg, Erin L.; Baker, Scott E.

ABSTRACT Fungi accumulate lipids in a manner dependent on the quantity and quality of the nitrogen source on which they are growing. In the oleaginous yeastYarrowia lipolytica, growth on a complex source of nitrogen enables rapid growth and limited accumulation of neutral lipids, while growth on a simple nitrogen source promotes lipid accumulation in large lipid droplets. Here we examined the roles of nitrogen catabolite repression and its regulation by GATA zinc finger transcription factors on lipid metabolism inY. lipolytica. Deletion of the GATA transcription factor genesgzf3andgzf2resulted in nitrogen source-specific growth defects and greater accumulation of lipids when the cells weremore » growing on a simple nitrogen source. Deletion ofgzf1, which is most similar to activators of genes repressed by nitrogen catabolite repression in filamentous ascomycetes, did not affect growth on the nitrogen sources tested. We examined gene expression of wild-type and GATA transcription factor mutants on simple and complex nitrogen sources and found that expression of enzymes involved in malate metabolism, beta-oxidation, and ammonia utilization are strongly upregulated on a simple nitrogen source. Deletion ofgzf3results in overexpression of genes with GATAA sites in their promoters, suggesting that it acts as a repressor, whilegzf2is required for expression of ammonia utilization genes but does not grossly affect the transcription level of genes predicted to be controlled by nitrogen catabolite repression. Both GATA transcription factor mutants exhibit decreased expression of genes controlled by carbon catabolite repression via the repressormig1, including genes for beta-oxidation, highlighting the complex interplay between regulation of carbon, nitrogen, and lipid metabolism. IMPORTANCENitrogen source is commonly used to control lipid production in industrial fungi. Here we identified regulators of nitrogen catabolite repression in the oleaginous yeast

Molecular Characterization of the Elaeis guineensis Medium-Chain Fatty Acid Diacylglycerol Acyltransferase DGAT1-1 by Heterologous Expression in Yarrowia lipolytica.

PubMed

Aymé, Laure; Jolivet, Pascale; Nicaud, Jean-Marc; Chardot, Thierry

2015-01-01

Diacylglycerol acyltransferases (DGAT) are involved in the acylation of sn-1,2-diacylglycerol. Palm kernel oil, extracted from Elaeis guineensis (oil palm) seeds, has a high content of medium-chain fatty acids mainly lauric acid (C12:0). A putative E. guineensis diacylglycerol acyltransferase gene (EgDGAT1-1) is expressed at the onset of lauric acid accumulation in the seed endosperm suggesting that it is a determinant of medium-chain triacylglycerol storage. To test this hypothesis, we thoroughly characterized EgDGAT1-1 activity through functional complementation of a Yarrowia lipolytica mutant strain devoid of neutral lipids. EgDGAT1-1 expression is sufficient to restore triacylglycerol accumulation in neosynthesized lipid droplets. A comparative functional study with Arabidopsis thaliana DGAT1 highlighted contrasting substrate specificities when the recombinant yeast was cultured in lauric acid supplemented medium. The EgDGAT1-1 expressing strain preferentially accumulated medium-chain triacylglycerols whereas AtDGAT1 expression induced long-chain triacylglycerol storage in Y. lipolytica. EgDGAT1-1 localized to the endoplasmic reticulum where TAG biosynthesis takes place. Reestablishing neutral lipid accumulation in the Y. lipolytica mutant strain did not induce major reorganization of the yeast microsomal proteome. Overall, our findings demonstrate that EgDGAT1-1 is an endoplasmic reticulum DGAT with preference for medium-chain fatty acid substrates, in line with its physiological role in palm kernel. The characterized EgDGAT1-1 could be used to promote medium-chain triacylglycerol accumulation in microbial-produced oil for industrial chemicals and cosmetics.

Molecular Characterization of the Elaeis guineensis Medium-Chain Fatty Acid Diacylglycerol Acyltransferase DGAT1-1 by Heterologous Expression in Yarrowia lipolytica

PubMed Central

Aymé, Laure; Jolivet, Pascale; Nicaud, Jean-Marc; Chardot, Thierry

2015-01-01

Diacylglycerol acyltransferases (DGAT) are involved in the acylation of sn-1,2-diacylglycerol. Palm kernel oil, extracted from Elaeis guineensis (oil palm) seeds, has a high content of medium-chain fatty acids mainly lauric acid (C12:0). A putative E. guineensis diacylglycerol acyltransferase gene (EgDGAT1-1) is expressed at the onset of lauric acid accumulation in the seed endosperm suggesting that it is a determinant of medium-chain triacylglycerol storage. To test this hypothesis, we thoroughly characterized EgDGAT1-1 activity through functional complementation of a Yarrowia lipolytica mutant strain devoid of neutral lipids. EgDGAT1-1 expression is sufficient to restore triacylglycerol accumulation in neosynthesized lipid droplets. A comparative functional study with Arabidopsis thaliana DGAT1 highlighted contrasting substrate specificities when the recombinant yeast was cultured in lauric acid supplemented medium. The EgDGAT1-1 expressing strain preferentially accumulated medium-chain triacylglycerols whereas AtDGAT1 expression induced long-chain triacylglycerol storage in Y. lipolytica. EgDGAT1-1 localized to the endoplasmic reticulum where TAG biosynthesis takes place. Reestablishing neutral lipid accumulation in the Y. lipolytica mutant strain did not induce major reorganization of the yeast microsomal proteome. Overall, our findings demonstrate that EgDGAT1-1 is an endoplasmic reticulum DGAT with preference for medium-chain fatty acid substrates, in line with its physiological role in palm kernel. The characterized EgDGAT1-1 could be used to promote medium-chain triacylglycerol accumulation in microbial-produced oil for industrial chemicals and cosmetics. PMID:26581109

Proteomic analysis of the response of α-ketoglutarate-producer Yarrowia lipolytica WSH-Z06 to environmental pH stimuli.

PubMed

Guo, Hongwei; Wan, Hui; Chen, Hongwen; Fang, Fang; Liu, Song; Zhou, Jingwen

2016-10-01

During bioproduction of short-chain carboxylates, a shift in pH is a common strategy for enhancing the biosynthesis of target products. Based on two-dimensional gel electrophoresis, comparative proteomics analysis of general and mitochondrial protein samples was used to investigate the cellular responses to environmental pH stimuli in the α-ketoglutarate overproducer Yarrowia lipolytica WSH-Z06. The lower environmental pH stimuli tensioned intracellular acidification and increased the level of reactive oxygen species (ROS). A total of 54 differentially expressed protein spots were detected, and 11 main cellular processes were identified to be involved in the cellular response to environmental pH stimuli. Slight decrease in cytoplasmic pH enhanced the cellular acidogenicity by elevating expression level of key enzymes in tricarboxylic acid cycle (TCA cycle). Enhanced energy biosynthesis, ROS elimination, and membrane potential homeostasis processes were also employed as cellular defense strategies to compete with environmental pH stimuli. Owing to its antioxidant role of α-ketoglutarate, metabolic flux shifted to α-ketoglutarate under lower pH by Y. lipolytica in response to acidic pH stimuli. The identified differentially expressed proteins provide clues for understanding the mechanisms of the cellular responses and for enhancing short-chain carboxylate production through metabolic engineering or process optimization strategies in combination with manipulation of environmental conditions.

Ylpex5 mutation partially suppresses the defective hyphal growth of a Yarrowia lipolytica ceramide synthase mutant, Yllac1, by recovering lipid raft polarization and vacuole morphogenesis.

PubMed

Bal, Jyotiranjan; Lee, Hye-Jeong; Cheon, Seon Ah; Lee, Kyung Jin; Oh, Doo-Byoung; Kim, Jeong-Yoon

2013-01-01

Sphingolipids are involved in cell differentiation and morphogenesis in eukaryotic cells. In this study, YlLac1p, a ceramide synthase required for glucosylceramide (GlcCer) synthesis, was found to be essential for hyphal growth in Yarrowia lipolytica. Y. lipolytica GlcCer was shown to be composed of a C16:0 fatty acid, which is hydroxylated at C2, and a C18:2 long chain base, which is unsaturated at both C4 and C8 and methylated at C9. Domain swapping analysis revealed that the entire TRAM/Lag1/CLN8 (TLC) domain, not the Lag1 motif, is crucial for the function of YlLac1p. YlDes1p, the C4 desaturase of the ceramide synthesized by YlLac1p, was also required for Y. lipolytica morphogenesis. Both Yllac1Δ and Yldes1Δ mutants neither polarize lipid rafts nor form normal vacuoles. Interestingly, mutation in YlPEX5, which encode a peroxisomal targeting signal receptor, partially suppressed the defective hyphal growth of Yllac1Δ. The Yllac1ΔYlpex5Δ mutant restored the ability to polarize lipid rafts and to form normal vacuoles, although it could not synthesize GlcCer. Taken together, our results suggest that GlcCer or GlcCer derivatives may be involved in hyphal morphogenesis in Y. lipolytica, at least in part, by affecting polarization of lipid rafts and vacuole morphogenesis. Copyright © 2012 Elsevier Inc. All rights reserved.

Dynamic behavior of Yarrowia lipolytica in response to pH perturbations: dependence of the stress response on the culture mode.

PubMed

Timoumi, Asma; Cléret, Mégane; Bideaux, Carine; Guillouet, Stéphane E; Allouche, Yohan; Molina-Jouve, Carole; Fillaudeau, Luc; Gorret, Nathalie

2017-01-01

Yarrowia lipolytica, a non-conventional yeast with a promising biotechnological potential, is able to undergo metabolic and morphological changes in response to environmental conditions. The effect of pH perturbations of different types (pulses, Heaviside) on the dynamic behavior of Y. lipolytica W29 strain was characterized under two modes of culture: batch and continuous. In batch cultures, different pH (4.5, 5.6 (optimal condition), and 7) were investigated in order to identify the pH inducing a stress response (metabolic and/or morphologic) in Y. lipolytica. Macroscopic behavior (kinetic parameters, yields, viability) of the yeast was slightly affected by pH. However, contrary to the culture at pH 5.6, a filamentous growth was induced in batch experiments at pH 4.5 and 7. Proportions of the filamentous subpopulation reached 84 and 93 % (v/v) under acidic and neutral conditions, respectively. Given the significant impact of neutral pH on morphology, pH perturbations from 5.6 to 7 were subsequently assayed in batch and continuous bioreactors. For both process modes, the growth dynamics remained fundamentally unaltered during exposure to stress. Nevertheless, morphological behavior of the yeast was dependent on the culture mode. Specifically, in batch bioreactors where cells proliferated at their maximum growth rate, mycelia were mainly formed. Whereas, in continuous cultures at controlled growth rates (from 0.03 to 0.20 h -1 ) even closed to the maximum growth rate of the stain (0.24 h -1 ), yeast-like forms predominated. This pointed out differences in the kinetic behavior of filamentous and yeast subpopulations, cell age distribution, and pH adaptive mechanisms between both modes of culture.

Optimization of a low-cost hyperosmotic medium and establishing the fermentation kinetics of erythritol production by Yarrowia lipolytica from crude glycerol.

PubMed

Yang, Li-Bo; Zhan, Xiao-Bei; Zhu, Li; Gao, Min-Jie; Lin, Chi-Chung

2016-05-18

The production of erythritol by Yarrowia lipolytica from low-cost substitutable substrates for high yield was investigated. Crude glycerol, urea, and NaCl related to osmotic pressure were the most significant factors affecting erythritol production. An artificial neural network model and genetic algorithm were used to search the optimal composition of the significant factors and locate the resulting erythritol yield. Medium with 232.39 g/L crude glycerol, 1.57 g/L urea, and 31.03 g/L NaCl led to predictive maximum erythritol concentration of 110.7 g/L. The erythritol concentration improved from 50.4 g/L to 109.2 g/L with the optimized medium, which was reproducible. Erythritol fermentation kinetics were investigated in a batch system. Multistep fermentation kinetic models with hyperosmotic inhibitory effects were developed. The resulting mathematical equations provided a good description of temporal variations such as microbial growth (X), substrate consumption (S), and product formation (P) in erythritol fermentation. The accordingly derived model is the first reported model for fermentative erythritol production from glycerol, providing useful information to optimize the growth of Y. lipolytica and contributing visual description for the erythritol fermentation process under high osmotic pressure, as well as improvement of productivity and efficiency.

Improvement of erythrose reductase activity, deletion of by-products and statistical media optimization for enhanced erythritol production from Yarrowia lipolytica mutant 49.

PubMed

Ghezelbash, Gholam Reza; Nahvi, Iraj; Emamzadeh, Rahman

2014-08-01

The purpose of the present investigation was to produce erythritol by Yarrowia lipolytica mutant without any by-products. Mutants of Y. lipolytica were generated by ultra-violet for enhancing erythrose reductase (ER) activity and erythritol production. The mutants showing the highest ER activity were screened by triphenyl tetrazolium chloride agar plate assay. Productivity of samples was analyzed by thin-layer chromatography and high-performance liquid chromatography equipped with the refractive index detector. One of the mutants named as mutant 49 gave maximum erythritol production without any other by-products (particularly glycerol). Erythritol production and specific ER activity in mutant 49 increased to 1.65 and 1.47 times, respectively, in comparison with wild-type strain. The ER gene of wild and mutant strains was sequenced and analyzed. A general comparison of wild and mutant gene sequences showed the replacement of Asp(270) with Glu(270) in ER protein. In order to enhance erythritol production, we used a three component-three level-one response Box-Behnken of response surface methodology model. The optimum medium composition for erythritol production was found to be (g/l) glucose 279.49, ammonium sulfate 9.28, and pH 5.41 with 39.76 erythritol production.

Yarrowia lipolytica NCIM 3589, a tropical marine yeast, degrades bromoalkanes by an initial hydrolytic dehalogenation step.

PubMed

Vatsal, Aakanksha; Zinjarde, Smita S; Kumar, Ameeta Ravi

2015-04-01

The widespread industrial use of organobromines which are known persistent organic pollutants has led to their accumulation in sediments and water bodies causing harm to animals and humans. While degradation of organochlorines by bacteria is well documented, information regarding degradation pathways of these recalcitrant organobromines is scarce. Hence, their fates and effects on the environment are of concern. The present study shows that a tropical marine yeast, Yarrowia lipolytica NCIM 3589 aerobically degrades bromoalkanes differing in carbon chain length and position of halogen substitution viz., 2-bromopropane (2-BP), 1-bromobutane (1-BB), 1,5 dibromopentane (1,5-DBP) and 1-bromodecane (1-BD) as seen by an increase in cell mass, release of bromide and concomitant decrease in concentration of brominated compound. The amount of bromoalkane degraded was 27.3, 21.9, 18.0 and 38.3 % with degradation rates of 0.076, 0.058, 0.046 and 0.117/day for 2-BP, 1-BB, 1,5-DBP and 1-BD, respectively. The initial product formed respectively were alcohols viz., 2-propanol, 1-butanol, 1-bromo, 5-pentanol and 1-decanol as detected by GC-MS. These were further metabolized to fatty acids viz., 2-propionic, 1-butyric and 1-decanoic acid eventually leading to carbon dioxide formation. Neither higher chain nor brominated fatty acids were detected. An inducible extracellular dehalogenase responsible for removal of bromide was detected with activities of 21.07, 18.82, 18.96 and 26.67 U/ml for 2-BP, 1-BB, 1,5-DBP and 1-BD, respectively. We report here for the first time the proposed aerobic pathway of bromoalkane degradation by an eukaryotic microbe Y. lipolytica 3589, involving an initial hydrolytic dehalogenation step.

[Activation of the alternative oxidase of Yarrowia lipolytica by adenosine 5'-monophosphate].

PubMed

Medentsev, A G; Arinbasarova, A Iu; Smirnova, N M; Akimenko, V K

2004-01-01

The study of the effect of nucleoside phosphates on the activity of cyanide-resistant oxidase in the mitochondria and the submitochondrial particles of Yarrowia lipolytica showed that adenosine monophosphate (5'-AMP, AMP) did not stimulate the respiration of the intact mitochondria. The incubation of the mitochondria at room temperature (25 degrees C) for 3-5 h or their treatment with ultrasound, phospholipase A, and detergent Triton X-100 at a low temperature inactivated the cyanide-resistant alternative oxidase. The inactivated alternative oxidase could be reactivated by AMP. The reactivating effect of AMP was enhanced by azolectin. Some other nucleoside phosphates also showed reactivating ability in the following descending order. AMP = GMP > GDP > GTP > XMP > IMP. The apparent reaction rate constant Km for AMP upon the reactivation of the alternative oxidase of mitochondria treated with Triton X-100 or incubated at 25 degrees C was 12.5 and 20 microM, respectively. The Km for AMP upon the reactivation of the alternative oxidase of submitochondrial particles was 15 microM. During the incubation of yeast cells under conditions promoting the development of alternative oxidase, the content of adenine nucleotides (AMP, ADP, and ATP) in the cells and their respiration tended to decrease. The subsequent addition of cyanide to the cells activated their respiration, diminished the intracellular content of ATP three times, and augmented the content of AMP five times. These data suggest that the stimulation of cell respiration by cyanide may be due to the activation of alternative oxidase by AMP.

Design of an efficient medium for heterologous protein production in Yarrowia lipolytica: case of human interferon alpha 2b.

PubMed

Gasmi, Najla; Ayed, Atef; Nicaud, Jean-Marc; Kallel, Héla

2011-05-20

The non conventional yeast Yarrowia lipolytica has aroused a strong industrial interest for heterologous protein production. However most of the studies describing recombinant protein production by this yeast rely on the use of complex media, such media are not convenient for large scale production particularly for products intended for pharmaceutical applications. In addition medium composition can also affect the production yield. Hence it is necessary to design an efficient medium for therapeutic protein expression by this host. Five different media, including four minimal media and a complex medium, were assessed in shake flasks for the production of human interferon alpha 2b (hIFN α2b) by Y. lipolytica under the control of POX2 promoter inducible with oleic acid. The chemically defined medium SM4 formulated by Invitrogen for Pichia pastoris growth was the most suitable. Using statistical experimental design this medium was further optimized. The selected minimal medium consisting in SM4 supplemented with 10 mg/l FeCl₃, 1 g/l glutamate, 5 ml/l PTM1 (Pichia Trace Metals) solution and a vitamin solution composed of myo-inositol, thiamin and biotin was called GNY medium. Compared to shake flask, bioreactor culture in GNY medium resulted in 416-fold increase of hIFN α2b production and 2-fold increase of the biological activity. Furthermore, SM4 enrichment with 5 ml/l PTM1 solution contributed to protect hIFN α2b against the degradation by the 28 kDa protease identified by zymography gel in culture supernatant. The screening of the inhibitory effect of the trace elements present in PTM1 solution on the activity of this protease was achieved using a Box-Behnken design. Statistical data analysis showed that FeCl₃ and MnSO₄ had the most inhibitory effect. We have designed an efficient medium for large scale production of heterologous proteins by Y. lipolytica. The optimized medium GNY is suitable for the production of hIFN α2b with the advantage that no

MHY1 Encodes a C2H2-Type Zinc Finger Protein That Promotes Dimorphic Transition in the Yeast Yarrowia lipolytica

PubMed Central

Hurtado, Cleofe A. R.; Rachubinski, Richard A.

1999-01-01

The yeast-to-hypha morphological transition (dimorphism) is typical of many pathogenic fungi. Dimorphism has been attributed to changes in temperature and nutritional status and is believed to constitute a mechanism of response to adverse conditions. We have isolated and characterized a gene, MHY1, whose transcription is dramatically increased during the yeast-to-hypha transition in Yarrowia lipolytica. Deletion of MHY1 is viable and has no effect on mating, but it does result in a complete inability of cells to undergo mycelial growth. MHY1 encodes a C2H2-type zinc finger protein, Mhy1p, which can bind putative cis-acting DNA stress response elements, suggesting that Mhy1p may act as a transcription factor. Interestingly, Mhy1p tagged with a hemagglutinin epitope was concentrated in the nuclei of actively growing cells found at the hyphal tip. PMID:10322005

Heterologous expression of lipases YLIP4, YLIP5, YLIP7, YLIP13, and YLIP15 from Yarrowia lipolytica MSR80 in Escherichia coli: Substrate specificity, kinetic comparison, and enantioselectivity.

PubMed

Syal, Poonam; Gupta, Rani

2017-11-01

Five lipase genes, ylip4, ylip5, ylip7, ylip13, and ylip15, from Yarrowia lipolytica MSR80 were cloned and expressed in the pEZZ18-HB101 system. The lipases shared maximum sequence identity with Candida galli lipase, whereas they shared structural similarity with YLIP2 of Y. lipolytica CLIB122. The enzymes, purified using IgG sepharose, had specific activities in the range of 7-25 U mg -1 . Biochemical characteristics of all the lipases varied with respect to thermostability, substrate specificity, and enantioselectivity. All the enzymes were most active at neutral or slightly alkaline pH and were stable in the pH range 3.0-8.0, except YLIP4, which showed 50% stability at pH 10.0. Temperature optima of all the lipases varied from 30 to 50 ºC. YLIP15 and YLIP13 were most thermostable with a t 1/2 of 138 and 112 Min, respectively, at 60 °C. The lipases exhibited varied substrate specificity on p-nitrophenyl esters ranging from short-chain specificity (YLIP15), mid-chain specificity (YLIP4, YLIP5, YLIP7), and long-chain specificity (YLIP13). Catalytic efficiency on p-nitrophenylcaprate was highest for YLIP13 (67 × 10 3 mM -1 min -1 ) and lowest for YLIP15 (6.7 × 10 3 mM -1 min -1 ). YLIP13 was S-enantioselective, and YLIP15 was R-enantioselective with enantiomeric excess of 53 and 36%, respectively. Of all five lipases, YLIP13 and YLIP15 could be considered as industrially important enzymes as they were thermostable and enantioselective. © 2016 International Union of Biochemistry and Molecular Biology, Inc.

Gene repression via multiplex gRNA strategy in Y. lipolytica.

PubMed

Zhang, Jin-Lai; Peng, Yang-Zi; Liu, Duo; Liu, Hong; Cao, Ying-Xiu; Li, Bing-Zhi; Li, Chun; Yuan, Ying-Jin

2018-04-20

The oleaginous yeast Yarrowia lipolytica is a promising microbial cell factory due to their biochemical characteristics and native capacity to accumulate lipid-based chemicals. To create heterogenous biosynthesis pathway and manipulate metabolic flux in Y. lipolytica, numerous studies have been done for developing synthetic biology tools for gene regulation. CRISPR interference (CRISPRi), as an emerging technology, has been applied for specifically repressing genes of interest. In this study, we established CRISPRi systems in Y. lipolytica based on four different repressors, that was DNase-deactivated Cpf1 (dCpf1) from Francisella novicida, deactivated Cas9 (dCas9) from Streptococcus pyogenes, and two fusion proteins (dCpf1-KRAB and dCas9-KRAB). Ten gRNAs that bound to different regions of gfp gene were designed and the results indicated that there was no clear correlation between the repression efficiency and targeting sites no matter which repressor protein was used. In order to rapidly yield strong gene repression, a multiplex gRNAs strategy based on one-step Golden-brick assembly technology was developed. High repression efficiency 85% (dCpf1) and 92% (dCas9) were achieved in a short time by making three different gRNAs towards gfp gene simultaneously, which avoided the need of screening effective gRNA loci in advance. Moreover, two genes interference including gfp and vioE and three genes repression including vioA, vioB and vioE in protodeoxy-violaceinic acid pathway were also realized. Taken together, successful CRISPRi-mediated regulation of gene expression via four different repressors dCpf1, dCas9, dCpf1-KRAB and dCas9-KRAB in Y. lipolytica is achieved. And we demonstrate a multiplexed gRNA targeting strategy can efficiently achieve transcriptional simultaneous repression of several targeted genes and different sites of one gene using the one-step Golden-brick assembly. This timesaving method promised to be a potent transformative tool valuable for

Harnessing the Effect of pH on Lipid Production in Batch Cultures of Yarrowia lipolytica SKY7.

PubMed

Kuttiraja, Mathiazhakan; Dhouha, Ayed; Tyagi, Rajeshwar Dayal

2018-04-01

The objective of this research was to investigate the kinetics of lipid production by Yarrowia lipolytica SKY7 in the crude glycerol-supplemented media with and without the control of pH. Lipid and citric acid production were improved with the pH control condition. There was no significant difference observed in the biomass concentration with or without the pH control. In the pH-controlled experiments, the biomass and lipid concentration reached 18 and 7.78 g/L, (45.5% w/w), respectively, with lipid yield (Yp/s) of 0.179 g/g at 60 h of fermentation. The lipid production was directly correlated with growth and the process was defined as growth associated. After 60 h of fermentation, the lipid degradation was noticed in the pH-controlled reactor whereas it occurred after 84 h in the pH-uncontrolled reactor. Apart from lipid, citric acid was produced as the major extracellular product in both fermentations but the much lower concentration in uncontrolled pH. Based on the experimental results, it is evident that controlling the pH will enhance the lipid production by 15% compared to pH-uncontrolled fermentation.

Production of oils and fats by oleaginous microorganisms with an emphasis given to the potential of the nonconventional yeast Yarrowia lipolytica.

PubMed

Carsanba, E; Papanikolaou, S; Erten, H

2018-05-15

Recently, there has been a great upsurge of interest in studies related to several aspects of microbial lipid production, which is one of the top topics in relevant research fields due to the high demand of these fatty materials in food, medical, oleochemical and biofuel industries. Lipid accumulation by the so-called "oleaginous microorganisms" can generate more than 20% w/w of oil in dry biomass and is governed by a plethora of parameters, such as medium pH, incubation temperature, nutrient limitation and C/N (carbon/nitrogen) ratio, which drastically affect the lipid production bioprocess. Until now, considerable work has been undertaken to find the cheapest substrate to enable lipid fermentation by oleaginous microorganisms. This review principally details information regarding microbial lipids, suitable production conditions and focuses attention on using the yeast Yarrowia lipolytica to achieve these objectives. Lipid production by this yeast is discussed and the necessary conditions and suitable substrates are reviewed.

Irradiation of Yarrowia lipolytica NRRL YB-567 creating novel strains with enhanced ammonia and oil production on protein and carbohydrate substrates.

PubMed

Lindquist, Mitch R; López-Núñez, Juan Carlos; Jones, Marjorie A; Cox, Elby J; Pinkelman, Rebecca J; Bang, Sookie S; Moser, Bryan R; Jackson, Michael A; Iten, Loren B; Kurtzman, Cletus P; Bischoff, Kenneth M; Liu, Siqing; Qureshi, Nasib; Tasaki, Kenneth; Rich, Joseph O; Cotta, Michael A; Saha, Badal C; Hughes, Stephen R

2015-11-01

Increased interest in sustainable production of renewable diesel and other valuable bioproducts is redoubling efforts to improve economic feasibility of microbial-based oil production. Yarrowia lipolytica is capable of employing a wide variety of substrates to produce oil and valuable co-products. We irradiated Y. lipolytica NRRL YB-567 with UV-C to enhance ammonia (for fertilizer) and lipid (for biodiesel) production on low-cost protein and carbohydrate substrates. The resulting strains were screened for ammonia and oil production using color intensity of indicators on plate assays. Seven mutant strains were selected (based on ammonia assay) and further evaluated for growth rate, ammonia and oil production, soluble protein content, and morphology when grown on liver infusion medium (without sugars), and for growth on various substrates. Strains were identified among these mutants that had a faster doubling time, produced higher maximum ammonia levels (enzyme assay) and more oil (Sudan Black assay), and had higher maximum soluble protein levels (Bradford assay) than wild type. When grown on plates with substrates of interest, all mutant strains showed similar results aerobically to wild-type strain. The mutant strain with the highest oil production and the fastest doubling time was evaluated on coffee waste medium. On this medium, the strain produced 0.12 g/L ammonia and 0.20 g/L 2-phenylethanol, a valuable fragrance/flavoring, in addition to acylglycerols (oil) containing predominantly C16 and C18 residues. These mutant strains will be investigated further for potential application in commercial biodiesel production.

D-stat culture for studying the metabolic shifts from oxidative metabolism to lipid accumulation and citric acid production in Yarrowia lipolytica.

PubMed

Ochoa-Estopier, Abril; Guillouet, Stéphane E

2014-01-20

Lipid accumulation in oleaginous yeasts is triggered by nutrient imbalance in the culture medium between the carbon source in excess and the nitrogen source in limiting concentration. However Yarrowia lipolytica when cultivated on glucose as the sole carbon source, mainly produces citric acid upon nitrogen limitation over lipid accumulation (only 5-10% triacylglycerol). Therefore for developing bioprocess for the production of triacylglycerol from renewable carbon source as glucose it is of first importance to control this imbalance in order to avoid citric acid production during TAG accumulation. Using D-stat cultivation system, where the N/C was linearly decreased using a constant change rate we were able to identify the N/C ratio inducing TAG accumulation (0.085NmolCmol(-1)) and citric acid (0.021NmolCmol(-1)). We therefore demonstrated that it was possible to accumulate lipids without excretion citric acid as long as the N/C was within this indicated range. Moreover enzyme specific activities measurement during the D-stat indicated that ATP-citrate lyase, malic enzyme and acetyl-coA carboxylase were strongly induced at the onset of lipid accumulation and showed different patterns when citric acid was excreted. Our results give relevant information for future industrial bioprocess development concerning the production of lipids using renewable carbohydrate substrates as an alternative way to produce synthons for fuel or chemical industry. By controlling the N/C over the fermentation process on glucose Y. lipolytica can accumulate lipids without excreting citric acid. Copyright © 2013 Elsevier B.V. All rights reserved.

Engineering Yarrowia lipolytica to Simultaneously Produce Lipase and Single Cell Protein from Agro-industrial Wastes for Feed.

PubMed

Yan, Jinyong; Han, Bingnan; Gui, Xiaohua; Wang, Guilong; Xu, Li; Yan, Yunjun; Madzak, Catherine; Pan, Dujie; Wang, Yaofeng; Zha, Genhan; Jiao, Liangcheng

2018-01-15

Lipases are scarcely exploited as feed enzymes in hydrolysis of lipids for increasing energy supply and improving nutrient use efficiency. In this work, we performed homologous overexpression, in vitro characterization and in vivo assessment of a lipase from the yeast Yarrowia lipolytica for feed purpose. Simultaneously, a large amount of yeast cell biomass was produced, for use as single cell protein, a potential protein-rich feed resource. Three kinds of low cost agro-industrial wastes were tested as substrates for simultaneous production of lipase and single cell protein (SCP) as feed additives: sugarcane molasses, waste cooking oil and crude glycerol from biodiesel production. Sugarcane molasses appeared as the most effective cheap medium, allowing production of 16420 U/ml of lipase and 151.2 g/L of single cell protein at 10 liter fermentation scale. In vitro characterization by mimicking a gastro-intestinal environment and determination of essential amino acids of the SCP, and in vivo oral feeding test on fish all revealed that lipase, SCP and their combination were excellent feed additives. Such simultaneous production of this lipase and SCP could address two main concerns of feed industry, poor utilization of lipid and shortage of protein resource at the same time.

SOA genes encode proteins controlling lipase expression in response to triacylglycerol utilization in the yeast Yarrowia lipolytica.

PubMed

Desfougères, Thomas; Haddouche, Ramdane; Fudalej, Franck; Neuvéglise, Cécile; Nicaud, Jean-Marc

2010-02-01

The oleaginous yeast Yarrowia lipolytica efficiently metabolizes hydrophobic substrates such as alkanes, fatty acids or triacylglycerol. This yeast has been identified in oil-polluted water and in lipid-rich food. The enzymes involved in lipid breakdown, for use as a carbon source, are known, but the molecular mechanisms controlling the expression of the genes encoding these enzymes are still poorly understood. The study of mRNAs obtained from cells grown on oleic acid identified a new group of genes called SOA genes (specific for oleic acid). SOA1 and SOA2 are two small genes coding for proteins with no known homologs. Single- and double-disrupted strains were constructed. Wild-type and mutant strains were grown on dextrose, oleic acid and triacylglycerols. The double mutant presents a clear phenotype consisting of a growth defect on tributyrin and triolein, but not on dextrose or oleic acid media. Lipase activity was 50-fold lower in this mutant than in the wild-type strain. The impact of SOA deletion on the expression of the main extracellular lipase gene (LIP2) was monitored using a LIP2-beta-galactosidase promoter fusion protein. These data suggest that Soa proteins are components of a molecular mechanism controlling lipase gene expression in response to extracellular triacylglycerol.

Integrated Approach To Producing High-Purity Trehalose from Maltose by the Yeast Yarrowia lipolytica Displaying Trehalose Synthase (TreS) on the Cell Surface.

PubMed

Li, Ning; Wang, Hengwei; Li, Lijuan; Cheng, Huiling; Liu, Dawen; Cheng, Hairong; Deng, Zixin

2016-08-10

An alternative strategy that integrated enzyme production, trehalose biotransformation, and bioremoval in one bioreactor was developed in this study, thus simplifying the traditional procedures used for trehalose production. The trehalose synthase gene from a thermophilic archaea, Picrophilus torridus, was first fused to the YlPir1 anchor gene and then inserted into the genome of Yarrowia lipolytica, thus yielding an engineered yeast strain. The trehalose yield reached 73% under optimal conditions. The thermal and pH stabilities of the displayed enzyme were improved compared to those of its free form purified from recombinant Escherichia coli. After biotransformation, the glucose byproduct and residual maltose were directly fermented to ethanol by a Saccharomyces cerevisiae strain. Ethanol can be separated by distillation, and high-purity trehalose can easily be obtained from the fermentation broth. The results show that this one-pot procedure is an efficient approach to the economical production of trehalose from maltose.

Green and sustainable succinic acid production from crude glycerol by engineered Yarrowia lipolytica via agricultural residue based in situ fibrous bed bioreactor.

PubMed

Li, Chong; Gao, Shi; Yang, Xiaofeng; Lin, Carol Sze Ki

2018-02-01

In situ fibrous bed bioreactor (isFBB) for efficient succinic acid (SA) production by Yarrowia lipolytica was firstly developed in our former study. In this study, agricultural residues including wheat straw, corn stalk and sugarcane bagasse were investigated for the improvement of isFBB, and sugarcane bagasse was demonstrated to be the best immobilization material. With crude glycerol as the sole carbon source, optimization for isFBB batch fermentation was carried out. Under the optimal conditions of 20g sugarcane bagasse as immobilization material, 120gL -1 crude glycerol as carbon source and 4Lmin -1 of aeration rate, the resultant SA concentration was 53.6gL -1 with an average productivity of 1.45gL -1 h -1 and a SA yield of 0.45gg -1 . By feeding crude glycerol, SA titer up to 209.7gL -1 was obtained from fed batch fermentation, which was the highest value that ever reported. Copyright © 2017 Elsevier Ltd. All rights reserved.

Mitochondria from Dipodascus (Endomyces) magnusii and Yarrowia lipolytica yeasts did not undergo a Ca²⁺-dependent permeability transition even under anaerobic conditions.

PubMed

Trendeleva, Tat'yana; Sukhanova, Evgeniya; Ural'skaya, Ludmila; Saris, Nils-Erik; Zvyagilskaya, Renata

2011-12-01

In this study we used tightly-coupled mitochondria from Yarrowia lipolytica and Dipodascus (Endomyces) magnusii yeasts. The two yeast strains are good alternatives to Saccharomyces cerevisiae, being aerobes containing well structured mitochondria (thus ensuring less structural limitation to observe their appreciable swelling) and fully competent respiratory chain with three invariantly functioning energy conservation points, including Complex I, that can be involved in induction of the canonical Ca²⁺/P(i)-dependent mitochondrial permeability transition (mPTP pore) with an increased open probability when electron flux increases(Fontaine et al. J Biol Chem 273: 25734–25740, 1998; Bernardi et al. FEBS J 273:2077–2099, 2006). High amplitude swelling and collapse of the membrane potential were used as parameters for demonstrating pore opening. Previously (Kovaleva et al. J Bioenerg Biomembr 41:239–249, 2009; Kovaleva et al. Biochemistry (Moscow) 75: 297–303, 2010) we have shown that mitochondria from Y.lipolytica and D. magnusii were very resistant to the Ca²⁺overload combined with varying concentrations of P(i),palmitic acid, SH-reagents, carboxyatractyloside (an inhibitor of ADP/ATP translocator), as well as depletion of intramitochondrial adenine nucleotide pools, deenergization of mitochondria, and shifting to acidic pH values in the presence of high [P(i)]. Here we subjected yeast mitochondria to other conditions known to induce an mPTP in animal and plant mitochondria, namely to Ca²⁺ overload under hypoxic conditions (anaerobiosis). We were unable to observe Ca²⁺-induced high permeability of the inner membrane of D. magnusii and Y. lipolytica yeast mitochondria under anaerobic conditions, thus suggesting that an mPTP-like pore, if it ever occurs in yeast mitochondria, is not coupled with the Ca²⁺ uptake. The results provide the first demonstration of ATP-dependent energization of yeast mitochondria under conditions of anaerobiosis.

Integrating Cellular and Bioprocess Engineering in the Non-Conventional Yeast Yarrowia lipolytica for Biodiesel Production: A Review

PubMed Central

Xie, Dongming

2017-01-01

As one of the major biofuels to replace fossil fuel, biodiesel has now attracted more and more attention due to its advantages in higher energy density and overall less greenhouse gas generation. Biodiesel (fatty acid alkyl esters) is produced by chemically or enzymatically catalyzed transesterification of lipids from microbial cells, microalgae, oil crops, or animal fats. Currently, plant oils or waste cooking oils/fats remain the major source for biodiesel production via enzymatic route, but the production capacity is limited either by the uncertain supplement of plant oils or by the low or inconsistent quality of waste oils/fats. In the past decades, significant progresses have been made on synthesis of microalgae oils directly from CO2 via a photosynthesis process, but the production cost from any current technologies is still too high to be commercialized due to microalgae’s slow growth rate on CO2, inefficiency in photo-bioreactors, lack of efficient contamination control methods, and high cost in downstream recovery. At the same time, many oleaginous microorganisms have been studied to produce lipids via the fatty acid synthesis pathway under aerobic fermentation conditions, among them one of the most studied is the non-conventional yeast, Yarrowia lipolytica, which is able to produce fatty acids at very high titer, rate, and yield from various economical substrates. This review summarizes the recent research progresses in both cellular and bioprocess engineering in Y. lipolytica to produce lipids at a low cost that may lead to commercial-scale biodiesel production. Specific technologies include the strain engineering for using various substrates, metabolic engineering in high-yield lipid synthesis, cell morphology study for efficient substrate uptake and product formation, free fatty acid formation and secretion for improved downstream recovery, and fermentation engineering for higher productivities and less operating cost. To further improve the

Use of Plackett-Burman design for rapid screening of nitrogen and carbon sources for the production of lipase in solid state fermentation by Yarrowia lipolytica from mustard oil cake (Brassica napus).

PubMed

Imandi, Sarat Babu; Karanam, Sita Kumari; Garapati, Hanumantha Rao

2013-01-01

Mustard oil cake (Brassica napus), the residue obtained after extraction of mustard oil from mustard oil seeds, was investigated for the production of lipase under solid state fermentation (SSF) using the marine yeast Yarrowia lipolytica NCIM 3589. Process parameters such as incubation time, biomass concentration, initial moisture content, carbon source concentration and nitrogen source concentration of the medium were optimized. Screening of ten nitrogen and five carbon sources has been accomplished with the help of Plackett-Burman design. The highest lipase activity of 57.89 units per gram of dry fermented substrate (U/gds) was observed with the substrate of mustard oil cake in four days of fermentation.

The ‘LipoYeasts’ project: using the oleaginous yeast Yarrowia lipolytica in combination with specific bacterial genes for the bioconversion of lipids, fats and oils into high‐value products

PubMed Central

Sabirova, Julia S.; Haddouche, R.; Van Bogaert, I. N.; Mulaa, F.; Verstraete, W.; Timmis, K. N.; Schmidt‐Dannert, C.; Nicaud, J. M.; Soetaert, W.

2011-01-01

Summary The oleochemical industry is currently still dominated by conventional chemistry, with biotechnology only starting to play a more prominent role, primarily with respect to the biosurfactants or lipases, e.g. as detergents, or for biofuel production. A major bottleneck for all further biotechnological applications is the problem of the initial mobilization of cheap and vastly available lipid and oil substrates, which are then to be transformed into high‐value biotechnological, nutritional or pharmacological products. Under the EU‐sponsored LipoYeasts project we are developing the oleaginous yeast Yarrowia lipolytica into a versatile and high‐throughput microbial factory that, by use of specific enzymatic pathways from hydrocarbonoclastic bacteria, efficiently mobilizes lipids by directing its versatile lipid metabolism towards the production of industrially valuable lipid‐derived compounds like wax esters (WE), isoprenoid‐derived compounds (carotenoids, polyenic carotenoid ester), polyhydroxyalkanoates (PHAs) and free hydroxylated fatty acids (HFAs). Different lipid stocks (petroleum, alkane, vegetable oil, fatty acid) and combinations thereof are being assessed as substrates in combination with different mutant and recombinant strains of Y. lipolytica, in order to modulate the composition and yields of the produced added‐value products. PMID:21255371

Improved performance of Yarrowia lipolytica lipase-catalyzed kinetic resolution of (R,S)-2-octanol by an integrated strategy of interfacial activation, bioimprinting and immobilization.

PubMed

Liu, Ying; Guo, Chen; Sun, Xi-Tong; Liu, Chun-Zhao

2013-08-01

Yarrowia lipolytica lipase (YLL) demonstrated an (R)-enantiopreference for efficient resolution of (R,S)-2-octanol. The activity, enantioselectivity, the ratio of substrate to enzyme, acetaldehyde tolerance, and operational stability of YLL were improved by an integrated strategy of interfacial activation, bioimprinting, and immobilization. In comparison with the control, both the enzymatic activity and enantioselectivity increased by a factor of 8.85 and 2.75 by the integrated strategy, respectively. Fifty-one percentage of conversion with 220 of enantioselectivity was obtained using the immobilized YLL prepared by the integrated strategy at a ratio of 104 of substrate to enzyme loaded. The immobilized YLL retained 97% of its initial activity without a decrease in enantioselectivity after 10 successive reuse cycles. Together these results will result in a promising strategy with the YYL for efficient resolution of (R,S)-2-octanol in practice. Copyright © 2013 Elsevier Ltd. All rights reserved.

Deciphering how LIP2 and POX2 promoters can optimally regulate recombinant protein production in the yeast Yarrowia lipolytica.

PubMed

Sassi, Hosni; Delvigne, Frank; Kar, Tambi; Nicaud, Jean-Marc; Coq, Anne-Marie Crutz-Le; Steels, Sebastien; Fickers, Patrick

2016-09-20

In recent years, the non-conventional model yeast species Yarrowia lipolytica has received much attention because it is a useful cell factory for producing recombinant proteins. In this species, expression vectors involving LIP2 and POX2 promoters have been developed and used successfully for protein production at yields similar to or even higher than those of other cell factories, such as Pichia pastoris. However, production processes involving these promoters can be difficult to manage, especially if carried out at large scales in fed-batch bioreactors, because they require hydrophobic inducers, such as oleic acid or methyl oleate. Thus, the challenge has become to reduce loads of hydrophobic substrates while simultaneously promoting recombinant protein production. One possible solution is to replace a portion of the inducer with a co-substrate that can serve as an alternative energy source. However, implementing such an approach would require detailed knowledge of how carbon sources impact promoter regulation, which is surprisingly still lacking for the LIP2 and POX2 promoters. This study's aim was thus to better characterize promoter regulation and cell metabolism in Y. lipolytica cultures grown in media supplemented with different carbon sources. pPOX2 induction could be detected when glucose or glycerol was used as sole carbon source, which meant these carbon source could not prevent promoter induction. In addition, when a mixture of glucose and oleic acid was used in complex medium, pPOX2 induction level was lower that that of pLIP2. In contrast, pLIP2 induction was absent when glucose was present in the culture medium, which meant that cell growth could occur without any recombinant gene expression. When a 40/60 mixture of glucose and oleic acid (w/w) was used, a tenfold increase in promoter induction, as compared to when an oleic-acid-only medium was observed. It was also clear that individual cells were adapting metabolically to use both glucose and oleic

Enhancement of methanol resistance of Yarrowia lipolytica lipase 2 using β-cyclodextrin as an additive: Insights from experiments and molecular dynamics simulation.

PubMed

Cao, Hao; Jiang, Yang; Zhang, Haiyang; Nie, Kaili; Lei, Ming; Deng, Li; Wang, Fang; Tan, Tianwei

2017-01-01

The methanol resistance of lipase is a critical parameter in enzymatic biodiesel production. In the present work, the methanol resistance of Yarrowia lipolytica Lipase 2 (YLLIP2) was significantly improved using β-cyclodextrin (β-CD) as an additive. According to the results, YLLIP2 with β-CD exhibited approximately 7000U/mg specific activity in 30wt% methanol for 60min compared with no activity without β-CD under the same conditions. Molecular dynamics (MD) simulation results indicated that the β-CD molecules weakened the conformational change of YLLIP2 and maintained a semi-open state of the lid by overcoming the interference caused by methanol molecules. Furthermore, the β-CD molecule could directly stabilize "pathway" regions (e.g., Asp61-Asp67) and indirectly stabilize "pathway" regions (e.g., Gly44-Phe50) by forming hydrogen bonds with "pathway" regions and nearby "pathway" regions, respectively. The regions stabilized by the β-CD molecule then prevented the closure of active pockets, thus retaining the enzymatic activity of YLLIP2 with β-CD in methanol solvent. Copyright © 2016. Published by Elsevier Inc.

Citric acid production from partly deproteinized whey under non-sterile culture conditions using immobilized cells of lactose-positive and cold-adapted Yarrowia lipolytica B9.

PubMed

Arslan, Nazli Pinar; Aydogan, Mehmet Nuri; Taskin, Mesut

2016-08-10

The present study was performed to produce citric acid (CA) from partly deproteinized cheese whey (DPCW) under non-sterile culture conditions using immobilized cells of the cold-adapted and lactose-positive yeast Yarrowia lipolytica B9. DPCW was prepared using the temperature treatment of 90°C for 15min. Sodium alginate was used as entrapping agent for cell immobilization. Optimum conditions for the maximum CA production (33.3g/L) in non-sterile DPCW medium were the temperature of 20°C, pH 5.5, additional lactose concentration of 20g/L, sodium alginate concentration of 2%, number of 150 beads/100mL and incubation time of 120h. Similarly, maximum citric acid/isocitric acid (CA/ICA) ratio (6.79) could be reached under these optimal conditions. Additional nitrogen and phosphorus sources decreased CA concentration and CA/ICA ratio. Immobilized cells were reused in three continuous reaction cycles without any loss in the maximum CA concentration. The unique combination of low pH and temperature values as well as cell immobilization procedure could prevent undesired microbial contaminants during CA production. This is the first work on CA production by cold-adapted microorganisms under non-sterile culture conditions. Besides, CA production using a lactose-positive strain of the yeast Y. lipolytica was investigated for the first time in the present study. Copyright © 2016 Elsevier B.V. All rights reserved.

Storage lipids of yeasts: a survey of nonpolar lipid metabolism in Saccharomyces cerevisiae, Pichia pastoris, and Yarrowia lipolytica.

PubMed

Koch, Barbara; Schmidt, Claudia; Daum, Günther

2014-09-01

Biosynthesis and storage of nonpolar lipids, such as triacylglycerols (TG) and steryl esters (SE), have gained much interest during the last decades because defects in these processes are related to severe human diseases. The baker's yeast Saccharomyces cerevisiae has become a valuable tool to study eukaryotic lipid metabolism because this single-cell microorganism harbors many enzymes and pathways with counterparts in mammalian cells. In this article, we will review aspects of TG and SE metabolism and turnover in the yeast that have been known for a long time and combine them with new perceptions of nonpolar lipid research. We will provide a detailed insight into the mechanisms of nonpolar lipid synthesis, storage, mobilization, and degradation in the yeast S. cerevisiae. The central role of lipid droplets (LD) in these processes will be addressed with emphasis on the prevailing view that this compartment is more than only a depot for TG and SE. Dynamic and interactive aspects of LD with other organelles will be discussed. Results obtained with S. cerevisiae will be complemented by recent investigations of nonpolar lipid research with Yarrowia lipolytica and Pichia pastoris. Altogether, this review article provides a comprehensive view of nonpolar lipid research in yeast. © 2014 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

Citric acid production from extract of Jerusalem artichoke tubers by the genetically engineered yeast Yarrowia lipolytica strain 30 and purification of citric acid.

PubMed

Wang, Ling-Fei; Wang, Zhi-Peng; Liu, Xiao-Yan; Chi, Zhen-Ming

2013-11-01

In this study, citric acid production from extract of Jerusalem artichoke tubers by the genetically engineered yeast Yarrowia lipolytica strain 30 was investigated. After the compositions of the extract of Jerusalem artichoke tubers for citric acid production were optimized, the results showed that natural components of extract of Jerusalem artichoke tubers without addition of any other components were suitable for citric acid production by the yeast strain. During 10 L fermentation using the extract containing 84.3 g L(-1) total sugars, 68.3 g L(-1) citric acid was produced and the yield of citric acid was 0.91 g g(-1) within 336 h. At the end of the fermentation, 9.2 g L(-1) of residual total sugar and 2.1 g L(-1) of reducing sugar were left in the fermented medium. At the same time, citric acid in the supernatant of the culture was purified. It was found that 67.2 % of the citric acid in the supernatant of the culture was recovered and purity of citric acid in the crystal was 96 %.

Draft Genome Sequence of the Dimorphic Yeast Yarrowia lipolytica Strain W29

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pomraning, Kyle R.; Baker, Scott E.

Here, we present the draft genome sequence of the dimorphic ascomycete yeastYarrowia lipolyticastrain W29 (ATCC 20460).Y. lipolyticais a commonly employed model for the industrial production of lipases, small molecules, and more recently for its ability to accumulate lipids.

Multi-responses optimization of simultaneous biosorption of cationic dyes by live yeast Yarrowia lipolytica 70562 from binary solution: Application of first order derivative spectrophotometry.

PubMed

Dil, Ebrahim Alipanahpour; Ghaedi, Mehrorang; Ghezelbash, Gholam Reza; Asfaram, Arash

2017-05-01

Present study is based on application of live yeast Yarrowia lipolytica 70562 as new biosorbent was investigated for the simultaneous biosorption of Crystal Violet (CV) and Brilliant Green (BG) from wastewater. The effect of operating parameters such as initial dye concentrations (6-14mgL -1 ), solution pH (4.0-8.0) and contact time (4-20h) was investigated by response surface methodology (RSM) for modeling and optimization of biosorption process and accordingly the best operational conditions was set as: initial CV and BG concentration of 8.0, and 10mgL -1 , pH of 7.0 and contact time of 16h. Above specified conditions lead to achievement of maximum biosorption of 98.823% and 99.927% for CV and BG dyes, respectively. The experimental equilibrium data well explained according to Langmuir isotherm model with maximum biosorption capacity of 65.359 and 56.497mgg -1 for BG and CV, respectively. The second order and intraparticle diffusion models as cooperative mechanism has high efficiency and performance for interpretation of real data. Copyright © 2017. Published by Elsevier Inc.

Comparative evaluation of 13 yeast species in the Yarrowia clade on lignocellulosic biomass hydrolysate and genetic engineering of inhibitor tolerant strains for lipid and biofuel production

USDA-ARS?s Scientific Manuscript database

Yarrowia lipolytica is an oleaginous yeast that has garnered interest for commercial production of single cell oil and other fatty acid-derived chemicals because of its GRAS status and genetic tractability. Three recent peer-reviewed studies have highlighted the possibility of lipid production by th...

Quantitative study of lipase secretion, extracellular lipolysis, and lipid storage in the yeast Yarrowia lipolytica grown in the presence of olive oil: analogies with lipolysis in humans.

PubMed

Najjar, Amal; Robert, Sylvie; Guérin, Clémence; Violet-Asther, Michèle; Carrière, Frédéric

2011-03-01

Lipase secretion, extracellular lipolysis, and fatty acid uptake were quantified in the yeast Yarrowia lipolytica grown in the presence of olive oil and/or glucose. Specific lipase assays, Western blot analysis, and ELISA indicated that most of the lipase activity measured in Y. lipolytica cultures resulted from the YLLIP2 lipase. Lipase production was triggered by olive oil and, during the first hours of culture, most of the lipase activity and YLLIP2 immunodetection remained associated with the yeast cells. YLLIP2 was then released in the culture medium before it was totally degraded by proteases. Olive oil triglycerides were largely degraded when the lipase was still attached to the cell wall. The fate of lipolysis products in the culture medium and inside the yeast cell, as well as lipid storage, was investigated simultaneously by quantitative TLC-FID and GC analysis. The intracellular levels of free fatty acids (FFA) and triglycerides increased transiently and were dependent on the carbon sources. A maximum fat storage of 37.8% w/w of yeast dry mass was observed with olive oil alone. A transient accumulation of saturated FFA was observed whereas intracellular triglycerides became enriched in unsaturated fatty acids. So far, yeasts have been mainly used for studying the intracellular synthesis, storage, and mobilization of neutral lipids. The present study shows that yeasts are also interesting models for studying extracellular lipolysis and fat uptake by the cell. The quantitative data obtained here allow for the first time to establish interesting analogies with gastrointestinal and vascular lipolysis in humans.

Mutants of Yarrowia lipolytica NCIM 3589 grown on waste cooking oil as a biofactory for biodiesel production.

PubMed

Katre, Gouri; Ajmera, Namasvi; Zinjarde, Smita; RaviKumar, Ameeta

2017-10-24

Oleaginous yeasts are fast emerging as a possible feedstock for biodiesel production. Yarrowia lipolytica, a model oleaginous yeast is known to utilize a variety of hydrophobic substrates for lipid accumulation including waste cooking oil (WCO). Approaches to increase lipid content in this yeast include metabolic engineering which requires manipulation of multiple genes in the lipid biosynthesis pathway. A classical and cost-effective approach, namely, random chemical mutagenesis on the yeast can lead to increased production of biodiesel as is explored here. In this study, chemical mutagenesis using the alkylating agent, N- methyl-N'-nitro-N-nitrosoguanidine (MNNG) as well as an additional treatment with cerulenin, a fatty acid synthase inhibitor generated 800 mutants of Y. lipolytica NCIM 3589 (761 MNNG treated and 39 MNNG + cerulenin treated). A three-stage screening using Sudan Black B plate technique, Nile red fluorimetry and total lipid extraction using solvent was performed, which enabled selection of ten high lipid yielding mutants. Time course studies of all the ten mutants were further undertaken in terms of biomass, lipid yield and lipid content to select three stable mutants (YlB6, YlC7 and YlE1) capable of growing and accumulating lipid on WCO, with lipid contents of 55, 60 and 67% as compared to 45% for the wild type. The mutants demonstrated increased volumetric lipid productivities (0.062, 0.044 and 0.041 g L -1  h -1 ) as compared to the wild type (0.033 g L -1  h -1 ). The fatty acid profile of the three mutants consisted of a high content of C16 and C18 saturated and monounsaturated fatty acids and was found to be suitable for biodiesel production. The fuel properties, namely, density, kinematic viscosity, total acid number, iodine value of the three mutants were evaluated and found to lie within the limits specified by internationally accepted standards. Additionally, it was noted that the mutants demonstrated better cetane numbers and

Phenol Is the Initial Product Formed during Growth and Degradation of Bromobenzene by Tropical Marine Yeast, Yarrowia lipolytica NCIM 3589 via an Early Dehalogenation Step.

PubMed

Vatsal, Aakanksha A; Zinjarde, Smita S; RaviKumar, Ameeta

2017-01-01

Bromobenzene (BrB), a hydrophobic, recalcitrant organic compound, is listed by the environmental protection agencies as an environmental and marine pollutant having hepatotoxic, mutagenic, teratogenic, and carcinogenic effects. The tropical marine yeast Yarrowia lipolytica 3589 was seen to grow aerobically on BrB and displayed a maximum growth rate (μ max ) of 0.04 h -1 . Furthermore, we also observed an increase in cell size and sedimentation velocity for the cells grown on BrB as compared to the glucose grown cells. The cells attached to the hydrophobic bromobenzene droplets through its hydrophobic and acid-base interactions. The BrB (0.5%, 47.6 mM) was utilized by the cells with the release of a corresponding amount of bromide (12.87 mM) and yielded a cell mass of 1.86 g/L after showing 34% degradation in 96 h. Maximum dehalogenase activity of 16.16 U/mL was seen in the cell free supernatant after 24 h of growth. Identification of metabolites formed as a result of BrB degradation, namely, phenol, catechol, cis, cis muconic acid, and carbon dioxide were determined by LC-MS and GC-MS. The initial attack on bromobenzene by Y. lipolytica cells lead to the transient accumulation of phenol as an early intermediate which is being reported for the first time. Degradation of phenol led to catechol which was degraded by the ortho- cleavage pathway forming cis, cis muconic acid and then to Krebs cycle intermediates eventually leading to CO 2 production. The study shows that dehalogenation via an extracellular dehalogenase occurs prior to ring cleavage with phenol as the preliminary degradative compound being produced. The yeast was also able to grow on the degradative products, i.e., phenol and catechol, to varying degrees which would be of potential relevance in the degradation and remediation of xenobiotic environmental bromoaromatic pollutants such as bromobenzene.

Oleaginous yeast Yarrowia lipolytica culture with synthetic and food waste-derived volatile fatty acids for lipid production.

PubMed

Gao, Ruiling; Li, Zifu; Zhou, Xiaoqin; Cheng, Shikun; Zheng, Lei

2017-01-01

The sustainability of microbial lipids production from traditional carbon sources, such as glucose or glycerol, is problematic given the high price of raw materials. Considerable efforts have been directed to minimize the cost and find new alternative carbon sources. Volatile fatty acids (VFAs) are especially attractive raw materials, because they can be produced from a variety of organic wastes fermentation. Therefore, the use of volatile fatty acids as carbon sources seems to be a feasible strategy for cost-effective microbial lipid production. Lipid accumulation in Y. lipolytica using synthetic and food waste-derived VFAs as substrates was systematically compared and evaluated in batch cultures. The highest lipid content obtained with acetic, butyric, and propionic acids reached 31.62 ± 0.91, 28.36 ± 0.74, and 28.91 ± 0.66%, respectively. High concentrations of VFA inhibited cell growth in the following order: butyric acid > propionic acid > acetic acid. Within a 30-day experimental period, Y. lipolytica could adapt up to 20 g/L acetic acid, whereas the corresponding concentration of propionic acid and butyric acid were 10 and 5 g/L, respectively. Cultures on a VFA mixture showed that the utilization of different types of VFA by Y. lipolytica was not synchronized but rather performed in a step-wise manner. Although yeast fermentation is an exothermic process, and the addition of VFA will directly affect the pH of the system by increasing environmental acidity, cultures at a cultivation temperature of 38 °C and uncontrolled pH demonstrated that Y. lipolytica had high tolerance in the high temperature and acidic environment when a low concentration (2.5 g/L) of either synthetic or food waste-derived VFA was used. However, batch cultures fed with food fermentate yielded lower lipid content (18.23 ± 1.12%) and lipid productivity (0.12 ± 0.02 g/L/day). The lipid composition obtained with synthetic and food waste-derived VFA was similar to

A comparative analysis of single cell and droplet-based FACS for improving production phenotypes: Riboflavin overproduction in Yarrowia lipolytica.

PubMed

Wagner, James M; Liu, Leqian; Yuan, Shuo-Fu; Venkataraman, Maya V; Abate, Adam R; Alper, Hal S

2018-04-23

Evolutionary approaches to strain engineering inherently require the identification of suitable selection techniques for the product and phenotype of interest. In this work, we undertake a comparative analysis of two related but functionally distinct methods of high-throughput screening: traditional single cell fluorescence activated cell sorting (single cell FACS) and microdroplet-enabled FACS (droplet FACS) using water/oil/water (w/o/w) emulsions. To do so, we first engineer and evolve the non-conventional yeast Yarrowia lipolytica for high extracellular production of riboflavin (vitamin B2), an innately fluorescent product. Following mutagenesis and adaptive evolution, a direct parity-matched comparison of these two selection strategies was conducted. Both single cell FACS and droplet FACS led to significant increases in total riboflavin titer (32 and 54 fold relative to the parental PO1f strain, respectively). However, single cell FACS favored intracellular riboflavin accumulation (with only 70% of total riboflavin secreted) compared with droplet FACS that favored extracellular product accumulation (with 90% of total riboflavin secreted). We find that for the test case of riboflavin, the extent of secretion and total production were highly correlated. The resulting differences in production modes and levels clearly demonstrate the significant impact that selection approaches can exert on final evolutionary outcomes in strain engineering. Moreover, we note that these results provide a cautionary tale when intracellular read-outs of product concentration (including signals from biosensors) are used as surrogates for total production of potentially secreted products. In this regard, these results demonstrate that extracellular production is best assayed through an encapsulation technique when performing high throughput screening. Copyright © 2018 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

Phenol Is the Initial Product Formed during Growth and Degradation of Bromobenzene by Tropical Marine Yeast, Yarrowia lipolytica NCIM 3589 via an Early Dehalogenation Step

PubMed Central

Vatsal, Aakanksha A.; Zinjarde, Smita S.; RaviKumar, Ameeta

2017-01-01

Bromobenzene (BrB), a hydrophobic, recalcitrant organic compound, is listed by the environmental protection agencies as an environmental and marine pollutant having hepatotoxic, mutagenic, teratogenic, and carcinogenic effects. The tropical marine yeast Yarrowia lipolytica 3589 was seen to grow aerobically on BrB and displayed a maximum growth rate (μmax) of 0.04 h-1. Furthermore, we also observed an increase in cell size and sedimentation velocity for the cells grown on BrB as compared to the glucose grown cells. The cells attached to the hydrophobic bromobenzene droplets through its hydrophobic and acid–base interactions. The BrB (0.5%, 47.6 mM) was utilized by the cells with the release of a corresponding amount of bromide (12.87 mM) and yielded a cell mass of 1.86 g/L after showing 34% degradation in 96 h. Maximum dehalogenase activity of 16.16 U/mL was seen in the cell free supernatant after 24 h of growth. Identification of metabolites formed as a result of BrB degradation, namely, phenol, catechol, cis, cis muconic acid, and carbon dioxide were determined by LC–MS and GC–MS. The initial attack on bromobenzene by Y. lipolytica cells lead to the transient accumulation of phenol as an early intermediate which is being reported for the first time. Degradation of phenol led to catechol which was degraded by the ortho- cleavage pathway forming cis, cis muconic acid and then to Krebs cycle intermediates eventually leading to CO2 production. The study shows that dehalogenation via an extracellular dehalogenase occurs prior to ring cleavage with phenol as the preliminary degradative compound being produced. The yeast was also able to grow on the degradative products, i.e., phenol and catechol, to varying degrees which would be of potential relevance in the degradation and remediation of xenobiotic environmental bromoaromatic pollutants such as bromobenzene. PMID:28690604

Co-expression of Exo-inulinase and Endo-inulinase Genes in the Oleaginous Yeast Yarrowia lipolytica for Efficient Single Cell Oil Production from Inulin.

PubMed

Shi, Nianci; Mao, Weian; He, Xiaoxia; Chi, Zhe; Chi, Zhenming; Liu, Guanglei

2018-05-01

Yarrowia lipolytica is a promising platform for the single cell oil (SCO) production. In this study, a transformant X+N8 in which exo- and endo-inulinase genes were co-expressed could produce an inulinase activity of 124.33 U/mL within 72 h. However, the inulinase activity of a transformant X2 carrying a single exo-inulinase gene was only 47.33 U/mL within 72 h. Moreover, the transformant X+N8 could accumulate 48.13% (w/w) SCO from inulin and the cell dry weight reached 13.63 g/L within 78 h, which were significantly higher than those of the transformant X2 (41.87% (w/w) and 11.23 g/L) under the same conditions. In addition, inulin hydrolysis and utilization of the transformant X+N8 were also more efficient than those of the transformant X2 during the fermentation process. These results demonstrated that the co-expression of the exo- and endo-inulinase genes significantly enhanced the SCO production from inulin due to the improvement of the inulinase activity and the synergistic action of exo- and endo-inulinase. Besides, over 95.01% of the fatty acids from the transformant X+N8 were C16-C18, especially C18:1 (53.10%), suggesting that the fatty acids could be used as feedstock for biodiesel production.

Targeted mutations and MD simulations of a methanol-stable lipase YLIP9 from Yarrowia lipolytica MSR80 to develop a biodiesel enzyme.

PubMed

Syal, Poonam; Verma, Ved Vrat; Gupta, Rani

2017-11-01

Biodiesel, an environment friendly alternative for fuels, contains methyl esters of long-chain fatty acids. Our group has reported a methanol-stable YLIP9 from Yarrowia lipolytica MSR80 that shows poor catalysis of long-chain fatty acids. To shift its substrate specificity, residues within lid and binding pocket were identified for sequential mutations using YLIP2 as the template. Of the two point mutations (Glu116Leu and Ser119Val) introduced in the lid, the former mutation (YLIP9L1) increased the catalytic rate by ∼2-fold without any change in substrate specificity. In this mutant, six binding pocket residues (Bp2-Bp7) were further mutated to obtain six double mutants. YLIP9L1Bp3 showed significant shift in substrate specificity towards long-chain pNPesters with 11-fold increase in catalytic efficiency than YLIP9. Double mutations also led to increased thermostability and lowered activation energy of YLIP9L1Bp3 thereby shifting its optimum temperature from 60°C to 50°C. In silico molecular dynamics simulations revealed improved lid flexibility and increased catalytic triad volume in YLIP9L1Bp3. The enzyme YLIP9L1Bp3 was methanol-stable having selectivity for long-chain fatty acids with improved catalytic efficiency. Its application as a biodiesel enzyme was validated by transesterification of palm oil in presence of methanol, where it showed 8-fold increase in conversion of oil to methyl esters. Copyright © 2017 Elsevier B.V. All rights reserved.

Development of New Genetic Manipulation Tools for Metabolic Engineering of Diatoms

DTIC Science & Technology

2008-08-28

protein L41 has been shown in a variety of yeasts to be involved in resistance to anisomycin and cycloheximide 7𔄂. A conserved mutation from proline...atipitis Yarrowia lipolytica Candida troplcalis S. cerevisiae A. nidulans Oryza sativa Homo sapiens P. tricornutum T. pseudonana...Pichia stipitis Yarrowia lipolytica Candida troplcalis S. cerevisiae A. nidulans Oryza sativa Homo sapiens P

An interfacial and comparative in vitro study of gastrointestinal lipases and Yarrowia lipolytica LIP2 lipase, a candidate for enzyme replacement therapy.

PubMed

Bénarouche, Anaïs; Point, Vanessa; Carrière, Frédéric; Cavalier, Jean-François

2014-07-01

Lipolytic activities of Yarrowia lipolytica LIP2 lipase (YLLIP2), human pancreatic (HPL) and dog gastric (DGL) lipases were first compared using lecithin-stabilized triacylglycerol (TAG) emulsions (Intralipid) at various pH and bile salt concentrations. Like DGL, YLLIP2 was able to hydrolyze TAG droplets covered by a lecithin monolayer, while HPL was not directly active on that substrate. These results were in good agreement with the respective kinetics of adsorption on phosphatidylcholine (PC) monomolecular films of the same three lipases, YLLIP2 being the most tensioactive lipase. YLLIP2 adsorption onto a PC monolayer spread at the air/water interface was influenced by pH-dependent changes in the enzyme/lipid interfacial association constant (KAds) which was optimum at pH 6.0 on long-chain egg PC monolayer, and at pH 5.0 on medium chain dilauroylphosphatidylcholine film. Using substrate monolayers (1,2-dicaprin, trioctanoin), YLLIP2 displayed the highest lipolytic activities on both substrates in the 25-35 mN m(-1) surface pressure range. YLLIP2 was active in a large pH range and displayed a pH-dependent activity profile combining DGL and HPL features at pH values found in the stomach (pH 3-5) and in the intestine (pH 6-7), respectively. The apparent maximum activity of YLLIP2 was observed at acidic pH 4-6 and was therefore well correlated with an efficient interfacial binding at these pH levels, whatever the type of interfaces (Intralipid emulsions, substrate or PC monolayers). All these findings support the use of YLLIP2 in enzyme replacement therapy for the treatment of pancreatic exocrine insufficiency, a pathological situation in which an acidification of intestinal contents occurs. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Wei; Wei, Hui; Alahuhta, Markus

In order to develop a direct microbial sugar conversion platform for the production of lipids, drop-in fuels and chemicals from cellulosic biomass substrate, we chose Yarrowia lipolytica as a viable demonstration strain. Y. lipolytica is known to accumulate lipids intracellularly and is capable of metabolizing sugars to produce lipids; however, it lacks the lignocellulose-degrading enzymes needed to break down biomass directly. While research is continuing on the development of a Y. lipolytica strain able to degrade cellulose, in this study, we present successful expression of several xylanases in Y. lipolytica. The XynII and XlnD expressing Yarrowia strains exhibited an abilitymore » to grow on xylan mineral plates. This was shown by Congo Red staining of halo zones on xylan mineral plates. Enzymatic activity tests further demonstrated active expression of XynII and XlnD in Y. lipolytica. Furthermore, synergistic action in converting xylan to xylose was observed when XlnD acted in concert with XynII. Finally, the successful expression of these xylanases in Yarrowia further advances us toward our goal to develop a direct microbial conversion process using this organism.« less

Production of Eight Different Hydride Complexes and Nitrite Release from 2,4,6-Trinitrotoluene by Yarrowia lipolytica▿ †

PubMed Central

Ziganshin, Ayrat M.; Gerlach, Robin; Borch, Thomas; Naumov, Anatoly V.; Naumova, Rimma P.

2007-01-01

2,4,6-Trinitrotoluene (TNT) transformation by the yeast strain Yarrowia lipolytica AN-L15 was shown to occur via two different pathways. Direct aromatic ring reduction was the predominant mechanism of TNT transformation, while nitro group reduction was observed to be a minor pathway. Although growth of Y. lipolytica AN-L15 was inhibited initially in the presence of TNT, TNT transformation was observed, indicating that the enzymes necessary for TNT reduction were present initially. Aromatic ring reduction resulted in the transient accumulation of eight different TNT-hydride complexes, which were characterized using high-performance liquid chromatography, UV-visible diode array detection, and negative-mode atmospheric pressure chemical ionization mass spectrometry (APCI-MS). APCI-MS analysis revealed three different groups of TNT-hydride complexes with molecular ions at m/z 227, 228, and 230, which correspond to TNT-mono- and dihydride complexes and protonated dihydride isomers, respectively. One of the three protonated dihydride complex isomers detected appears to release nitrite in the presence of strain AN-L15. This release of nitrite is of particular interest since it can provide a pathway towards complete degradation and detoxification of TNT. PMID:17933928

The Gene YALI0E20207g from Yarrowia lipolytica Encodes an N-Acetylglucosamine Kinase Implicated in the Regulated Expression of the Genes from the N-Acetylglucosamine Assimilatory Pathway

PubMed Central

Flores, Carmen-Lisset; Gancedo, Carlos

2015-01-01

The non-conventional yeast Yarrowia lipolytica possesses an ORF, YALI0E20207g, which encodes a protein with an amino acid sequence similar to hexokinases from different organisms. We have cloned that gene and determined several enzymatic properties of its encoded protein showing that it is an N-acetylglucosamine (NAGA) kinase. This conclusion was supported by the lack of growth in NAGA of a strain carrying a YALI0E20207g deletion. We named this gene YlNAG5. Expression of YlNAG5 as well as that of the genes encoding the enzymes of the NAGA catabolic pathway—identified by a BLAST search—was induced by this sugar. Deletion of YlNAG5 rendered that expression independent of the presence of NAGA in the medium and reintroduction of the gene restored the inducibility, indicating that YlNag5 participates in the transcriptional regulation of the NAGA assimilatory pathway genes. Expression of YlNAG5 was increased during sporulation and homozygous Ylnag5/Ylnag5 diploid strains sporulated very poorly as compared with a wild type isogenic control strain pointing to a participation of the protein in the process. Overexpression of YlNAG5 allowed growth in glucose of an Ylhxk1glk1 double mutant and produced, in a wild type background, aberrant morphologies in different media. Expression of the gene in a Saccharomyces cerevisiae hxk1 hxk2 glk1 triple mutant restored ability to grow in glucose. PMID:25816199

Characterization of the two intracellular lipases of Y. lipolytica encoded by TGL3 and TGL4 genes: new insights into the role of intracellular lipases and lipid body organisation.

PubMed

Dulermo, Thierry; Tréton, Brigitte; Beopoulos, Athanasios; Kabran Gnankon, Affoué Philomène; Haddouche, Ramdane; Nicaud, Jean-Marc

2013-09-01

Eukaryotes store lipids in a specialised organelle, the lipid body (LB), mainly as triglycerides (TAGs). Both the rates of synthesis and degradation contribute to the control of the accumulation of TAGs. The synthesis of TAGs in yeasts has been well documented, especially in the model yeast Saccharomyces cerevisiae and in the oleaginous yeast Yarrowia lipolytica. However, descriptions of the processes involved in TAG degradation are more scarce and mostly for S. cerevisiae. Here, we report the characterisation of two Y. lipolytica genes, YlTGL3 and YlTGL4, encoding intracellular lipases involved in TAG degradation. The two proteins are localised in lipid bodies, and YlTgl4 was mainly found at the interface between LBs. Surprisingly, the spatial organisation of YlTgl3 and YlTgl4 depends on the culture medium and on the physiological phase of the cell. Inactivation of one or both genes doubles the lipid accumulation capacity of Y. lipolytica, increasing the cell's capacity to accumulate TAGs. The amino acid sequence of YlTgl4 contains the consensus sequence motif (G/A)XSXG, typical of serine hydrolases, whereas YlTgl3 does not. Single and double mutants are unable to degrade TAGs, and higher expression of YlTgl4 correlates with TAG degradation. Therefore, we propose that YlTgl4 is the main lipase responsible for TAG degradation and that YlTgl3 may act as a positive regulator of YlTgl4 rather than a functional lipase. Thus, contrary to S. cerevisiae, Y. lipolytica possesses two intracellular lipases with distinct roles and with distinct localisations in the LB. © 2013. Published by Elsevier B.V. All rights reserved.

Evaluation of the Composition of Culture Medium for Yeast Biomass Production Using Raw Glycerol from Biodiesel Synthesis

PubMed Central

dos Santos, Elisane Odriosolla; Michelon, Mariano; Furlong, Eliana Badiale; Burkert, Janaína Fernandes de Medeiros; Kalil, Susana Juliano; Burkert, Carlos André Veiga

2012-01-01

The work herewith investigated the production of yeast biomass as a source of protein, using Yarrowia lipolytica NRRL YB-423 and raw glycerol from biodiesel synthesis as the main carbon source. A significant influence of glycerol concentration, initial pH and yeast extract concentration on biomass and protein content was observed according to the 2v5-1 fractional design. These factors were further evaluated using a central composite design and response surface methodology, and an empirical model for protein content was established and validated. The biomass of Yarrowia lipolytica NRRL YB-423 reached 19.5 ± 1.0 g/L in shaken flasks cultivation, with a protein content of 20.1 ± 0.6% (w/w). PMID:24031849

21 CFR 173.165 - Candida lipolytica.

Code of Federal Regulations, 2010 CFR

2010-04-01

... description for Candida lipolytica variety lipolytica listed in “The Yeasts—A Toxonomic Study,” 2d Ed. (1970... equivalent). Activate as follows: Slurry 900 grams of silica gel reagent with 2 liters of purified water in a 3-liter beaker. Cool the mixture and pour into a 80 × 900 chromatographic column with coarse fritted...

21 CFR 173.165 - Candida lipolytica.

Code of Federal Regulations, 2011 CFR

2011-04-01

... description for Candida lipolytica variety lipolytica listed in “The Yeasts—A Toxonomic Study,” 2d Ed. (1970... equivalent). Activate as follows: Slurry 900 grams of silica gel reagent with 2 liters of purified water in a 3-liter beaker. Cool the mixture and pour into a 80 × 900 chromatographic column with coarse fritted...

Sustainable conversion of coffee and other crop wastes to biofuels and bioproducts using coupled biochemical and thermochemical processes in a multi-stage biorefinery concept.

PubMed

Hughes, Stephen R; López-Núñez, Juan Carlos; Jones, Marjorie A; Moser, Bryan R; Cox, Elby J; Lindquist, Mitch; Galindo-Leva, Luz Angela; Riaño-Herrera, Néstor M; Rodriguez-Valencia, Nelson; Gast, Fernando; Cedeño, David L; Tasaki, Ken; Brown, Robert C; Darzins, Al; Brunner, Lane

2014-10-01

The environmental impact of agricultural waste from the processing of food and feed crops is an increasing concern worldwide. Concerted efforts are underway to develop sustainable practices for the disposal of residues from the processing of such crops as coffee, sugarcane, or corn. Coffee is crucial to the economies of many countries because its cultivation, processing, trading, and marketing provide employment for millions of people. In coffee-producing countries, improved technology for treatment of the significant amounts of coffee waste is critical to prevent ecological damage. This mini-review discusses a multi-stage biorefinery concept with the potential to convert waste produced at crop processing operations, such as coffee pulping stations, to valuable biofuels and bioproducts using biochemical and thermochemical conversion technologies. The initial bioconversion stage uses a mutant Kluyveromyces marxianus yeast strain to produce bioethanol from sugars. The resulting sugar-depleted solids (mostly protein) can be used in a second stage by the oleaginous yeast Yarrowia lipolytica to produce bio-based ammonia for fertilizer and are further degraded by Y. lipolytica proteases to peptides and free amino acids for animal feed. The lignocellulosic fraction can be ground and treated to release sugars for fermentation in a third stage by a recombinant cellulosic Saccharomyces cerevisiae, which can also be engineered to express valuable peptide products. The residual protein and lignin solids can be jet cooked and passed to a fourth-stage fermenter where Rhodotorula glutinis converts methane into isoprenoid intermediates. The residues can be combined and transferred into pyrocracking and hydroformylation reactions to convert ammonia, protein, isoprenes, lignins, and oils into renewable gas. Any remaining waste can be thermoconverted to biochar as a humus soil enhancer. The integration of multiple technologies for treatment of coffee waste has the potential to

Origins of cell-to-cell bioprocessing diversity and implications of the extracellular environment revealed at the single-cell level

DOE PAGES

Vasdekis, A. E.; Silverman, A. M.; Stephanopoulos, G.

2015-12-14

We probed the lipid expression dynamics of the oleaginous yeast Yarrowia Lipolytica. We observed that neutral lipid expression is sporadic. By performing single-cell analysis, we found that such noise emanates from the metabolic reaction level. Our results provide an alternative insight into the regulation and phenotypic variability of lipogenesis.

Complementation and Genetic Recombination in Candida lipolytica

PubMed Central

Bassel, John; Warfei, Jean; Mortimer, Robert

1971-01-01

Nutritional requirements were introduced into wild-type, heterothallic strains of Candida lipolytica by exposing the cells to X rays. Complementing hybrids were recovered from mixtures of the auxotrophic strains, and genetic recombination was observed in individually isolated ascospores from the hybrid strains. PMID:5122814

Adhesion of Bacillus subtilis and Pseudoalteromonas lipolytica to steel in a seawater environment and their effects on corrosion.

PubMed

Guo, Zhangwei; Liu, Tao; Cheng, Y Frank; Guo, Na; Yin, Yansheng

2017-09-01

In a marine environment, Bacillus subtilis and Pseudoalteromonas lipolytica are commonly found in the biofilms adherent to low-alloy engineering steel, and they have distinct effects on corrosion. In the present work, this phenomenon was investigated through the study of various materials characterization methods, electrochemical techniques, and contact angle measurements. It was found that the surface film formed on the steel in the presence of B. subtilis was compact, uniform, free of cracks, and hydrophobic. However, the film formed in the presence of P. lipolytica was loose, rough, heterogeneous, and hydrophilic. The main components of the films formed in the presence of B. subtilis and P. lipolytica were polysaccharides/TasA amyloid fibers and proteins/carboxylic acid, respectively. The composition, structure, and properties of the surface films formed on the steel were associated with different effects on corrosion. The presence of B. subtilis enhances the steel's resistance to corrosion, whereas corrosion was increased by the presence of P. lipolytica. In short, the compact and hydrophobic biofilm of B. subtilis appears to inhibit the corrosion of steel, while the loose, hydrophilic film of P. lipolytica tends to induce pitting corrosion. Copyright © 2017 Elsevier B.V. All rights reserved.

Occurrence and diversity of marine yeasts in Antarctica environments

NASA Astrophysics Data System (ADS)

Zhang, Xue; Hua, Mingxia; Song, Chunli; Chi, Zhenming

2012-03-01

A total of 28 yeast strains were obtained from the sea sediment of Antarctica. According to the results of routine identification and molecular characterization, the strains belonged to species of Yarrowia lipolytica, Debaryomyces hansenii, Rhodotorula slooffiae, Rhodotorula mucilaginosa, Sporidiobolus salmonicolor, Aureobasidium pullulans, Mrakia frigida and Guehomyces pullulans, respectively. The Antarctica yeasts have wide potential applications in biotechnology, for some of them can produce β-galactosidase and killer toxins.

Clinical, microbiological, and experimental animal studies of Candida lipolytica.

PubMed Central

Walsh, T J; Salkin, I F; Dixon, D M; Hurd, N J

1989-01-01

Candida lipolytica was recovered from six patients in three different clinical centers. The index isolate caused a persistent fungemia with catheter-associated Candida thrombophlebitis, the second isolate was from a polymicrobial sinusitis, and the remaining four isolates were involved in tissue colonization. These and 20 other isolates were consistent in their morphological and physiological characteristics. All formed true hyphae and blastoconidia on cornmeal-Tween 80 agar and all assimilated glucose, glycerol, and erythritol. In a murine model of disseminated candidiasis, the index isolate that caused clinical fungemia caused no mortality and produced only two lesions on a kidney, as determined at necropsy. The nine isolates selected for in vitro antifungal susceptibility studies had intermediate susceptibilities to amphotericin B but were susceptible to ketoconazole. We conclude that C. lipolytica is a weakly virulent pathogen which may require an intravascular foreign body to cause fungemia. Images PMID:2745702

The growth, properties and interactions of yeasts and bacteria associated with the maturation of Camembert and blue-veined cheeses.

PubMed

Addis, E; Fleet, G H; Cox, J M; Kolak, D; Leung, T

2001-09-19

The growth of yeasts and bacteria were monitored during the maturation of Camembert and blue-veined cheese produced in Australia. Yeasts were prominent throughout maturation, growing to 10(5)-10(9)/g, depending on the manufacturer. Debaryomyces hansenii predominated, but there were lesser, inconsistent contributions from Yarrowia lipolytica. Of the non-lactic acid bacteria, Acinetobacter species were significant during the maturation of Camembert but not blue-veined cheeses, and grew to 10(6)-10(8) cfu/g. Staphylococcus and Micrococcus species were consistently isolated from the cheeses with Staphylococcus xylosus growing to 10(5)-10(9) cfu/g, depending on the product. Lactic acid bacteria (10(7)-10(9) cfu/g) were present throughout maturation but were not identified. Interactions between the various yeasts and bacterial isolates were examined. Several strains of D. hansenii exhibited killer activity but not against Y. lipolytica. None of the yeasts were antagonistic towards the bacteria but some strains of D. hansenii enhanced the growth of Y. lipolytica and S. xylosus. The yeast and bacterial isolates exhibited various degrees of extracellular proteolytic and lipolytic activities.

Influence of selected factors on browning of Camembert cheese.

PubMed

Carreira, Alexandra; Dillinger, Klaus; Eliskases-Lechner, Frieda; Loureiro, Virgílio; Ginzinger, Wolfgang; Rohm, Harald

2002-05-01

Experimental Camembert cheeses were made to investigate the effects on browning of the following factors: inoculation with Yarrowia lipolytica, the use of Penicillium candidum strains with different proteolytic activity, the addition of tyrosine, and the addition of Mn2+ thus leading to 16 different variants of cheese. Two physical colour parameters were used to describe browning, depending on the location in the cheeses: a whiteness index for the outside browning (mould mycelium), and a brownness index for the inside browning (surface of the cheese body). Mn2+ promoted a significant increase of browning at both locations, whereas Yar. lipolytica had the opposite effect. Outside browning was significantly more intense when using the Pen. candidum strain with higher proteolytic activity. A significant interaction was found between Yar. lipolytica and Pen. candidum. The yeast had no effect in combination with a low proteolytic strain of Pen. candidum, but significantly reduced proteolysis and browning in combination with a high proteolytic strain of Pen. candidum. We further confirmed that both strains of Pen. candidum were able to produce brown pigments from tyrosine and thus both are presumably responsible for the browning activity in this type of cheese.

Metabolic Engineering of Oleaginous Yeasts for Fatty Alcohol Production

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Wei; Wei, Hui; Knoshaug, Eric

To develop pathways for advanced biological upgrading of sugars to hydrocarbons, we are seeking biological approaches to produce high carbon efficiency intermediates amenable to separations and catalytic upgrading to hydrocarbon fuels. In this study, we successfully demonstrated fatty alcohol production by oleaginous yeasts Yarrowia lipolytica and Lipomyces starkeyi by expressing a bacteria-derived fatty acyl-CoA reductase (FAR). Moreover, we find higher extracellular distribution of fatty alcohols produced by FAR-expressing L. starkeyi strain as compared to Y. lipolytica strain, which would benefit the downstream product recovery process. In both oleaginous yeasts, long chain length saturated fatty alcohols were predominant, accounting for moremore » than 85% of the total fatty alcohols produced. To the best of our knowledge, this is the first report of fatty alcohol production in L. starkeyi. Taken together, our work demonstrates that in addition to Y. lipolytica, L. starkeyi can also serve as a platform organism for production of fatty acid-derived biofuels and bioproducts via metabolic engineering. We believe strain and process development both will significantly contribute to our goal of producing scalable and cost-effective fatty alcohols from renewable biomass.« less

Evolutionary relationships among pathogenic Candida species and relatives.

PubMed Central

Barns, S M; Lane, D J; Sogin, M L; Bibeau, C; Weisburg, W G

1991-01-01

Small subunit rRNA sequences have been determined for 10 of the most clinically important pathogenic species of the yeast genus Candida (including Torulopsis [Candida] glabrata and Yarrowia [Candida] lipolytica) and for Hansenula polymorpha. Phylogenetic analyses of these sequences and those of Saccharomyces cerevisiae, Kluyveromyces marxianus var. lactis, and Aspergillus fumigatus indicate that Candida albicans, C. tropicalis, C. parapsilosis, and C. viswanathii form a subgroup within the genus. The remaining significant pathogen, T. glabrata, falls into a second, distinct subgroup and is specifically related to S. cerevisiae and more distantly related to C. kefyr (psuedotropicalis) and K. marxianus var. lactis. The 18S rRNA sequence of Y. lipolytica has evolved rapidly in relation to the other Candida sequences examined and appears to be only distantly related to them. As anticipated, species of several other genera appear to bear specific relationships to members of the genus Candida. PMID:2007550

Metrological aspects of enzyme production

NASA Astrophysics Data System (ADS)

Kerber, T. M.; Dellamora-Ortiz, G. M.; Pereira-Meirelles, F. V.

2010-05-01

Enzymes are frequently used in biotechnology to carry out specific biological reactions, either in industrial processes or for the production of bioproducts and drugs. Microbial lipases are an important group of biotechnologically valuable enzymes that present widely diversified applications. Lipase production by microorganisms is described in several published papers; however, none of them refer to metrological evaluation and the estimation of the uncertainty in measurement. Moreover, few of them refer to process optimization through experimental design. The objectives of this work were to enhance lipase production in shaken-flasks with Yarrowia lipolytica cells employing experimental design and to evaluate the uncertainty in measurement of lipase activity. The highest lipolytic activity obtained was about three- and fivefold higher than the reported activities of CRMs BCR-693 and BCR-694, respectively. Lipase production by Y. lipolytica cells aiming the classification as certified reference material is recommended after further purification and stability studies.

High-oleate yeast oil without polyunsaturated fatty acids.

PubMed

Tsakraklides, Vasiliki; Kamineni, Annapurna; Consiglio, Andrew L; MacEwen, Kyle; Friedlander, Jonathan; Blitzblau, Hannah G; Hamilton, Maureen A; Crabtree, Donald V; Su, Austin; Afshar, Jonathan; Sullivan, John E; LaTouf, W Greg; South, Colin R; Greenhagen, Emily H; Shaw, A Joe; Brevnova, Elena E

2018-01-01

Oleate-enriched triacylglycerides are well-suited for lubricant applications that require high oxidative stability. Fatty acid carbon chain length and degree of desaturation are key determinants of triacylglyceride properties and the ability to manipulate fatty acid composition in living organisms is critical to developing a source of bio-based oil tailored to meet specific application requirements. We sought to engineer the oleaginous yeast Yarrowia lipolytica for production of high-oleate triacylglyceride oil. We studied the effect of deletions and overexpressions in the fatty acid and triacylglyceride synthesis pathways to identify modifications that increase oleate levels. Oleic acid accumulation in triacylglycerides was promoted by exchanging the native ∆9 fatty acid desaturase and glycerol-3-phosphate acyltransferase with heterologous enzymes, as well as deletion of the Δ12 fatty acid desaturase and expression of a fatty acid elongase. By combining these engineering steps, we eliminated polyunsaturated fatty acids and created a Y. lipolytica strain that accumulates triglycerides with > 90% oleate content. High-oleate content and lack of polyunsaturates distinguish this triacylglyceride oil from plant and algal derived oils. Its composition renders the oil suitable for applications that require high oxidative stability and further demonstrates the potential of Y. lipolytica as a producer of tailored lipid profiles.

Yeast mitochondrial HMG proteins: DNA-binding properties of the most evolutionarily divergent component of mitochondrial nucleoids.

PubMed

Bakkaiova, Jana; Marini, Victoria; Willcox, Smaranda; Nosek, Jozef; Griffith, Jack D; Krejci, Lumir; Tomaska, Lubomir

2015-12-08

Yeast mtDNA is compacted into nucleoprotein structures called mitochondrial nucleoids (mt-nucleoids). The principal mediators of nucleoid formation are mitochondrial high-mobility group (HMG)-box containing (mtHMG) proteins. Although these proteins are some of the fastest evolving components of mt-nucleoids, it is not known whether the divergence of mtHMG proteins on the level of their amino acid sequences is accompanied by diversification of their biochemical properties. In the present study we performed a comparative biochemical analysis of yeast mtHMG proteins from Saccharomyces cerevisiae (ScAbf2p), Yarrowia lipolytica (YlMhb1p) and Candida parapsilosis (CpGcf1p). We found that all three proteins exhibit relatively weak binding to intact dsDNA. In fact, ScAbf2p and YlMhb1p bind quantitatively to this substrate only at very high protein to DNA ratios and CpGcf1p shows only negligible binding to dsDNA. In contrast, the proteins exhibit much higher preference for recombination intermediates such as Holliday junctions (HJ) and replication forks (RF). Therefore, we hypothesize that the roles of the yeast mtHMG proteins in maintenance and compaction of mtDNA in vivo are in large part mediated by their binding to recombination/replication intermediates. We also speculate that the distinct biochemical properties of CpGcf1p may represent one of the prerequisites for frequent evolutionary tinkering with the form of the mitochondrial genome in the CTG-clade of hemiascomycetous yeast species. © 2016 Authors.

Improvement of soil characteristics and growth of Dorycnium pentaphyllum by amendment with agrowastes and inoculation with AM fungi and/or the yeast Yarowia lipolytica.

PubMed

Medina, A; Vassileva, M; Caravaca, F; Roldán, A; Azcón, R

2004-08-01

The effectiveness of two microbiologically treated agrowastes [dry olive cake (DOC) and/or sugar beet (SB)] on plant growth, soil enzymatic activities and other soil characteristics was determined in a natural soil from a desertified area. Dorycnium pentaphyllum, a legume plant adapted to stress situations, was the test plant to evaluate the effect of inoculation of native arbuscular mycorrhizal (AM) fungi and/or Yarowia lipolytica (a dry soil adapted yeast) on amended and non-amended soils. Plant growth and nutrition, symbiotic developments and soil enzymatic activities were limited in non-amended soil where microbial inoculations did not improve plant development. The lack of nodules formation and AM colonization can explain the limited plant growth in this natural soil. The effectiveness and performance of inocula applied was only evident in amended soils. AM colonization and spores number in natural soil were increased by amendments and the inoculation with Y. lipolytica promoted this value. The effect of the inoculations on plant N-acquisition was only important in AM-inoculated plants growing in SB medium. Enzymatic activities as urease and protease activities were particularly increased in DOC amended soil meanwhile dehydrogenase activity was greatest in treatments inoculated with Y. lipolytica in SB added soil. The biological activities in rhizosphere of agrowaste amended soil, used as indices of changes in soil properties and fertility, were affected not only by the nature of amendments but also by the inoculant applied. All these results show that the lignocellulosic agrowastes treated with a selected microorganism and its further interaction with beneficial microbial groups (native AM fungi and/or Y. lipolytica) is a useful tool to modify soil physico-chemical, biological and fertility properties that enhance the plant performance probably by making nutrients more available to plants.

Screening of a thiamine-auxotrophic yeast for alpha-ketoglutaric acid overproduction.

PubMed

Zhou, Jingwen; Zhou, Haiyan; Du, Guocheng; Liu, Liming; Chen, Jian

2010-09-01

To obtain a thiamine-auxotrophic yeast strain that overproduces alpha-ketoglutaric acid (alpha-KG) from glycerol and to investigate nutrient effects on alpha-KG production. Yeast strain WSH-Z06, a thiamine auxotroph that gave high yields of alpha-KG from glycerol, was obtained by screening for ampicillin/kanamycin resistance and thiamine auxotrophy. The strain was identified as Yarrowia lipolytica based on physiological, chemical, and phylogenetic analysis. The ability of the strain to convert glycerol to alpha-KG was analysed by investigating the effects of nutritional factors, including thiamine, riboflavin, nitrogen sources, and calcium ion. Thiamine and calcium ion concentration had the greatest effect on alpha-KG accumulation. Under optimal conditions, a yield of 39.2 g l(-1)alpha-KG was obtained from 100 g l(-1) glycerol, with 16.84 g l(-1) pyruvate as a by-product. The current work provides a method for screening for an alpha-KG overproducer. Nutrients have a significant impact on alpha-KG production in the yeast strain presented here. The alpha-KG-overproducing yeast strain Y. lipolytica WSH-Z06 is a promising parent strain for further metabolic engineering to lower by-product accumulation and accelerate glycerol utilization.

Physiological uncoupling of mitochondrial oxidative phosphorylation. Studies in different yeast species.

PubMed

Guerrero-Castillo, Sergio; Araiza-Olivera, Daniela; Cabrera-Orefice, Alfredo; Espinasa-Jaramillo, Juan; Gutiérrez-Aguilar, Manuel; Luévano-Martínez, Luís A; Zepeda-Bastida, Armando; Uribe-Carvajal, Salvador

2011-06-01

Under non-phosphorylating conditions a high proton transmembrane gradient inhibits the rate of oxygen consumption mediated by the mitochondrial respiratory chain (state IV). Slow electron transit leads to production of reactive oxygen species (ROS) capable of participating in deleterious side reactions. In order to avoid overproducing ROS, mitochondria maintain a high rate of O(2) consumption by activating different exquisitely controlled uncoupling pathways. Different yeast species possess one or more uncoupling systems that work through one of two possible mechanisms: i) Proton sinks and ii) Non-pumping redox enzymes. Proton sinks are exemplified by mitochondrial unspecific channels (MUC) and by uncoupling proteins (UCP). Saccharomyces. cerevisiae and Debaryomyces hansenii express highly regulated MUCs. Also, a UCP was described in Yarrowia lipolytica which promotes uncoupled O(2) consumption. Non-pumping alternative oxido-reductases may substitute for a pump, as in S. cerevisiae or may coexist with a complete set of pumps as in the branched respiratory chains from Y. lipolytica or D. hansenii. In addition, pumps may suffer intrinsic uncoupling (slipping). Promising models for study are unicellular parasites which can turn off their aerobic metabolism completely. The variety of energy dissipating systems in eukaryote species is probably designed to control ROS production in the different environments where each species lives.

Evolution of the carboxylate Jen transporters in fungi.

PubMed

Lodi, Tiziana; Diffels, Julie; Goffeau, André; Baret, Philippe V

2007-08-01

Synteny analysis is combined with sequence similarity and motif identification to trace the evolution of the putative monocarboxylate (lactate/pyruvate) transporters Jen1p and the dicarboxylate (succinate/fumarate/malate) transporters Jen2p in Hemiascomycetes yeasts and Euascomycetes fungi. It is concluded that a precursor form of Jen1p, named here preJen1p, arose by the duplication of an ancestral Jen2p, during the speciation of Yarrowia lipolytica, which was transferred into a new syntenic context. The Jen1p transporters differentiated from preJen1p in Kluyveromyces lactis, before the Whole Genome Duplication (WGD), and are conserved as a single copy in the Saccharomyces species. In contrast, the ancestral Jen2p was definitively lost just prior to the WGD and is absent in Saccharomyces.

Biodegradation of micropollutant naproxen with a selected fungal strain and identification of metabolites.

PubMed

Aracagök, Y Doruk; Göker, Hakan; Cihangir, Nilüfer

2017-05-01

Pharmaceuticals are widely used for treating human and animal diseases. Naproxen [(S) 6-methoxy-α-methyl-2-naphthalene acetic acid] and its sodium salt are members of the α-arylpropionic acid group of nonsteroidal anti-inflammatory drugs. Due to excessive usage of naproxen, this drug has been determined even in drinking water. In this study, four fungal strains Phanerochaete chrysosporium, Funalia trogii, Aspergillus niger, and Yarrowia lipolytica were investigated in terms of naproxen removal abilities. According to LC/MS data, A. niger was found the most efficient strain with 98% removal rate. Two main by-products of fungal transformation, O-desmethylnaproxen and 7-hydroxynaproxen, were identified by using LC/MS, 1HNMR, and 13CNMR. Our results showed that O-demethylation and hydroxylation of naproxen is catalyzed by cytochrome P450 enzyme system.

Molecular and biochemical characterisation of two aspartic proteinases TcAP1 and TcAP2 from Theobroma cacao seeds.

PubMed

Laloi, Maryse; McCarthy, James; Morandi, Olivia; Gysler, Christof; Bucheli, Peter

2002-09-01

Aspartic proteinase (EC 3.4.23) activity plays a pivotal role in the degradation of Theobroma cacao L. seed proteins during the fermentation step of cacao bean processing. Therefore, this enzyme is believed to be critical for the formation of the peptide and amino acid cocoa flavor precursors that occurs during fermentation. Using cDNA cloning and northern blot analysis, we show here that there are at least two distinct aspartic proteinase genes ( TcAP1 and TcAP2) expressed during cacao seed development. Both genes are expressed early during seed development and their mRNA levels decrease towards the end of seed maturation. TcAP2 is expressed at a much higher level than TcAP1, although the expression of TcAP1 increases slightly during germination. The proteins encoded by TcAP1 and TcAP2 are relatively different from each other (73% identity). This, and the fact that the two corresponding genes have different expression patterns, suggests that the TcAP1 and TcAP2 proteins may have different functions in the maturing seeds and during germination. Because the TcAP2 gene is expressed at a much higher level during seed development than TcAP1, it is likely that the TcAP2 protein is primarily responsible for the majority of the industrially important protein hydrolysis that occurs during cacao bean fermentation. Finally, TcAP2 has been functionally expressed in the yeast Yarrowia lipolytica. The secreted recombinant protein is able to hydrolyse bovine haemoglobin at acidic pH and is sensitive to pepstatin A, confirming that TcAP2 encodes an aspartic proteinase, and strongly suggests that this gene encodes the well-characterized aspartic proteinase of mature cacao seeds.

Investigation of the effect of biologically active threo-Ds-isocitric acid on oxidative stress in Paramecium caudatum.

PubMed

Morgunov, Igor G; Karpukhina, Olga V; Kamzolova, Svetlana V; Samoilenko, Vladimir A; Inozemtsev, Anatoly N

2018-01-02

The effect of biologically active form (threo-Ds-) of isocitric acid (ICA) on oxidative stress was studied using the infusorian Paramecium caudatum stressed by hydrogen peroxide and salts of some heavy metals (Cu, Pb, Zn, and Cd). ICA at concentrations between 0.5 and 10 mM favorably influenced the infusorian cells with oxidative stress induced by the toxicants studied. The maximal antioxidant effect of ICA was observed at its concentration 10 mM irrespective of the toxicant used (either H 2 O 2 or heavy metal ions). ICA was found to be a more active antioxidant than ascorbic acid. Biologically active pharmaceutically pure threo-Ds-ICA was produced through cultivation of the yeast Yarrowia lipolytica and isolated from the culture liquid in the form of crystalline monopotassium salt with a purity of 99.9%.

Antifungal activity of the lipopeptides produced by Bacillus amyloliquefaciens anti-CA against Candida albicans isolated from clinic.

PubMed

Song, Bo; Rong, Yan-Jun; Zhao, Ming-Xin; Chi, Zhen-Ming

2013-08-01

The bacterium Bacillus amyloliquefaciens anti-CA isolated from mangrove system was found to be able to actively kill Candida albicans isolated from clinic. The bacterial strain anti-CA could produce high level of bioactive substance, amylase and protease in the cheap medium containing 2.0 % soybean meal, 2.0 % wheat flour, pH 6.5 within 26 h. After purification, the main bioactive substance was confirmed to be a cyclic lipopeptide containing a heptapeptide, L-Asp→L-Leu→L-Leu→L-Val→L-Val→L-Glu→L-Leu and a 3-OH fatty acid (15 carbons). In addition to C. albicans, the purified lipopeptide can also kill many yeast strains including Metschnikowia bicuspidata, Candida tropicalis, Yarrowia lipolytica and Saccharomyces cerevisiae. After treated by the purified lipopeptide, both the whole cells and protoplasts of C. albicans were destroyed.

Metabolic engineering of microbial competitive advantage for industrial fermentation processes.

PubMed

Shaw, A Joe; Lam, Felix H; Hamilton, Maureen; Consiglio, Andrew; MacEwen, Kyle; Brevnova, Elena E; Greenhagen, Emily; LaTouf, W Greg; South, Colin R; van Dijken, Hans; Stephanopoulos, Gregory

2016-08-05

Microbial contamination is an obstacle to widespread production of advanced biofuels and chemicals. Current practices such as process sterilization or antibiotic dosage carry excess costs or encourage the development of antibiotic resistance. We engineered Escherichia coli to assimilate melamine, a xenobiotic compound containing nitrogen. After adaptive laboratory evolution to improve pathway efficiency, the engineered strain rapidly outcompeted a control strain when melamine was supplied as the nitrogen source. We additionally engineered the yeasts Saccharomyces cerevisiae and Yarrowia lipolytica to assimilate nitrogen from cyanamide and phosphorus from potassium phosphite, and they outcompeted contaminating strains in several low-cost feedstocks. Supplying essential growth nutrients through xenobiotic or ecologically rare chemicals provides microbial competitive advantage with minimal external risks, given that engineered biocatalysts only have improved fitness within the customized fermentation environment. Copyright © 2016, American Association for the Advancement of Science.

Zinc Finger Transcription Factors Displaced SREBP Proteins as the Major Sterol Regulators during Saccharomycotina Evolution

PubMed Central

Maguire, Sarah L.; Wang, Can; Holland, Linda M.; Brunel, François; Neuvéglise, Cécile; Nicaud, Jean-Marc; Zavrel, Martin; White, Theodore C.; Wolfe, Kenneth H.; Butler, Geraldine

2014-01-01

In most eukaryotes, including the majority of fungi, expression of sterol biosynthesis genes is regulated by Sterol-Regulatory Element Binding Proteins (SREBPs), which are basic helix-loop-helix transcription activators. However, in yeasts such as Saccharomyces cerevisiae and Candida albicans sterol synthesis is instead regulated by Upc2, an unrelated transcription factor with a Gal4-type zinc finger. The SREBPs in S. cerevisiae (Hms1) and C. albicans (Cph2) have lost a domain, are not major regulators of sterol synthesis, and instead regulate filamentous growth. We report here that rewiring of the sterol regulon, with Upc2 taking over from SREBP, likely occurred in the common ancestor of all Saccharomycotina. Yarrowia lipolytica, a deep-branching species, is the only genome known to contain intact and full-length orthologs of both SREBP (Sre1) and Upc2. Deleting YlUPC2, but not YlSRE1, confers susceptibility to azole drugs. Sterol levels are significantly reduced in the YlUPC2 deletion. RNA-seq analysis shows that hypoxic regulation of sterol synthesis genes in Y. lipolytica is predominantly mediated by Upc2. However, YlSre1 still retains a role in hypoxic regulation; growth of Y. lipolytica in hypoxic conditions is reduced in a Ylupc2 deletion and is abolished in a Ylsre1/Ylupc2 double deletion, and YlSre1 regulates sterol gene expression during hypoxia adaptation. We show that YlSRE1, and to a lesser extent YlUPC2, are required for switching from yeast to filamentous growth in hypoxia. Sre1 appears to have an ancestral role in the regulation of filamentation, which became decoupled from its role in sterol gene regulation by the arrival of Upc2 in the Saccharomycotina. PMID:24453983

Presence and changes in populations of yeasts on raw and processed poultry products stored at refrigeration temperature.

PubMed

Ismail, S A; Deak, T; El-Rahman, H A; Yassien, M A; Beuchat, L R

2000-12-05

A study was undertaken to determine populations and profiles of yeast species on fresh and processed poultry products upon purchase from retail supermarkets and after storage at 5 degrees C until shelf life expiration, and to assess the potential role of these yeasts in product spoilage. Fifty samples representing 15 commercial raw, marinated, smoked, or roasted chicken and turkey products were analyzed. Yeast populations were determined by plating on dichloran rose bengal chloramphenicol (DRBC) agar and tryptone glucose yeast extract (TGY) agar. Proteolytic activity was determined using caseinate and gelatin agars and lipolytic activity was determined on plate count agar supplemented with tributyrin. Populations of aerobic microorganisms were also determined. Initial populations of yeasts (log10 cfu/g) ranged from less than 1 (detection limit) to 2.89, and increased by the expiration date to 0.37-5.06, indicating the presence of psychrotrophic species. Highest initial populations were detected in raw chicken breast, wings, and ground chicken, as well as in turkey necks and legs, whereas roasted chicken and turkey products contained less than 1 log10 cfu/g. During storage, yeast populations increased significantly (P < or = 0.05) in whole chicken, ground chicken, liver, heart and gizzard, and in ground turkey and turkey sausage. Isolates (152 strains) of yeasts from poultry products consisted of 12 species. Yarrowia lipolytica and Candida zeylanoides were predominant, making up 39 and 26% of the isolates, respectively. Six different species of basidiomycetous yeasts representing 24% of the isolates were identified. Most Y. lipolytica strains showed strong proteolytic and lipolytic activities, whereas C. zeylanoides was weakly lipolytic. Results suggest that yeasts, particularly Y. lipolytica, may play a more prominent role than previously recognized in the spoilage of fresh and processed poultry stored at 5 degrees C.

Interactions between yeasts and bacteria in the smear surface-ripened cheeses.

PubMed

Corsetti, A; Rossi, J; Gobbetti, M

2001-09-19

In the initial phase of ripening, the microflora of bacterial smear surface-ripened cheeses such as Limburger, Taleggio, Brick, Münster and Saint-Paulin and that of surface mould-ripened cheeses such as Camembert and Brie may be similar, but at the end of the ripening, bacteria such as Brevibacterium spp., Arthrobacter spp., Micrococcus spp., Corynebacterium spp. and moulds such as Penicillium camemberti are, respectively, the dominant microorganisms. Yeasts such as Candida spp., Cryptococcus spp., Debaryomyces spp., Geotrichum candidum, Pichia spp., Rhodotorula spp., Saccharomyces spp. and Yarrowia lipolytica are often and variably isolated from the smear surface-ripened cheeses. Although not dominant within the microorganisms of the smear surface-ripened cheeses, yeasts establish significant interactions with moulds and especially bacteria, including surface bacteria and lactic acid bacteria. Some aspects of the interactions between yeasts and bacteria in such type of cheeses are considered in this paper.

Human recombinant lysosomal enzymes produced in microorganisms.

PubMed

Espejo-Mojica, Ángela J; Alméciga-Díaz, Carlos J; Rodríguez, Alexander; Mosquera, Ángela; Díaz, Dennis; Beltrán, Laura; Díaz, Sergio; Pimentel, Natalia; Moreno, Jefferson; Sánchez, Jhonnathan; Sánchez, Oscar F; Córdoba, Henry; Poutou-Piñales, Raúl A; Barrera, Luis A

2015-01-01

Lysosomal storage diseases (LSDs) are caused by accumulation of partially degraded substrates within the lysosome, as a result of a function loss of a lysosomal protein. Recombinant lysosomal proteins are usually produced in mammalian cells, based on their capacity to carry out post-translational modifications similar to those observed in human native proteins. However, during the last years, a growing number of studies have shown the possibility to produce active forms of lysosomal proteins in other expression systems, such as plants and microorganisms. In this paper, we review the production and characterization of human lysosomal proteins, deficient in several LSDs, which have been produced in microorganisms. For this purpose, Escherichia coli, Saccharomyces cerevisiae, Pichia pastoris, Yarrowia lipolytica, and Ogataea minuta have been used as expression systems. The recombinant lysosomal proteins expressed in these hosts have shown similar substrate specificities, and temperature and pH stability profiles to those produced in mammalian cells. In addition, pre-clinical results have shown that recombinant lysosomal enzymes produced in microorganisms can be taken-up by cells and reduce the substrate accumulated within the lysosome. Recently, metabolic engineering in yeasts has allowed the production of lysosomal enzymes with tailored N-glycosylations, while progresses in E. coli N-glycosylations offer a potential platform to improve the production of these recombinant lysosomal enzymes. In summary, microorganisms represent convenient platform for the production of recombinant lysosomal proteins for biochemical and physicochemical characterization, as well as for the development of ERT for LSD. Copyright © 2015 Elsevier Inc. All rights reserved.

Efficient resource recycling from liquid digestate by microalgae-yeast mixed culture and the assessment of key gene transcription related to nitrogen assimilation in microalgae.

PubMed

Qin, Lei; Liu, Lu; Wang, Zhongming; Chen, Weining; Wei, Dong

2018-05-18

To determine the feasibility of microalgae-yeast mixed culture using the liquid digestate of dairy wastewater (LDDW) for biofuels and single cell protein (SCP) production, the cell growth, nutrient removal and outputs evaluation of the mono and mixed culture of Chlorella vulgaris and Yarrowia lipolytica in LDDW were investigated by adding glycerol as carbon source. The results showed that the mixed culture could enhance the biological utilization efficiency of nitrogen and phosphorus, and obtain higher yield of biomass (1.62 g/L), lipid (0.31 g/L), protein (0.51 g/L), and higher heating value (34.06 KJ/L). Compared with the mono culture of C. vulgaris, a decline of the transcription level in nitrate reductase and glutamine synthetase II genes in C. vulgaris was observed in the mixed culture when ammonia was sufficient. The results suggest the possibility of using the mixed culture for the efficient treatment of LDDW and resources recycling. Copyright © 2018. Published by Elsevier Ltd.

Molecular characterization and expression of microbial inulinase genes.

PubMed

Liu, Guang-Lei; Chi, Zhe; Chi, Zhen-Ming

2013-05-01

Many genes encoding exo- and endo-inulinases from bacteria, yeasts and filamentous fungi have been cloned and characterized. All the inulinases have several conserved motifs, such as WMND(E)PNGL, RDP, EC(V)P, SVEVF, Q and FS(T), which play an important role in inulinase catalysis and substrate binding. However, the exo-inulinases produced by yeasts has no conserved motif SVEVF and the yeasts do not produce any endo-inulinase. Exo- and endo-inulinases found in different microorganisms cluster separately at distant positions from each other. Most of the cloned inulinase genes have been expressed in Yarrowia lipolytica, Saccharomyces cerevisiae, Pichia pastoris, Klyuveromyces lactis and Escherichia coli, respectively. The recombinant inulinases produced and the engineered hosts using the cloned inulinase genes have many potential applications. Expression of most of the inulinase genes is repressed by glucose and fructose and induced by inulin and sucrose. However, the detailed mechanisms of the repression and induction are still unknown.

Metabolic engineering of yeast for lignocellulosic biofuel production.

PubMed

Jin, Yong-Su; Cate, Jamie Hd

2017-12-01

Production of biofuels from lignocellulosic biomass remains an unsolved challenge in industrial biotechnology. Efforts to use yeast for conversion face the question of which host organism to use, counterbalancing the ease of genetic manipulation with the promise of robust industrial phenotypes. Saccharomyces cerevisiae remains the premier host for metabolic engineering of biofuel pathways, due to its many genetic, systems and synthetic biology tools. Numerous engineering strategies for expanding substrate ranges and diversifying products of S. cerevisiae have been developed. Other yeasts generally lack these tools, yet harbor superior phenotypes that could be exploited in the harsh processes required for lignocellulosic biofuel production. These include thermotolerance, resistance to toxic compounds generated during plant biomass deconstruction, and wider carbon consumption capabilities. Although promising, these yeasts have yet to be widely exploited. By contrast, oleaginous yeasts such as Yarrowia lipolytica capable of producing high titers of lipids are rapidly advancing in terms of the tools available for their metabolic manipulation. Copyright © 2017 Elsevier Ltd. All rights reserved.

Synthetic biology and molecular genetics in non-conventional yeasts: Current tools and future advances.

PubMed

Wagner, James M; Alper, Hal S

2016-04-01

Coupling the tools of synthetic biology with traditional molecular genetic techniques can enable the rapid prototyping and optimization of yeast strains. While the era of yeast synthetic biology began in the well-characterized model organism Saccharomyces cerevisiae, it is swiftly expanding to include non-conventional yeast production systems such as Hansenula polymorpha, Kluyveromyces lactis, Pichia pastoris, and Yarrowia lipolytica. These yeasts already have roles in the manufacture of vaccines, therapeutic proteins, food additives, and biorenewable chemicals, but recent synthetic biology advances have the potential to greatly expand and diversify their impact on biotechnology. In this review, we summarize the development of synthetic biological tools (including promoters and terminators) and enabling molecular genetics approaches that have been applied in these four promising alternative biomanufacturing platforms. An emphasis is placed on synthetic parts and genome editing tools. Finally, we discuss examples of synthetic tools developed in other organisms that can be adapted or optimized for these hosts in the near future. Copyright © 2015 Elsevier Inc. All rights reserved.

Comparison of nitrogen depletion and repletion on lipid production in yeast and fungal species

DOE PAGES

Yang, Shihui; Wang, Wei; Wei, Hui; ...

2016-08-29

Although it is well known that low nitrogen stimulates lipid accumulation, especially for algae and some oleaginous yeast, few studies have been conducted in fungal species, especially on the impact of different nitrogen deficiency strategies. In this study, we use two promising consolidated bioprocessing (CBP) candidates to examine the impact of two nitrogen deficiency strategies on lipid production, which are the extensively investigated oleaginous yeast Yarrowia lipolytica, and the commercial cellulase producer Trichoderma reesei. We first utilized bioinformatics approaches to reconstruct the fatty acid metabolic pathway and demonstrated the presence of a triacylglycerol (TAG) biosynthesis pathway in Trichoderma reesei. Wemore » then examined the lipid production of Trichoderma reesei and Y. lipomyces in different media using two nitrogen deficiency strategies of nitrogen natural repletion and nitrogen depletion through centrifugation. Our results demonstrated that nitrogen depletion was better than nitrogen repletion with about 30% lipid increase for Trichoderma reesei and Y. lipomyces, and could be an option to improve lipid production in both oleaginous yeast and filamentous fungal species. The resulting distinctive lipid composition profiles indicated that the impacts of nitrogen depletion on yeast were different from those for fungal species. Under three types of C/N ratio conditions, C16 and C18 fatty acids were the predominant forms of lipids for both Trichoderma reesei and Y. lipolytica. In addition, while the overall fatty acid methyl ester (FAME) profiles of Trichoderma reesei were similar, the overall FAME profiles of Y. lipolytica observed a shift. The fatty acid metabolic pathway reconstructed in this work supports previous reports of lipid production in T. reesei, and provides a pathway for future omics studies and metabolic engineering efforts. Further investigation to identify the genetic targets responsible for the effect of nitrogen depletion

Comparison of nitrogen depletion and repletion on lipid production in yeast and fungal species

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yang, Shihui; Wang, Wei; Wei, Hui

Although it is well known that low nitrogen stimulates lipid accumulation, especially for algae and some oleaginous yeast, few studies have been conducted in fungal species, especially on the impact of different nitrogen deficiency strategies. In this study, we use two promising consolidated bioprocessing (CBP) candidates to examine the impact of two nitrogen deficiency strategies on lipid production, which are the extensively investigated oleaginous yeast Yarrowia lipolytica, and the commercial cellulase producer Trichoderma reesei. We first utilized bioinformatics approaches to reconstruct the fatty acid metabolic pathway and demonstrated the presence of a triacylglycerol (TAG) biosynthesis pathway in Trichoderma reesei. Wemore » then examined the lipid production of Trichoderma reesei and Y. lipomyces in different media using two nitrogen deficiency strategies of nitrogen natural repletion and nitrogen depletion through centrifugation. Our results demonstrated that nitrogen depletion was better than nitrogen repletion with about 30% lipid increase for Trichoderma reesei and Y. lipomyces, and could be an option to improve lipid production in both oleaginous yeast and filamentous fungal species. The resulting distinctive lipid composition profiles indicated that the impacts of nitrogen depletion on yeast were different from those for fungal species. Under three types of C/N ratio conditions, C16 and C18 fatty acids were the predominant forms of lipids for both Trichoderma reesei and Y. lipolytica. In addition, while the overall fatty acid methyl ester (FAME) profiles of Trichoderma reesei were similar, the overall FAME profiles of Y. lipolytica observed a shift. The fatty acid metabolic pathway reconstructed in this work supports previous reports of lipid production in T. reesei, and provides a pathway for future omics studies and metabolic engineering efforts. Further investigation to identify the genetic targets responsible for the effect of nitrogen depletion

A novel phagocytic receptor (CgNimC) from Pacific oyster Crassostrea gigas with lipopolysaccharide and gram-negative bacteria binding activity.

PubMed

Wang, Weilin; Liu, Rui; Zhang, Tao; Zhang, Ran; Song, Xuan; Wang, Lingling; Song, Linsheng

2015-03-01

Phagocytosis is an evolutionarily conserved process to ingest the invading microbes and apoptotic or necrotic corpses, playing vital roles in defensing invaders and maintenance of normal physiological conditions. In the present study, a new Nimrod family phagocytic receptor with three EGF-like domains was identified in Pacific oyster Crassostrea gigas (designated CgNimC). CgNimC shared homology with other identified multiple EGF-like domain containing proteins. The mRNA transcripts of CgNimC were mainly distributed in mantle and hemocytes. Its relative expression level in hemocytes was significantly (P < 0.01) up-regulated after the injection of bacteria Vibrio anguillarum. Different to the NimC in Drosophila and Anopheles gambiae, the recombinant protein of CgNimC (rCgNimC) could bind directly to two gram-negative bacteria V. anguillarum and Vibrio splendidus, but not to gram-positive bacteria Staphylococci aureus, Micrococcus luteus or fungi Yarrowia lipolytica and Pichia pastoris. The affinity of rCgNimC toward M. luteus and Y. lipolytica was enhanced when the microorganisms were pre-incubated with the cell free hemolymph. rCgNimC exhibited higher affinity to lipopolysaccharide (LPS) and relatively lower affinity to peptidoglycan (PGN), while no affinity to glucan (GLU). After the CgNimC receptor was blocked by anti-rCgNimC antibody in vitro, the phagocytic rate of hemocytes toward two gram-negative bacteria V. anguillarum and V. splendidus was reduced significantly (P < 0.05), but no significant change of phagocytic rate was observed toward M. luteus and Y. lipolytica. All these results implied that CgNimC, with significant binding capability to LPS and gram-negative bacteria, was a novel phagocytic receptor involved in immune response of Pacific oyster. Further, it was speculated that receptors of Nimrod family might function as a phagocytic receptor to recognize PAMPs on the invaders and its recognition could be promoted by opsonization of molecules in

Microbial Interactions within a Cheese Microbial Community▿ †

PubMed Central

Mounier, Jérôme; Monnet, Christophe; Vallaeys, Tatiana; Arditi, Roger; Sarthou, Anne-Sophie; Hélias, Arnaud; Irlinger, Françoise

2008-01-01

The interactions that occur during the ripening of smear cheeses are not well understood. Yeast-yeast interactions and yeast-bacterium interactions were investigated within a microbial community composed of three yeasts and six bacteria found in cheese. The growth dynamics of this community was precisely described during the ripening of a model cheese, and the Lotka-Volterra model was used to evaluate species interactions. Subsequently, the effects on ecosystem functioning of yeast omissions in the microbial community were evaluated. It was found both in the Lotka-Volterra model and in the omission study that negative interactions occurred between yeasts. Yarrowia lipolytica inhibited mycelial expansion of Geotrichum candidum, whereas Y. lipolytica and G. candidum inhibited Debaryomyces hansenii cell viability during the stationary phase. However, the mechanisms involved in these interactions remain unclear. It was also shown that yeast-bacterium interactions played a significant role in the establishment of this multispecies ecosystem on the cheese surface. Yeasts were key species in bacterial development, but their influences on the bacteria differed. It appeared that the growth of Arthrobacter arilaitensis or Hafnia alvei relied less on a specific yeast function because these species dominated the bacterial flora, regardless of which yeasts were present in the ecosystem. For other bacteria, such as Leucobacter sp. or Brevibacterium aurantiacum, growth relied on a specific yeast, i.e., G. candidum. Furthermore, B. aurantiacum, Corynebacterium casei, and Staphylococcus xylosus showed reduced colonization capacities in comparison with the other bacteria in this model cheese. Bacterium-bacterium interactions could not be clearly identified. PMID:17981942

Detergent assisted lipid extraction from wet yeast biomass for biodiesel: A response surface methodology approach.

PubMed

Yellapu, Sravan Kumar; Bezawada, Jyothi; Kaur, Rajwinder; Kuttiraja, Mathiazhakan; Tyagi, Rajeshwar D

2016-10-01

The lipid extraction from the microbial biomass is a tedious and high cost dependent process. In the present study, detergent assisted lipids extraction from the culture of the yeast Yarrowia lipolytica SKY-7 was carried out. Response surface methodology (RSM) was used to investigate the effect of three principle parameters (N-LS concentration, time and temperature) on microbial lipid extraction efficiency % (w/w). The results obtained by statistical analysis showed that the quadratic model fits in all cases. Maximum lipid recovery of 95.3±0.3% w/w was obtained at the optimum level of process variables [N-LS concentration 24.42mg (equal to 48mgN-LS/g dry biomass), treatment time 8.8min and reaction temperature 30.2°C]. Whereas the conventional chloroform and methanol extraction to achieve total lipid recovery required 12h at 60°C. The study confirmed that oleaginous yeast biomass treatment with N-lauroyl sarcosine would be a promising approach for industrial scale microbial lipid recovery. Copyright © 2016 Elsevier Ltd. All rights reserved.

Yeast synthetic biology for the production of recombinant therapeutic proteins.

PubMed

Kim, Hyunah; Yoo, Su Jin; Kang, Hyun Ah

2015-02-01

The production of recombinant therapeutic proteins is one of the fast-growing areas of molecular medicine and currently plays an important role in treatment of several diseases. Yeasts are unicellular eukaryotic microbial host cells that offer unique advantages in producing biopharmaceutical proteins. Yeasts are capable of robust growth on simple media, readily accommodate genetic modifications, and incorporate typical eukaryotic post-translational modifications. Saccharomyces cerevisiae is a traditional baker's yeast that has been used as a major host for the production of biopharmaceuticals; however, several nonconventional yeast species including Hansenula polymorpha, Pichia pastoris, and Yarrowia lipolytica have gained increasing attention as alternative hosts for the industrial production of recombinant proteins. In this review, we address the established and emerging genetic tools and host strains suitable for recombinant protein production in various yeast expression systems, particularly focusing on current efforts toward synthetic biology approaches in developing yeast cell factories for the production of therapeutic recombinant proteins. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permission@oup.com.

Accumulation and metabolism of selenium by yeast cells.

PubMed

Kieliszek, Marek; Błażejak, Stanisław; Gientka, Iwona; Bzducha-Wróbel, Anna

2015-07-01

This paper examines the process of selenium bioaccumulation and selenium metabolism in yeast cells. Yeast cells can bind elements in ionic from the environment and permanently integrate them into their cellular structure. Up to now, Saccharomyces cerevisiae, Candida utilis, and Yarrowia lipolytica yeasts have been used primarily in biotechnological studies to evaluate binding of minerals. Yeast cells are able to bind selenium in the form of both organic and inorganic compounds. The process of bioaccumulation of selenium by microorganisms occurs through two mechanisms: extracellular binding by ligands of membrane assembly and intracellular accumulation associated with the transport of ions across the cytoplasmic membrane into the cell interior. During intracellular metabolism of selenium, oxidation, reduction, methylation, and selenoprotein synthesis processes are involved, as exemplified by detoxification processes that allow yeasts to survive under culture conditions involving the elevated selenium concentrations which were observed. Selenium yeasts represent probably the best absorbed form of this element. In turn, in terms of wide application, the inclusion of yeast with accumulated selenium may aid in lessening selenium deficiency in a diet.

CE separation of proteins and yeasts dynamically modified by PEG pyrenebutanoate with fluorescence detection.

PubMed

Horká, Marie; Růzicka, Filip; Holá, Veronika; Slais, Karel

2007-07-01

The optimized protocols of the bioanalytes separation, proteins and yeasts, dynamically modified by the nonionogenic tenside PEG pyrenebutanoate, were applied in CZE and CIEF with the acidic gradient in pH range 2-5.5, both with fluorescence detection. PEG pyrenebutanoate was used as a buffer additive for a dynamic modification of proteins and/or yeast samples. The narrow peaks of modified analytes were detected. The values of the pI's of the labeled proteins were calculated using new fluorescent pI markers in CIEF and they were found to be comparable with pI's of the native compounds. As an example of the possible use of the suggested CIEF technique, the mixed cultures of yeasts, Candida albicans, Candida glabrata, Candida kefyr, Candida krusei, Candida lusitaniae, Candida parapsilosis, Candida tropicalis, Candida zeylanoides, Geotrichum candidum, Saccharomyces cerevisiae, Trichosporon asahii and Yarrowia lipolytica, were reproducibly focused and separated with high sensitivity. Using UV excitation for the on-column fluorometric detection, the minimum detectable amounts of analytes, femtograms of proteins and down to ten cells injected on the separation capillary, were estimated.

A critical tyrosine residue determines the uncoupling protein-like activity of the yeast mitochondrial oxaloacetate carrier.

PubMed

Luévano-Martínez, Luis A; Barba-Ostria, Carlos; Araiza-Olivera, Daniela; Chiquete-Félix, Natalia; Guerrero-Castillo, Sergio; Rial, Eduardo; Georgellis, Dimitris; Uribe-Carvajal, Salvador

2012-04-01

The mitochondrial Oac (oxaloacetate carrier) found in some fungi and plants catalyses the uptake of oxaloacetate, malonate and sulfate. Despite their sequence similarity, transport specificity varies considerably between Oacs. Indeed, whereas ScOac (Saccharomyces cerevisiae Oac) is a specific anion-proton symporter, the YlOac (Yarrowia lipolytica Oac) has the added ability to transport protons, behaving as a UCP (uncoupling protein). Significantly, we identified two amino acid changes at the matrix gate of YlOac and ScOac, tyrosine to phenylalanine and methionine to leucine. We studied the role of these amino acids by expressing both wild-type and specifically mutated Oacs in an Oac-null S. cerevisiae strain. No phenotype could be associated with the methionine to leucine substitution, whereas UCP-like activity was dependent on the presence of the tyrosine residue normally expressed in the YlOac, i.e. Tyr-ScOac mediated proton transport, whereas Phe-YlOac lost its protonophoric activity. These findings indicate that the UCP-like activity of YlOac is determined by the tyrosine residue at position 146.

Characterization of self-generated variants in Pseudoalteromonas lipolytica biofilm with increased antifouling activities.

PubMed

Zeng, Zhenshun; Guo, Xing-Pan; Li, Baiyuan; Wang, Pengxia; Cai, Xingsheng; Tian, Xinpeng; Zhang, Si; Yang, Jin-Long; Wang, Xiaoxue

2015-12-01

Pseudoalteromonas is widespread in various marine environments, and most strains can affect invertebrate larval settlement and metamorphosis by forming biofilms. However, the impact and the molecular basis of population diversification occurring in Pseudoalteromonas biofilms are poorly understood. Here, we show that morphological diversification is prevalent in Pseudoalteromonas species during biofilm formation. Two types of genetic variants, wrinkled (frequency of 12±5%) and translucent (frequency of 5±3%), were found in Pseudoalteromonas lipolytica biofilms. The inducing activities of biofilms formed by the two variants on larval settlement and metamorphosis of the mussel Mytilus coruscus were significantly decreased, suggesting strong antifouling activities. Using whole-genome re-sequencing combined with genetic manipulation, two genes were identified to be responsible for the morphology alternations. A nonsense mutation in AT00_08765 led to a wrinkled morphology due to the overproduction of cellulose, whereas a point mutation in AT00_17125 led to a translucent morphology via a reduction in capsular polysaccharide production. Taken together, the results suggest that the microbial behavior on larval settlement and metamorphosis in marine environment could be affected by the self-generated variants generated during the formation of marine biofilms, thereby rendering potential application in biocontrol of marine biofouling.

Rapid fabrication of three-dimensional structures for dielectrophoretic sorting of lipid-containing organisms

NASA Astrophysics Data System (ADS)

Schor, Alisha R.; Buie, Cullen R.

2016-10-01

In this work, we demonstrate a microfluidic particle sorter consisting of three-dimensional, conducting microposts. Our sorter uses dielectrophoresis (DEP) to sort high- and low-lipid phenotypes of the yeast Yarrowia lipolytica. Y. lipolytica is one of the many microorganisms being explored as a hydrocarbon source for biodiesel, Omega-3 additives, and other products derived from fatty acids. A rapid, non-destructive, lipid-based sorting tool would accelerate the commercialization of these products. Our device consists of an array of 105, 25 μm wide gold microposts that span the height of a 15 μm channel. This array generates an electric field in a microfluidic device that is uniform through the channel height, but has a custom-shaped non-uniformity in the horizontal directions. This is crucial in order to achieve continuous sorting using DEP, as it ensures all cells are exposed to the same conditions throughout the channel height. By using very low currents (100 μA), we are able to electroplate these post arrays in fewer than 15 min. This is an order of magnitude improvement over previous reports of electroplated microstructures. With an applied signal of 250 MHz, 2.6 V pp in our device, we separate a heterogeneous population with a purity of 97.8% in the low-lipid stream and 71.4% in the high-lipid stream. The high-lipid stream purity can be improved by adjusting the spacing of the array. This unique protocol for the rapid fabrication of 3D microstructures has enabled the creation of a non-invasive sorting tool for genetically engineered, lipid-producing organisms. The ability to screen organisms based on lipid content will alleviate one of the major bottlenecks in commercialization of microbial biofuels.

Utilization of inulin-containing waste in industrial fermentations to produce biofuels and bio-based chemicals.

PubMed

Hughes, Stephen R; Qureshi, Nasib; López-Núñez, Juan Carlos; Jones, Marjorie A; Jarodsky, Joshua M; Galindo-Leva, Luz Ángela; Lindquist, Mitchell R

2017-04-01

Inulins are polysaccharides that belong to an important class of carbohydrates known as fructans and are used by many plants as a means of storing energy. Inulins contain 20 to several thousand fructose units joined by β-2,1 glycosidic bonds, typically with a terminal glucose unit. Plants with high concentrations of inulin include: agave, asparagus, coffee, chicory, dahlia, dandelion, garlic, globe artichoke, Jerusalem artichoke, jicama, onion, wild yam, and yacón. To utilize inulin as its carbon and energy source directly, a microorganism requires an extracellular inulinase to hydrolyze the glycosidic bonds to release fermentable monosaccharides. Inulinase is produced by many microorganisms, including species of Aspergillus, Kluyveromyces, Penicillium, and Pseudomonas. We review various inulinase-producing microorganisms and inulin feedstocks with potential for industrial application as well as biotechnological efforts underway to develop sustainable practices for the disposal of residues from processing inulin-containing crops. A multi-stage biorefinery concept is proposed to convert cellulosic and inulin-containing waste produced at crop processing operations to valuable biofuels and bioproducts using Kluyveromyces marxianus, Yarrowia lipolytica, Rhodotorula glutinis, and Saccharomyces cerevisiae as well as thermochemical treatments.

Genome-scale metabolic modeling of Mucor circinelloides and comparative analysis with other oleaginous species.

PubMed

Vongsangnak, Wanwipa; Klanchui, Amornpan; Tawornsamretkit, Iyarest; Tatiyaborwornchai, Witthawin; Laoteng, Kobkul; Meechai, Asawin

2016-06-01

We present a novel genome-scale metabolic model iWV1213 of Mucor circinelloides, which is an oleaginous fungus for industrial applications. The model contains 1213 genes, 1413 metabolites and 1326 metabolic reactions across different compartments. We demonstrate that iWV1213 is able to accurately predict the growth rates of M. circinelloides on various nutrient sources and culture conditions using Flux Balance Analysis and Phenotypic Phase Plane analysis. Comparative analysis of three oleaginous genome-scale models, including M. circinelloides (iWV1213), Mortierella alpina (iCY1106) and Yarrowia lipolytica (iYL619_PCP) revealed that iWV1213 possesses a higher number of genes involved in carbohydrate, amino acid, and lipid metabolisms that might contribute to its versatility in nutrient utilization. Moreover, the identification of unique and common active reactions among the Zygomycetes oleaginous models using Flux Variability Analysis unveiled a set of gene/enzyme candidates as metabolic engineering targets for cellular improvement. Thus, iWV1213 offers a powerful metabolic engineering tool for multi-level omics analysis, enabling strain optimization as a cell factory platform of lipid-based production. Copyright © 2016 Elsevier B.V. All rights reserved.

Comparative Lipidomic Profiling of S. cerevisiae and Four Other Hemiascomycetous Yeasts

PubMed Central

Hein, Eva-Maria; Hayen, Heiko

2012-01-01

Glycerophospholipids (GP) are the building blocks of cellular membranes and play essential roles in cell compartmentation, membrane fluidity or apoptosis. In addition, GPs are sources for multifunctional second messengers. Whereas the genome and proteome of the most intensively studied eukaryotic model organism, the baker’s yeast (Saccharomyces cerevisiae), are well characterized, the analysis of its lipid composition is still at the beginning. Moreover, different yeast species can be distinguished on the DNA, RNA and protein level, but it is currently unknown if they can also be differentiated by determination of their GP pattern. Therefore, the GP compositions of five different yeast strains, grown under identical environmental conditions, were elucidated using high performance liquid chromatography coupled to negative electrospray ionization-hybrid linear ion trap-Fourier transform ion cyclotron resonance mass spectrometry in single and multistage mode. Using this approach, relative quantification of more than 100 molecular species belonging to nine GP classes was achieved. The comparative lipidomic profiling of Saccharomyces cerevisiae, Saccharomyces bayanus, Kluyveromyces thermotolerans, Pichia angusta, and Yarrowia lipolytica revealed characteristic GP profiles for each strain. However, genetically related yeast strains show similarities in their GP compositions, e.g., Saccharomyces cerevisiae and Saccharomyces bayanus. PMID:24957378

Oleaginous yeasts: Promising platforms for the production of oleochemicals and biofuels.

PubMed

Adrio, José L

2017-09-01

Oleaginous yeasts have a unique physiology that makes them the best suited hosts for the production of lipids, oleochemicals, and diesel-like fuels. Their high lipogenesis, capability of growing on many different carbon sources (including lignocellulosic sugars), easy large-scale cultivation, and an increasing number of genetic tools are some of the advantages that have encouraged their use to develop sustainable processes. This mini-review summarizes the metabolic engineering strategies developed in oleaginous yeasts within the last 2 years to improve process metrics (titer, yield, and productivity) for the production of lipids, free fatty acids, fatty acid-based chemicals (e.g., fatty alcohols, fatty acid ethyl esters), and alkanes. During this short period of time, tremendous progress has been made in Yarrowia lipolytica, the model oleaginous yeast, which has been engineered to improve lipid production by different strategies including increasing lipogenic pathway flux and biosynthetic precursors, and blocking degradation pathways. Moreover, remarkable advances have also been reported in Rhodosporidium toruloides and Lipomyces starkey despite the limited genetic tools available for these two very promising hosts. Biotechnol. Bioeng. 2017;114: 1915-1920. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

Genome and metabolic engineering in non-conventional yeasts: Current advances and applications.

PubMed

Löbs, Ann-Kathrin; Schwartz, Cory; Wheeldon, Ian

2017-09-01

Microbial production of chemicals and proteins from biomass-derived and waste sugar streams is a rapidly growing area of research and development. While the model yeast Saccharomyces cerevisia e is an excellent host for the conversion of glucose to ethanol, production of other chemicals from alternative substrates often requires extensive strain engineering. To avoid complex and intensive engineering of S. cerevisiae, other yeasts are often selected as hosts for bioprocessing based on their natural capacity to produce a desired product: for example, the efficient production and secretion of proteins, lipids, and primary metabolites that have value as commodity chemicals. Even when using yeasts with beneficial native phenotypes, metabolic engineering to increase yield, titer, and production rate is essential. The non-conventional yeasts Kluyveromyces lactis, K. marxianus, Scheffersomyces stipitis, Yarrowia lipolytica, Hansenula polymorpha and Pichia pastoris have been developed as eukaryotic hosts because of their desirable phenotypes, including thermotolerance, assimilation of diverse carbon sources, and high protein secretion. However, advanced metabolic engineering in these yeasts has been limited. This review outlines the challenges of using non-conventional yeasts for strain and pathway engineering, and discusses the developed solutions to these problems and the resulting applications in industrial biotechnology.

The expression of glycerol facilitators from various yeast species improves growth on glycerol of Saccharomyces cerevisiae.

PubMed

Klein, Mathias; Islam, Zia-Ul; Knudsen, Peter Boldsen; Carrillo, Martina; Swinnen, Steve; Workman, Mhairi; Nevoigt, Elke

2016-12-01

Glycerol is an abundant by-product during biodiesel production and additionally has several assets compared to sugars when used as a carbon source for growing microorganisms in the context of biotechnological applications. However, most strains of the platform production organism Saccharomyces cerevisiae grow poorly in synthetic glycerol medium. It has been hypothesized that the uptake of glycerol could be a major bottleneck for the utilization of glycerol in S. cerevisiae . This species exclusively relies on an active transport system for glycerol uptake. This work demonstrates that the expression of predicted glycerol facilitators (Fps1 homologues) from superior glycerol-utilizing yeast species such as Pachysolen tannophilus , Komagataella pastoris , Yarrowia lipolytica and Cyberlindnera jadinii significantly improves the growth performance on glycerol of the previously selected glycerol-consuming S. cerevisiae wild-type strain (CBS 6412-13A). The maximum specific growth rate increased from 0.13 up to 0.18 h -1 and a biomass yield coefficient of 0.56 g DW /g glycerol was observed. These results pave the way for exploiting the assets of glycerol in the production of fuels, chemicals and pharmaceuticals based on baker's yeast.

Characterization of the newly isolated ω-oxidizing yeast Candida sorbophila DS02 and its potential applications in long-chain dicarboxylic acid production.

PubMed

Lee, Heeseok; Sugiharto, Yohanes Eko Chandra; Lee, Seunghoon; Park, Gyuyeon; Han, Changpyo; Jang, Hyeran; Jeon, Wooyoung; Park, Heejoon; Ahn, Jungoh; Kang, Kyungbo; Lee, Hongwoen

2017-08-01

α, ω-Dicarboxylic acids (DCAs) are multipurpose chemicals widely used in polymers, perfumes, plasticizers, lubricants, and adhesives. The biotransformation of DCAs from alkanes and fatty acids by microorganisms has attracted recent interest, since synthesis via chemical oxidation causes problems in terms of the environment and safety. We isolated an ω-oxidizing yeast from a wastewater disposal facility of a petrochemical factory by chemostat enrichment culture. The haploid strain identified as Candida sorbophila DS02 grew on glucose and dodecane, exhibiting greater cell shrinkage on the latter. In flask cultures with mixed alkanes (C10-16) and fatty acid methyl esters (C10-16), DS02 used mixed alkanes simultaneously unlike Candida tropicalis and Yarrowia lipolytica and showed high substrate resistance. In flask cultures with acrylic acid-a known inhibitor of β-oxidation-DS02 produced 0.28 g/l dodecanedioic acid (DDDA) from dodecane, similar to wild-type C. tropicalis ATCC 20336. In fed-batch fermentation, DS02 produced 9.87 g/l DDDA, which was 5.7-fold higher than wild-type C. tropicalis. These results suggest that C. sorbophila strain DS02 has potential applications for the large-scale production of DCA.

Cloning, expression and characterization of a lipase gene from marine bacterium Pseudoalteromonas lipolytica SCSIO 04301

NASA Astrophysics Data System (ADS)

Su, Hongfei; Mai, Zhimao; Zhang, Si

2016-12-01

A lipase gene, lip1233, isolated from Pseudoalteromonas lipolytica SCSIO 04301, was cloned and expressed in E. coli. The enzyme comprised 810 amino acid residues with a deduced molecular weight of 80 kDa. Lip1233 was grouped into the lipase family X because it contained a highly conserved motif GHSLG. The recombinant enzyme was purified with Ni-NTA affinity chromatography. The optimal temperature and pH value of Lip1233 were 45°C and 8.0, respectively. It retained more than 70% of original activity after being incubated in pH ranging from 6.0 to 9.5 for 30 min. It was stable when the temperature was below 45°C, but was unstable when the temperature was above 55°C. Most metal ions tested had no significant effect on the activity of Lip1233. Lip1233 remained more than original activity in some organic solvents at the concentration of 30% (v/v). It retained more than 30% activity after incubated in pure organic solvents for 12 h, while in hexane the activity was nearly 100%. Additionally, Lip1233 exhibited typical halotolerant characteristic as it was active under 4M NaCl. Lip1233 powder could catalyze efficiently the synthesis of fructose esters in hexane at 40°C. These characteristics demonstrated that Lip1233 is applicable to elaborate food processing and organic synthesis.

Subunit mass fingerprinting of mitochondrial complex I.

PubMed

Morgner, Nina; Zickermann, Volker; Kerscher, Stefan; Wittig, Ilka; Abdrakhmanova, Albina; Barth, Hans-Dieter; Brutschy, Bernhard; Brandt, Ulrich

2008-10-01

We have employed laser induced liquid bead ion desorption (LILBID) mass spectrometry to determine the total mass and to study the subunit composition of respiratory chain complex I from Yarrowia lipolytica. Using 5-10 pmol of purified complex I, we could assign all 40 known subunits of this membrane bound multiprotein complex to peaks in LILBID subunit fingerprint spectra by comparing predicted protein masses to observed ion masses. Notably, even the highly hydrophobic subunits encoded by the mitochondrial genome were easily detectable. Moreover, the LILBID approach allowed us to spot and correct several errors in the genome-derived protein sequences of complex I subunits. Typically, the masses of the individual subunits as determined by LILBID mass spectrometry were within 100 Da of the predicted values. For the first time, we demonstrate that LILBID spectrometry can be successfully applied to a complex I band eluted from a blue-native polyacrylamide gel, making small amounts of large multiprotein complexes accessible for subunit mass fingerprint analysis even if they are membrane bound. Thus, the LILBID subunit mass fingerprint method will be of great value for efficient proteomic analysis of complex I and its assembly intermediates, as well as of other water soluble and membrane bound multiprotein complexes.

Comparison of volatile sulphur compound production by cheese-ripening yeasts from methionine and methionine-cysteine mixtures.

PubMed

López Del Castillo-Lozano, M; Delile, A; Spinnler, H E; Bonnarme, P; Landaud, S

2007-07-01

Production of volatile sulphur compounds (VSC) was assessed in culture media supplemented with L-methionine or L-methionine/L-cysteine mixtures, using five cheese-ripening yeasts: Debaryomyces hansenii DH47(8), Kluyveromyces lactis KL640, Geotrichum candidum GC77, Yarrowia lipolytica YL200 and Saccharomyces cerevisiae SC45(3). All five yeasts produced VSC with L-methionine or L-methionine/L-cysteine, but different VSC profiles were found. GC77 and YL200 produced dimethyldisulphide and trace levels of dimethyltrisulphide while DH47(8), KL640 and SC45(3) produced mainly methionol and low levels of methional. S-methylthioacetate was produced by all the yeasts but at different concentrations. DH47(8), KL640 and SC45(3) also produced other minor VSC including 3-methylthiopropyl acetate, ethyl-3-methylthiopropanoate, a thiophenone, and an oxathiane. However, VSC production diminished in a strain-dependent behaviour when L-cysteine was supplemented, even at a low concentration (0.2 g l(-1)). This effect was due mainly to a significant decrease in L-methionine consumption in all the yeasts except YL200. Hydrogen sulphide produced by L-cysteine catabolism did not seem to contribute to VSC generation at the acid pH of yeast cultures. The significance of such results in the cheese-ripening context is discussed.

Harnessing biodiesel-producing microbes: from genetic engineering of lipase to metabolic engineering of fatty acid biosynthetic pathway.

PubMed

Yan, Jinyong; Yan, Yunjun; Madzak, Catherine; Han, Bingnan

2017-02-01

Microbial production routes, notably whole-cell lipase-mediated biotransformation and fatty-acids-derived biosynthesis, offer new opportunities for synthesizing biodiesel. They compare favorably to immobilized lipase and chemically catalyzed processes. Genetically modified whole-cell lipase-mediated in vitro route, together with in vivo and ex vivo microbial biosynthesis routes, constitutes emerging and rapidly developing research areas for effective production of biodiesel. This review presents recent advances in customizing microorganisms for producing biodiesel, via genetic engineering of lipases and metabolic engineering (including system regulation) of fatty-acids-derived pathways. Microbial hosts used include Escherichia coli, Saccharomyces cerevisiae, Pichia pastoris and Aspergillus oryzae. These microbial cells can be genetically modified to produce lipases under different forms: intracellularly expressed, secreted or surface-displayed. They can be metabolically redesigned and systematically regulated to obtain balanced biodiesel-producing cells, as highlighted in this study. Such genetically or metabolically modified microbial cells can support not only in vitro biotransformation of various common oil feedstocks to biodiesel, but also de novo biosynthesis of biodiesel from glucose, glycerol or even cellulosic biomass. We believe that the genetically tractable oleaginous yeast Yarrowia lipolytica could be developed to an effective biodiesel-producing microbial cell factory. For this purpose, we propose several engineered pathways, based on lipase and wax ester synthase, in this promising oleaginous host.

Comparison of Yeasts as Hosts for Recombinant Protein Production.

PubMed

Vieira Gomes, Antonio Milton; Souza Carmo, Talita; Silva Carvalho, Lucas; Mendonça Bahia, Frederico; Parachin, Nádia Skorupa

2018-04-29

Recombinant protein production emerged in the early 1980s with the development of genetic engineering tools, which represented a compelling alternative to protein extraction from natural sources. Over the years, a high level of heterologous protein was made possible in a variety of hosts ranging from the bacteria Escherichia coli to mammalian cells. Recombinant protein importance is represented by its market size, which reached $1654 million in 2016 and is expected to reach $2850.5 million by 2022. Among the available hosts, yeasts have been used for producing a great variety of proteins applied to chemicals, fuels, food, and pharmaceuticals, being one of the most used hosts for recombinant production nowadays. Historically, Saccharomyces cerevisiae was the dominant yeast host for heterologous protein production. Lately, other yeasts such as Komagataella sp., Kluyveromyces lactis , and Yarrowia lipolytica have emerged as advantageous hosts. In this review, a comparative analysis is done listing the advantages and disadvantages of using each host regarding the availability of genetic tools, strategies for cultivation in bioreactors, and the main techniques utilized for protein purification. Finally, examples of each host will be discussed regarding the total amount of protein recovered and its bioactivity due to correct folding and glycosylation patterns.

Immobilization of Yarrowia lipolytica Lipase on Macroporous Resin Using Different Methods: Characterization of the Biocatalysts in Hydrolysis Reaction.

PubMed

Sun, Jingjing; Chen, Yiling; Sheng, Jun; Sun, Mi

2015-01-01

To improve the reusability and organic solvent tolerance of microbial lipase and expand the application of lipase (hydrolysis, esterification, and transesterification), we immobilized marine microbial lipase using different methods and determined the properties of immobilized lipases. Considering the activity and cost of immobilized lipase, the concentration of lipase was fixed at 2 mg/mL. The optimal temperature of immobilized lipases was 40°C and 5°C higher than free lipase. The activities of immobilized lipases were much higher than free lipase at alkaline pH (more than 50% at pH 12). The free lipase lost most activity (35.3%) and immobilized lipases retained more than 46.4% of their initial activity after 3 h heat treatment at 70°C. At alkaline pH, immobilized lipases were more stable than free lipase (more than 60% residue activity at pH 11 for 3 h). Immobilized lipases retained 80% of their activity after 5 cycles and increased enzyme activity (more than 108.7%) after 3 h treatment in tert-butanol. Immobilization of lipase which improved reusability of lipase and provided a chance to expand the application of marine microbial lipase in organic system expanded the application range of lipase to catalyze hydrolysis and esterification in harsh condition.

Fine Structure of Tibetan Kefir Grains and Their Yeast Distribution, Diversity, and Shift

PubMed Central

Lu, Man; Wang, Xingxing; Sun, Guowei; Qin, Bing; Xiao, Jinzhou; Yan, Shuling; Pan, Yingjie; Wang, Yongjie

2014-01-01

Tibetan kefir grains (TKGs), a kind of natural starter for fermented milk in Tibet, China, host various microorganisms of lactic acid bacteria, yeasts, and occasionally acetic acid bacteria in a polysaccharide/protein matrix. In the present study, the fine structure of TKGs was studied to shed light on this unusual symbiosis with stereomicroscopy and thin sections. The results reveal that TKGs consist of numerous small grain units, which are characterized by a hollow globular structure with a diameter between 2.0 and 9.0 mm and a wall thickness of approximately 200 µm. A polyhedron-like net structure, formed mainly by the bacteria, was observed in the wall of the grain units, which has not been reported previously to our knowledge. Towards the inside of the grain unit, the polyhedron-like net structures became gradually larger in diameter and fewer in number. Such fine structures may play a crucial role in the stability of the grains. Subsequently, the distribution, diversity, and shift of yeasts in TKGs were investigated based on thin section, scanning electron microscopy, cloning and sequencing of D1/D2 of the 26S rRNA gene, real-time quantitative PCR, and in situ hybridization with specific fluorescence-labeled oligonucleotide probes. These show that (i) yeasts appear to localize on the outer surface of the grains and grow normally together to form colonies embedded in the bacterial community; (ii) the diversity of yeasts is relatively low on genus level with three dominant species – Saccharomyces cerevisiae, Kluyveromyces marxianus, and Yarrowia lipolytica; (iii) S. cerevisiae is the stable predominant yeast species, while the composition of Kluyveromyces and Yarrowia are subject to change over time. Our results indicate that TKGs are relatively stable in structure, and culture conditions to some extent shape the microbial community and interaction in kefir grains. These findings pave the way for further study of the specific symbiotic associations between S

Candida lipolytica UCP0988 Biosurfactant: Potential as a Bioremediation Agent and in Formulating a Commercial Related Product

PubMed Central

Santos, Danyelle K. F.; Resende, Ana H. M.; de Almeida, Darne G.; Soares da Silva, Rita de Cássia F.; Rufino, Raquel D.; Luna, Juliana M.; Banat, Ibrahim M.; Sarubbo, Leonie A.

2017-01-01

The aim of the present study was to investigate the potential application of the biosurfactant from Candida lipolytica grown in low-cost substrates, which has previously been produced and characterized under optimized conditions as an adjunct material to enhance the remediation processes of hydrophobic pollutants and heavy metals generated by the oil industry and propose the formulation of a safe and stable remediation agent. In tests carried out with seawater, the crude biosurfactant demonstrated 80% oil spreading efficiency. The dispersion rate was 50% for the biosurfactant at a concentration twice that of the CMC. The biosurfactant removed 70% of motor oil from contaminated cotton cloth in detergency tests. The crude biosurfactant also removed 30–40% of Cu and Pb from standard sand, while the isolated biosurfactant removed ~30% of the heavy metals. The conductivity of solutions containing Cd and Pb was sharply reduced after biosurfactants' addition. A product was prepared through adding 0.2% potassium sorbate as preservative and tested over 120 days. The formulated biosurfactant was analyzed for emulsification and surface tension under different pH values, temperatures, and salt concentrations and tested for toxicity against the fish Poecilia vivipara. The results showed that the formulation had no toxicity and did not cause significant changes in the tensoactive capacity of the biomolecule while maintaining activity demonstrating suitability for potential future commercial product formulation. PMID:28507538

Balanced Biochemical Reactions: A New Approach to Unify Chemical and Biochemical Thermodynamics

PubMed Central

Sabatini, Antonio; Vacca, Alberto; Iotti, Stefano

2012-01-01

A novel procedure is presented which, by balancing elements and electric charge of biochemical reactions which occur at constant pH and pMg, allows assessing the thermodynamics properties of reaction ΔrG ′0, ΔrH ′0, ΔrS ′0 and the change in binding of hydrogen and magnesium ions of these reactions. This procedure of general applicability avoids the complex calculations required by the use of the Legendre transformed thermodynamic properties of formation ΔfG ′0, ΔfH ′0 and ΔfS ′0 hitherto considered an obligatory prerequisite to deal with the thermodynamics of biochemical reactions. As a consequence, the term “conditional” is proposed in substitution of “Legendre transformed” to indicate these thermodynamics properties. It is also shown that the thermodynamic potential G is fully adequate to give a criterion of spontaneous chemical change for all biochemical reactions and then that the use of the Legendre transformed G′ is unnecessary. The procedure proposed can be applied to any biochemical reaction, making possible to re-unify the two worlds of chemical and biochemical thermodynamics, which so far have been treated separately. PMID:22247780

The requirement of sterol glucoside for pexophagy in yeast is dependent on the species and nature of peroxisome inducers.

PubMed

Nazarko, Taras Y; Polupanov, Andriy S; Manjithaya, Ravi R; Subramani, Suresh; Sibirny, Andriy A

2007-01-01

Sterol glucosyltransferase, Ugt51/Atg26, is essential for both micropexophagy and macropexophagy of methanol-induced peroxisomes in Pichia pastoris. However, the role of this protein in pexophagy in other yeast remained unclear. We show that oleate- and amine-induced peroxisomes in Yarrowia lipolytica are degraded by Atg26-independent macropexophagy. Surprisingly, Atg26 was also not essential for macropexophagy of oleate- and amine-induced peroxisomes in P. pastoris, suggesting that the function of sterol glucoside (SG) in pexophagy is both species and peroxisome inducer specific. However, the rates of degradation of oleate- and amine-induced peroxisomes in P. pastoris were reduced in the absence of SG, indicating that P. pastoris specifically uses sterol conversion by Atg26 to enhance selective degradation of peroxisomes. However, methanol-induced peroxisomes apparently have lost the redundant ability to be degraded without SG. We also show that the P. pastoris Vac8 armadillo repeat protein is not essential for macropexophagy of methanol-, oleate-, or amine-induced peroxisomes, which makes PpVac8 the first known protein required for the micropexophagy, but not for the macropexophagy, machinery. The uniqueness of Atg26 and Vac8 functions under different pexophagy conditions demonstrates that not only pexophagy inducers, such as glucose or ethanol, but also the inducers of peroxisomes, such as methanol, oleate, or primary amines, determine the requirements for subsequent pexophagy in yeast.

Oxidative phosphorylation in Debaryomyces hansenii: physiological uncoupling at different growth phases.

PubMed

Cabrera-Orefice, Alfredo; Guerrero-Castillo, Sergio; Díaz-Ruíz, Rodrigo; Uribe-Carvajal, Salvador

2014-07-01

Physiological uncoupling of mitochondrial oxidative phosphorylation (OxPhos) was studied in Debaryomyces hansenii. In other species, such as Yarrowia lipolytica and Saccharomyces cerevisiae, OxPhos can be uncoupled through differential expression of branched respiratory chain enzymes or by opening of a mitochondrial unspecific channel (ScMUC), respectively. However D. hansenii mitochondria, which contain both a branched respiratory chain and a mitochondrial unspecific channel (DhMUC), selectively uncouple complex I-dependent rate of oxygen consumption in the stationary growth phase. The uncoupled complex I-dependent respiration was only 20% of the original activity. Inhibition was not due to inactivation of complex I, lack of protein expression or to differential expression of alternative oxidoreductases. Furthermore, all other respiratory chain activities were normal. Decrease of complex I-dependent respiration was due to NAD(+) loss from the matrix, probably through an open of DhMUC. When NAD(+) was added back, coupled complex I-activity was recovered. NAD(+) re-uptake was independent of DhMUC opening and seemed to be catalyzed by a NAD(+)-specific transporter, which was sensitive to bathophenanthroline, bromocresol purple or pyridoxal-5'-phosphate as described for S. cerevisiae mitochondrial NAD(+) transporters. Loss of NAD(+) from the matrix through an open MUC is proposed as an additional mechanism to uncouple OxPhos. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

"Bligh and Dyer" and Folch Methods for Solid-Liquid-Liquid Extraction of Lipids from Microorganisms. Comprehension of Solvatation Mechanisms and towards Substitution with Alternative Solvents.

PubMed

Breil, Cassandra; Abert Vian, Maryline; Zemb, Thomas; Kunz, Werner; Chemat, Farid

2017-03-27

Bligh and Dyer (B & D) or Folch procedures for the extraction and separation of lipids from microorganisms and biological tissues using chloroform/methanol/water have been used tens of thousands of times and are "gold standards" for the analysis of extracted lipids. Based on the Conductor-like Screening MOdel for realistic Solvatation (COSMO-RS), we select ethanol and ethyl acetate as being potentially suitable for the substitution of methanol and chloroform. We confirm this by performing solid-liquid extraction of yeast ( Yarrowia lipolytica IFP29 ) and subsequent liquid-liquid partition-the two steps of routine extraction. For this purpose, we consider similar points in the ternary phase diagrams of water/methanol/chloroform and water/ethanol/ethyl acetate, both in the monophasic mixtures and in the liquid-liquid miscibility gap. Based on high performance thin-layer chromatography (HPTLC) to obtain the distribution of lipids classes, and gas chromatography coupled with a flame ionisation detector (GC/FID) to obtain fatty acid profiles, this greener solvents pair is found to be almost as effective as the classic methanol-chloroform couple in terms of efficiency and selectivity of lipids and non-lipid material. Moreover, using these bio-sourced solvents as an alternative system is shown to be as effective as the classical system in terms of the yield of lipids extracted from microorganism tissues, independently of their apparent hydrophilicity.

“Bligh and Dyer” and Folch Methods for Solid–Liquid–Liquid Extraction of Lipids from Microorganisms. Comprehension of Solvatation Mechanisms and towards Substitution with Alternative Solvents

PubMed Central

Breil, Cassandra; Abert Vian, Maryline; Zemb, Thomas; Kunz, Werner; Chemat, Farid

2017-01-01

Bligh and Dyer (B & D) or Folch procedures for the extraction and separation of lipids from microorganisms and biological tissues using chloroform/methanol/water have been used tens of thousands of times and are “gold standards” for the analysis of extracted lipids. Based on the Conductor-like Screening MOdel for realistic Solvatation (COSMO-RS), we select ethanol and ethyl acetate as being potentially suitable for the substitution of methanol and chloroform. We confirm this by performing solid–liquid extraction of yeast (Yarrowia lipolytica IFP29) and subsequent liquid–liquid partition—the two steps of routine extraction. For this purpose, we consider similar points in the ternary phase diagrams of water/methanol/chloroform and water/ethanol/ethyl acetate, both in the monophasic mixtures and in the liquid–liquid miscibility gap. Based on high performance thin-layer chromatography (HPTLC) to obtain the distribution of lipids classes, and gas chromatography coupled with a flame ionisation detector (GC/FID) to obtain fatty acid profiles, this greener solvents pair is found to be almost as effective as the classic methanol–chloroform couple in terms of efficiency and selectivity of lipids and non-lipid material. Moreover, using these bio-sourced solvents as an alternative system is shown to be as effective as the classical system in terms of the yield of lipids extracted from microorganism tissues, independently of their apparent hydrophilicity. PMID:28346372

The modulation of extracellular superoxide dismutase in the specifically enhanced cellular immune response against secondary challenge of Vibrio splendidus in Pacific oyster (Crassostrea gigas).

PubMed

Liu, Conghui; Zhang, Tao; Wang, Lingling; Wang, Mengqiang; Wang, Weilin; Jia, Zhihao; Jiang, Shuai; Song, Linsheng

2016-10-01

Extracellular superoxide dismutase (EcSOD) is a copper-containing glycoprotein playing an important role in antioxidant defense of living cells exposed to oxidative stress, and also participating in microorganism internalization and cell adhesion in invertebrates. EcSOD from oyster (designated CgEcSOD) had been previously reported to bind lipopolysaccharides (LPS) and act as a bridge molecule in Vibrio splendidus internalization. Its mRNA expression pattern, PAMP binding spectrum and microorganism binding capability were examined in the present study. The mRNA expression of CgEcSOD in hemocytes was significantly up-regulated at the initial phase and decreased sharply at 48 h post V. splendidus stimulation. The recombinant CgEcSOD protein (rCgEcSOD) could bind LPS, PGN and poly (I:C), as well as various microorganisms including Micrococcus luteus, Staphylococcus aureus, Escherichia coli, Vibrio anguillarum, V. splendidus, Pastoris pastoris and Yarrowia lipolytica at the presence of divalent metal ions Cu(2+). After the secondary V. splendidus stimulation, the mRNA and protein of CgEcSOD were both down-regulated significantly. The results collectively indicated that CgEcSOD could not only function in the immune recognition, but also might contribute to the immune priming of oyster by inhibiting the foreign microbe invasion through a specific down-regulation. Copyright © 2016 Elsevier Ltd. All rights reserved.

The secretory pathway: exploring yeast diversity.

PubMed

Delic, Marizela; Valli, Minoska; Graf, Alexandra B; Pfeffer, Martin; Mattanovich, Diethard; Gasser, Brigitte

2013-11-01

Protein secretion is an essential process for living organisms. In eukaryotes, this encompasses numerous steps mediated by several hundred cellular proteins. The core functions of translocation through the endoplasmic reticulum membrane, primary glycosylation, folding and quality control, and vesicle-mediated secretion are similar from yeasts to higher eukaryotes. However, recent research has revealed significant functional differences between yeasts and mammalian cells, and even among diverse yeast species. This review provides a current overview of the canonical protein secretion pathway in the model yeast Saccharomyces cerevisiae, highlighting differences to mammalian cells as well as currently unresolved questions, and provides a genomic comparison of the S. cerevisiae pathway to seven other yeast species where secretion has been investigated due to their attraction as protein production platforms, or for their relevance as pathogens. The analysis of Candida albicans, Candida glabrata, Kluyveromyces lactis, Pichia pastoris, Hansenula polymorpha, Yarrowia lipolytica, and Schizosaccharomyces pombe reveals that many - but not all - secretion steps are more redundant in S. cerevisiae due to duplicated genes, while some processes are even absent in this model yeast. Recent research obviates that even where homologous genes are present, small differences in protein sequence and/or differences in the regulation of gene expression may lead to quite different protein secretion phenotypes. © 2013 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

13C Metabolic Flux Analysis for Systematic Metabolic Engineering of S. cerevisiae for Overproduction of Fatty Acids

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ghosh, Amit; Ando, David; Gin, Jennifer

Efficient redirection of microbial metabolism into the abundant production of desired bioproducts remains non-trivial. Here, we used flux-based modeling approaches to improve yields of fatty acids in Saccharomyces cerevisiae. We combined 13C labeling data with comprehensive genome-scale models to shed light onto microbial metabolism and improve metabolic engineering efforts. We concentrated on studying the balance of acetyl-CoA, a precursor metabolite for the biosynthesis of fatty acids. A genome-wide acetyl-CoA balance study showed ATP citrate lyase from Yarrowia lipolytica as a robust source of cytoplasmic acetyl-CoA and malate synthase as a desirable target for downregulation in terms of acetyl-CoA consumption. Thesemore » genetic modifications were applied to S. cerevisiae WRY2, a strain that is capable of producing 460 mg/L of free fatty acids. With the addition of ATP citrate lyase and downregulation of malate synthase, the engineered strain produced 26% more free fatty acids. Further increases in free fatty acid production of 33% were obtained by knocking out the cytoplasmic glycerol-3-phosphate dehydrogenase, which flux analysis had shown was competing for carbon flux upstream with the carbon flux through the acetyl-CoA production pathway in the cytoplasm. In total, the genetic interventions applied in this work increased fatty acid production by ~70%.« less

13C Metabolic Flux Analysis for Systematic Metabolic Engineering of S. cerevisiae for Overproduction of Fatty Acids

DOE PAGES

Ghosh, Amit; Ando, David; Gin, Jennifer; ...

2016-10-05

Efficient redirection of microbial metabolism into the abundant production of desired bioproducts remains non-trivial. Here, we used flux-based modeling approaches to improve yields of fatty acids in Saccharomyces cerevisiae. We combined 13C labeling data with comprehensive genome-scale models to shed light onto microbial metabolism and improve metabolic engineering efforts. We concentrated on studying the balance of acetyl-CoA, a precursor metabolite for the biosynthesis of fatty acids. A genome-wide acetyl-CoA balance study showed ATP citrate lyase from Yarrowia lipolytica as a robust source of cytoplasmic acetyl-CoA and malate synthase as a desirable target for downregulation in terms of acetyl-CoA consumption. Thesemore » genetic modifications were applied to S. cerevisiae WRY2, a strain that is capable of producing 460 mg/L of free fatty acids. With the addition of ATP citrate lyase and downregulation of malate synthase, the engineered strain produced 26% more free fatty acids. Further increases in free fatty acid production of 33% were obtained by knocking out the cytoplasmic glycerol-3-phosphate dehydrogenase, which flux analysis had shown was competing for carbon flux upstream with the carbon flux through the acetyl-CoA production pathway in the cytoplasm. In total, the genetic interventions applied in this work increased fatty acid production by ~70%.« less

Safety assessment of EPA-rich triglyceride oil produced from yeast: genotoxicity and 28-day oral toxicity in rats.

PubMed

Belcher, Leigh A; MacKenzie, Susan A; Donner, Maria; Sykes, Greg P; Frame, Steven R; Gillies, Peter J

2011-02-01

The 28-day repeat-dose oral and genetic toxicity of eicosapentaenoic acid triglyceride oil (EPA oil) produced from genetically modified Yarrowia lipolytica yeast were assessed. Groups of rats received 0 (olive oil), 940, 1880, or 2820 mg EPA oil/kg/day, or fish oil (sardine/anchovy source) by oral gavage. Lower total serum cholesterol was seen in all EPA and fish oil groups. Liver weights were increased in the medium and high-dose EPA (male only), and fish oil groups but were considered non-adverse physiologically adaptive responses. Increased thyroid follicular cell hypertrophy was observed in male high-dose EPA and fish oil groups, and was considered to be an adaptive response to high levels of polyunsaturated fatty acids. No adverse test substance-related effects were observed on body weight, nutritional, or other clinical or anatomic pathology parameters. The oil was not mutagenic in the in vitro Ames or mouse lymphoma assay, and was not clastogenic in the in vivo mouse micronucleus test. In conclusion, exposure for 28 days to EPA oil derived from yeast did not produce adverse effects at doses up to 2820 mg/kg/day and was not genotoxic. The safety profile of the EPA oil in these tests was comparable to a commercial fish oil. Copyright © 2010 Elsevier Inc. All rights reserved.

Application of metabolic controls for the maximization of lipid production in semicontinuous fermentation.

PubMed

Xu, Jingyang; Liu, Nian; Qiao, Kangjian; Vogg, Sebastian; Stephanopoulos, Gregory

2017-07-03

Acetic acid can be generated through syngas fermentation, lignocellulosic biomass degradation, and organic waste anaerobic digestion. Microbial conversion of acetate into triacylglycerols for biofuel production has many advantages, including low-cost or even negative-cost feedstock and environmental benefits. The main issue stems from the dilute nature of acetate produced in such systems, which is costly to be processed on an industrial scale. To tackle this problem, we established an efficient bioprocess for converting dilute acetate into lipids, using the oleaginous yeast Yarrowia lipolytica in a semicontinuous system. The implemented design used low-strength acetic acid in both salt and acid forms as carbon substrate and a cross-filtration module for cell recycling. Feed controls for acetic acid and nitrogen based on metabolic models and online measurement of the respiratory quotient were used. The optimized process was able to sustain high-density cell culture using acetic acid of only 3% and achieved a lipid titer, yield, and productivity of 115 g/L, 0.16 g/g, and 0.8 g⋅L -1 ⋅h -1 , respectively. No carbon substrate was detected in the effluent stream, indicating complete utilization of acetate. These results represent a more than twofold increase in lipid production metrics compared with the current best-performing results using concentrated acetic acid as carbon feed.

Assessment of Antimicrobial and Antioxidant Activities of Nepeta trachonitica: Analysis of Its Phenolic Compounds Using HPLC-MS/MS

PubMed Central

Köksal, Ekrem; Tohma, Hatice; Kılıç, Ömer; Alan, Yusuf; Aras, Abdülmelik; Gülçin, İlhami; Bursal, Ercan

2017-01-01

Continuing our work on the sources of natural bioactive compounds, we evaluated the antimicrobial and antioxidant activities of Nepeta trachonitica as well as its major phenolic content using the high-performance liquid chromatography-mass spectrometry/mass spectrometry (HPLC-MS/MS) technique. For antioxidant activity, ferric reducing antioxidant power (FRAP) and cupric ion reducing antioxidant capacity (CUPRAC) methods were performed to measure the reducing power and 1,1-diphenyl-2-picrylhydrazyl (DPPH) assay was employed to evaluate the radical scavenging activity of the sample. For antimicrobial activity, three Gram-positive and four Gram-negative microbial species as well as three fungi species were tested. N. trachonitica appeared to have reasonable antioxidant activity and decent antimicrobial activity as indicated by the inhibition of the organisms’ growth. The most susceptible species were Bacillus subtilis ATCC 6633 and Escherichia coli ATCC 11229 among the organisms tested. Ethanol extract of the plant has the highest effect on Saccharomyces cerevisiae but no effect on Yarrowia lipolytica. The HPLC-MS/MS analysis showed that at least 11 major phenolic compounds of N. trachonitica exist, the major ones being rosmarinic acid, chlorogenic acid and quinic acid. The obtained results suggest that N. trachonitica could be a promising source for food and nutraceutical industries because of its antimicrobial and antioxidant properties and phenolic compounds. PMID:28505129

A surprisingly large RNase P RNA in Candida glabrata

PubMed Central

KACHOURI, RYM; STRIBINSKIS, VILIUS; ZHU, YANGLONG; RAMOS, KENNETH S.; WESTHOF, ERIC; LI, YONG

2005-01-01

We have found an extremely large ribonuclease P (RNase P) RNA (RPR1) in the human pathogen Candida glabrata and verified that this molecule is expressed and present in the active enzyme complex of this hemiascomycete yeast. A structural alignment of the C. glabrata sequence with 36 other hemiascomycete RNase P RNAs (abbreviated as P RNAs) allows us to characterize the types of insertions. In addition, 15 P RNA sequences were newly characterized by searching in the recently sequenced genomes Candida albicans, C. glabrata, Debaryomyces hansenii, Eremothecium gossypii, Kluyveromyces lactis, Kluyveromyces waltii, Naumovia castellii, Saccharomyces kudriavzevii, Saccharomyces mikatae, and Yarrowia lipolytica; and by PCR amplification for other Candida species (Candida guilliermondii, Candida krusei, Candida parapsilosis, Candida stellatoidea, and Candida tropicalis). The phylogenetic comparative analysis identifies a hemiascomycete secondary structure consensus that presents a conserved core in all species with variable insertions or deletions. The most significant variability is found in C. glabrata P RNA in which three insertions exceeding in total 700 nt are present in the Specificity domain. This P RNA is more than twice the length of any other homologous P RNAs known in the three domains of life and is eight times the size of the smallest. RNase P RNA, therefore, represents one of the most diversified noncoding RNAs in terms of size variation and structural diversity. PMID:15987816

Changes in volatile profile of soybean residue (okara) upon solid-state fermentation by yeasts.

PubMed

Vong, Weng Chan; Liu, Shao-Quan

2017-01-01

Soybean residue (okara), a by-product of soymilk, is produced in large volumes by the soy food industry and is often discarded due to its undesirable flavour. As it contains a considerable amount of protein and fats, biotransformation of okara to improve its flavour presents an opportunity for alternative utilisation. This paper evaluated 10 yeasts in the solid-state fermentation of okara based on their volatile profiles as analysed with HS-SPME GC-MS/FID. Four 'dairy yeasts' (Geotrichum candidum, Yarrowia lipolytica, Debaryomyces hansenii and Kluyveromyces lactis) and six 'wine yeasts' (Saccharomyces cerevisiae, Lachancea thermotolerans, Metschnikowia pulcherrima, Pichia kluyveri, Torulaspora delbrueckii, and Williopsis saturnus) were studied. The main off-odourants in okara, hexanal and trans-2-hexenal, significantly decreased after fermentation due to their bioconversion into methyl ketones and/or esters. The okara fermented by dairy yeasts contained greater proportions of methyl ketones, while that by wine yeasts contained more ethyl and acetyl esters. Notably, the okara fermented by W. saturnus contained 13 esters and the total GC-FID peak area of esters was about 380 times that in fresh okara, leading to a perceptible fruity note. Okara can be exploited as an inexpensive substrate for bioflavour extraction and/or a more pleasant food ingredient via yeast fermentation. © 2016 Society of Chemical Industry. © 2016 Society of Chemical Industry.

The RNA polymerase III-dependent family of genes in hemiascomycetes: comparative RNomics, decoding strategies, transcription and evolutionary implications

PubMed Central

Marck, Christian; Kachouri-Lafond, Rym; Lafontaine, Ingrid; Westhof, Eric; Dujon, Bernard; Grosjean, Henri

2006-01-01

We present the first comprehensive analysis of RNA polymerase III (Pol III) transcribed genes in ten yeast genomes. This set includes all tRNA genes (tDNA) and genes coding for SNR6 (U6), SNR52, SCR1 and RPR1 RNA in the nine hemiascomycetes Saccharomyces cerevisiae, Saccharomyces castellii, Candida glabrata, Kluyveromyces waltii, Kluyveromyces lactis, Eremothecium gossypii, Debaryomyces hansenii, Candida albicans, Yarrowia lipolytica and the archiascomycete Schizosaccharomyces pombe. We systematically analysed sequence specificities of tRNA genes, polymorphism, variability of introns, gene redundancy and gene clustering. Analysis of decoding strategies showed that yeasts close to S.cerevisiae use bacterial decoding rules to read the Leu CUN and Arg CGN codons, in contrast to all other known Eukaryotes. In D.hansenii and C.albicans, we identified a novel tDNA-Leu (AAG), reading the Leu CUU/CUC/CUA codons with an unusual G at position 32. A systematic ‘p-distance tree’ using the 60 variable positions of the tRNA molecule revealed that most tDNAs cluster into amino acid-specific sub-trees, suggesting that, within hemiascomycetes, orthologous tDNAs are more closely related than paralogs. We finally determined the bipartite A- and B-box sequences recognized by TFIIIC. These minimal sequences are nearly conserved throughout hemiascomycetes and were satisfactorily retrieved at appropriate locations in other Pol III genes. PMID:16600899

Nanoparticles as biochemical sensors

PubMed Central

El-Ansary, Afaf; Faddah, Layla M

2010-01-01

There is little doubt that nanoparticles offer real and new opportunities in many fields, such as biomedicine and materials science. Such particles are small enough to enter almost all areas of the body, including cells and organelles, potentially leading to new approaches in nanomedicine. Sensors for small molecules of biochemical interest are of critical importance. This review is an attempt to trace the use of nanomaterials in biochemical sensor design. The possibility of using nanoparticles functionalized with antibodies as markers for proteins will be elucidated. Moreover, capabilities and applications for nanoparticles based on gold, silver, magnetic, and semiconductor materials (quantum dots), used in optical (absorbance, luminescence, surface enhanced Raman spectroscopy, surface plasmon resonance), electrochemical, and mass-sensitive sensors will be highlighted. The unique ability of nanosensors to improve the analysis of biochemical fluids is discussed either through considering the use of nanoparticles for in vitro molecular diagnosis, or in the biological/biochemical analysis for in vivo interaction with the human body. PMID:24198472

Misleading biochemical laboratory test results

PubMed Central

Nanji, Amin A.

1984-01-01

This article reviews the general and specific factors that interfere with the performance of common biochemical laboratory tests and the interpretation of their results. The clinical status of the patient, drug interactions, and in-vivo and in-vitro biochemical interactions and changes may alter the results obtained from biochemical analysis of blood constituents. Failure to recognize invalid laboratory test results may lead to injudicious and dangerous management of patients. PMID:6375845

Biochemical transformation of coals

DOEpatents

Lin, Mow S.; Premuzic, Eugene T.

1999-03-23

A method of biochemically transforming macromolecular compounds found in solid carbonaceous materials, such as coal is provided. The preparation of new microorganisms, metabolically weaned through challenge growth processes to biochemically transform solid carbonaceous materials at extreme temperatures, pressures, pH, salt and toxic metal concentrations is also disclosed.

Biochemical transformation of coals

DOEpatents

Lin, M.S.; Premuzic, E.T.

1999-03-23

A method of biochemically transforming macromolecular compounds found in solid carbonaceous materials, such as coal is provided. The preparation of new microorganisms, metabolically weaned through challenge growth processes to biochemically transform solid carbonaceous materials at extreme temperatures, pressures, pH, salt and toxic metal concentrations is also disclosed. 7 figs.

Biochemical-Pathway Diversity in Archaebacteria

DTIC Science & Technology

1990-08-30

Classification) (U) Biochemical-pathway diversity in Archaebacteria 12 PERSONAL AUTHOR(S) I Jensen, Roy-A. i3o. TYPE OF REN" RT 12b. Tki~ 0’E D-30-9 4...by block numtb.sj FIEL I ROU I SIGRLJP Archaebacteria , biochemical diversity, prephenate 06 03. 1 dehydratase, aromatic amino acid biosynthesis t...1988 RE10SE: lo assess the extent to which the archaebacteria possess unique biochemical features of aromatic amino acid biosynthesis and regulation and

Biochemical abnormalities in neonatal seizures.

PubMed

Sood, Arvind; Grover, Neelam; Sharma, Roshan

2003-03-01

The presence of seizure does not constitute a diagnoses but it is a symptom of an underlying central nervous system disorder due to systemic or biochemical disturbances. Biochemical disturbances occur frequently in the neonatal seizures either as an underlying cause or as an associated abnormality. In their presence, it is difficult to control seizure and there is a risk of further brain damage. Early recognition and treatment of biochemical disturbances is essential for optimal management and satisfactory long term outcome. The present study was conducted in the department of pediatrics in IGMC Shimla on 59 neonates. Biochemical abnormalities were detected in 29 (49.15%) of cases. Primary metabolic abnormalities occurred in 10(16.94%) cases of neonatal seizures, most common being hypocalcaemia followed by hypoglycemia, other metabolic abnormalities include hypomagnesaemia and hyponateremia. Biochemical abnormalities were seen in 19(38.77%) cases of non metabolic seizure in neonates. Associated metabolic abnormalities were observed more often with Hypoxic-ischemic-encephalopathy (11 out of 19) cases and hypoglycemia was most common in this group. No infant had hyponateremia, hyperkelemia or low zinc level.

Novel efficient promoter of the mitochondrial porin, voltage-dependent anion channel (VDAC), in the genome of the Yarrowia lipolytica yeast.

PubMed

Kulanbaewa, F F; Sekova, V Yu; Isakova, E P; Deryabina, Y I; Nikolaev, A V

2016-09-01

This article presents the characteristics of the highly inducible promoter of the gene encoding the mitochondrial porin, the voltage-dependent anion channel (VDAC). This promoter is recommended for use in new genetic constructs both in basic research for assessing the adaptive strategy of lower eukaryotes under adverse conditions and in designing new highly competitive transformants producing economically important compounds (proteins, lipids, and organic acids) on its basis.

Expression and secretion of fungal endoglucanase II and chimeric cellobiohydrolase I in the oleaginous yeast Lipomyces starkeyi

DOE PAGES

Xu, Qi; Knoshaug, Eric P.; Wang, Wei; ...

2017-07-24

Lipomyces starkeyi is one of the leading lipid-producing microorganisms reported to date; its genetic transformation was only recently reported. Our aim is to engineer L. starkeyi to serve in consolidated bioprocessing (CBP) to produce lipid or fatty acid-related biofuels directly from abundant and low-cost lignocellulosic substrates. To evaluate L. starkeyi in this role, we first conducted a genome analysis, which revealed the absence of key endo- and exocellulases in this yeast, prompting us to select and screen four signal peptides for their suitability for the overexpression and secretion of cellulase genes. To compensate for the cellulase deficiency, we chose twomore » prominent cellulases, Trichoderma reesei endoglucanase II (EG II) and a chimeric cellobiohydrolase I (TeTrCBH I) formed by fusion of the catalytic domain from Talaromyces emersonii CBH I with the linker peptide and cellulose-binding domain from T. reesei CBH I. The systematically tested signal peptides included three peptides from native L. starkeyi and one from Yarrowia lipolytica. We found that all four signal peptides permitted secretion of active EG II. We also determined that three of these signal peptides worked for expression of the chimeric CBH I; suggesting that our design criteria for selecting these signal peptides was effective. Encouragingly, the Y. lipolytica signal peptide was able to efficiently guide secretion of the chimeric TeTrCBH I protein from L. starkeyi. The purified chimeric TeTrCBH I showed high activity against the cellulose in pretreated corn stover and the purified EG II showed high endocellulase activity measured by the CELLG3 (Megazyme) method. Our results suggest that L. starkeyi is capable of expressing and secreting core fungal cellulases. Moreover, the purified EG II and chimeric TeTrCBH I displayed significant and potentially useful enzymatic activities, demonstrating that engineered L. starkeyi has the potential to function as an oleaginous CBP strain for

Expression and secretion of fungal endoglucanase II and chimeric cellobiohydrolase I in the oleaginous yeast Lipomyces starkeyi

DOE Office of Scientific and Technical Information (OSTI.GOV)

Xu, Qi; Knoshaug, Eric P.; Wang, Wei

Lipomyces starkeyi is one of the leading lipid-producing microorganisms reported to date; its genetic transformation was only recently reported. Our aim is to engineer L. starkeyi to serve in consolidated bioprocessing (CBP) to produce lipid or fatty acid-related biofuels directly from abundant and low-cost lignocellulosic substrates. To evaluate L. starkeyi in this role, we first conducted a genome analysis, which revealed the absence of key endo- and exocellulases in this yeast, prompting us to select and screen four signal peptides for their suitability for the overexpression and secretion of cellulase genes. To compensate for the cellulase deficiency, we chose twomore » prominent cellulases, Trichoderma reesei endoglucanase II (EG II) and a chimeric cellobiohydrolase I (TeTrCBH I) formed by fusion of the catalytic domain from Talaromyces emersonii CBH I with the linker peptide and cellulose-binding domain from T. reesei CBH I. The systematically tested signal peptides included three peptides from native L. starkeyi and one from Yarrowia lipolytica. We found that all four signal peptides permitted secretion of active EG II. We also determined that three of these signal peptides worked for expression of the chimeric CBH I; suggesting that our design criteria for selecting these signal peptides was effective. Encouragingly, the Y. lipolytica signal peptide was able to efficiently guide secretion of the chimeric TeTrCBH I protein from L. starkeyi. The purified chimeric TeTrCBH I showed high activity against the cellulose in pretreated corn stover and the purified EG II showed high endocellulase activity measured by the CELLG3 (Megazyme) method. Our results suggest that L. starkeyi is capable of expressing and secreting core fungal cellulases. Moreover, the purified EG II and chimeric TeTrCBH I displayed significant and potentially useful enzymatic activities, demonstrating that engineered L. starkeyi has the potential to function as an oleaginous CBP strain for

Chemical signaling and insect attraction is a conserved trait in yeasts.

PubMed

Becher, Paul G; Hagman, Arne; Verschut, Vasiliki; Chakraborty, Amrita; Rozpędowska, Elżbieta; Lebreton, Sébastien; Bengtsson, Marie; Flick, Gerhard; Witzgall, Peter; Piškur, Jure

2018-03-01

Yeast volatiles attract insects, which apparently is of mutual benefit, for both yeasts and insects. However, it is unknown whether biosynthesis of metabolites that attract insects is a basic and general trait, or if it is specific for yeasts that live in close association with insects. Our goal was to study chemical insect attractants produced by yeasts that span more than 250 million years of evolutionary history and vastly differ in their metabolism and lifestyle. We bioassayed attraction of the vinegar fly Drosophila melanogaster to odors of phylogenetically and ecologically distinct yeasts grown under controlled conditions. Baker's yeast Saccharomyces cerevisiae , the insect-associated species Candida californica , Pichia kluyveri and Metschnikowia andauensis , wine yeast Dekkera bruxellensis , milk yeast Kluyveromyces lactis , the vertebrate pathogens Candida albicans and Candida glabrata , and oleophilic Yarrowia lipolytica were screened for fly attraction in a wind tunnel. Yeast headspace was chemically analyzed, and co-occurrence of insect attractants in yeasts and flowering plants was investigated through a database search. In yeasts with known genomes, we investigated the occurrence of genes involved in the synthesis of key aroma compounds. Flies were attracted to all nine yeasts studied. The behavioral response to baker's yeast was independent of its growth stage. In addition to Drosophila , we tested the basal hexapod Folsomia candida (Collembola) in a Y-tube assay to the most ancient yeast, Y. lipolytica, which proved that early yeast signals also function on clades older than neopteran insects. Behavioral and chemical data and a search for selected genes of volatile metabolites underline that biosynthesis of chemical signals is found throughout the yeast clade and has been conserved during the evolution of yeast lifestyles. Literature and database reviews corroborate that yeast signals mediate mutualistic interactions between insects and yeasts

Expression and secretion of fungal endoglucanase II and chimeric cellobiohydrolase I in the oleaginous yeast Lipomyces starkeyi.

PubMed

Xu, Qi; Knoshaug, Eric P; Wang, Wei; Alahuhta, Markus; Baker, John O; Yang, Shihui; Vander Wall, Todd; Decker, Stephen R; Himmel, Michael E; Zhang, Min; Wei, Hui

2017-07-24

Lipomyces starkeyi is one of the leading lipid-producing microorganisms reported to date; its genetic transformation was only recently reported. Our aim is to engineer L. starkeyi to serve in consolidated bioprocessing (CBP) to produce lipid or fatty acid-related biofuels directly from abundant and low-cost lignocellulosic substrates. To evaluate L. starkeyi in this role, we first conducted a genome analysis, which revealed the absence of key endo- and exocellulases in this yeast, prompting us to select and screen four signal peptides for their suitability for the overexpression and secretion of cellulase genes. To compensate for the cellulase deficiency, we chose two prominent cellulases, Trichoderma reesei endoglucanase II (EG II) and a chimeric cellobiohydrolase I (TeTrCBH I) formed by fusion of the catalytic domain from Talaromyces emersonii CBH I with the linker peptide and cellulose-binding domain from T. reesei CBH I. The systematically tested signal peptides included three peptides from native L. starkeyi and one from Yarrowia lipolytica. We found that all four signal peptides permitted secretion of active EG II. We also determined that three of these signal peptides worked for expression of the chimeric CBH I; suggesting that our design criteria for selecting these signal peptides was effective. Encouragingly, the Y. lipolytica signal peptide was able to efficiently guide secretion of the chimeric TeTrCBH I protein from L. starkeyi. The purified chimeric TeTrCBH I showed high activity against the cellulose in pretreated corn stover and the purified EG II showed high endocellulase activity measured by the CELLG3 (Megazyme) method. Our results suggest that L. starkeyi is capable of expressing and secreting core fungal cellulases. Moreover, the purified EG II and chimeric TeTrCBH I displayed significant and potentially useful enzymatic activities, demonstrating that engineered L. starkeyi has the potential to function as an oleaginous CBP strain for biofuel

40 CFR 158.2010 - Biochemical pesticides data requirements.

Code of Federal Regulations, 2012 CFR

2012-07-01

... conditions are identified within the test notes. Definitions that apply to all biochemical data requirements... 40 Protection of Environment 25 2012-07-01 2012-07-01 false Biochemical pesticides data...) PESTICIDE PROGRAMS DATA REQUIREMENTS FOR PESTICIDES Biochemical Pesticides § 158.2010 Biochemical pesticides...

40 CFR 158.2010 - Biochemical pesticides data requirements.

Code of Federal Regulations, 2013 CFR

2013-07-01

... conditions are identified within the test notes. Definitions that apply to all biochemical data requirements... 40 Protection of Environment 25 2013-07-01 2013-07-01 false Biochemical pesticides data...) PESTICIDE PROGRAMS DATA REQUIREMENTS FOR PESTICIDES Biochemical Pesticides § 158.2010 Biochemical pesticides...

40 CFR 158.2010 - Biochemical pesticides data requirements.

Code of Federal Regulations, 2014 CFR

2014-07-01

... conditions are identified within the test notes. Definitions that apply to all biochemical data requirements... 40 Protection of Environment 24 2014-07-01 2014-07-01 false Biochemical pesticides data...) PESTICIDE PROGRAMS DATA REQUIREMENTS FOR PESTICIDES Biochemical Pesticides § 158.2010 Biochemical pesticides...

40 CFR 158.2010 - Biochemical pesticides data requirements.

Code of Federal Regulations, 2010 CFR

2010-07-01

... conditions are identified within the test notes. Definitions that apply to all biochemical data requirements... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Biochemical pesticides data...) PESTICIDE PROGRAMS DATA REQUIREMENTS FOR PESTICIDES Biochemical Pesticides § 158.2010 Biochemical pesticides...

40 CFR 158.2010 - Biochemical pesticides data requirements.

Code of Federal Regulations, 2011 CFR

2011-07-01

... conditions are identified within the test notes. Definitions that apply to all biochemical data requirements... 40 Protection of Environment 24 2011-07-01 2011-07-01 false Biochemical pesticides data...) PESTICIDE PROGRAMS DATA REQUIREMENTS FOR PESTICIDES Biochemical Pesticides § 158.2010 Biochemical pesticides...

Biochemical Network Stochastic Simulator (BioNetS): software for stochastic modeling of biochemical networks.

PubMed

Adalsteinsson, David; McMillen, David; Elston, Timothy C

2004-03-08

Intrinsic fluctuations due to the stochastic nature of biochemical reactions can have large effects on the response of biochemical networks. This is particularly true for pathways that involve transcriptional regulation, where generally there are two copies of each gene and the number of messenger RNA (mRNA) molecules can be small. Therefore, there is a need for computational tools for developing and investigating stochastic models of biochemical networks. We have developed the software package Biochemical Network Stochastic Simulator (BioNetS) for efficiently and accurately simulating stochastic models of biochemical networks. BioNetS has a graphical user interface that allows models to be entered in a straightforward manner, and allows the user to specify the type of random variable (discrete or continuous) for each chemical species in the network. The discrete variables are simulated using an efficient implementation of the Gillespie algorithm. For the continuous random variables, BioNetS constructs and numerically solves the appropriate chemical Langevin equations. The software package has been developed to scale efficiently with network size, thereby allowing large systems to be studied. BioNetS runs as a BioSpice agent and can be downloaded from http://www.biospice.org. BioNetS also can be run as a stand alone package. All the required files are accessible from http://x.amath.unc.edu/BioNetS. We have developed BioNetS to be a reliable tool for studying the stochastic dynamics of large biochemical networks. Important features of BioNetS are its ability to handle hybrid models that consist of both continuous and discrete random variables and its ability to model cell growth and division. We have verified the accuracy and efficiency of the numerical methods by considering several test systems.

The underlying pathway structure of biochemical reaction networks

PubMed Central

Schilling, Christophe H.; Palsson, Bernhard O.

1998-01-01

Bioinformatics is yielding extensive, and in some cases complete, genetic and biochemical information about individual cell types and cellular processes, providing the composition of living cells and the molecular structure of its components. These components together perform integrated cellular functions that now need to be analyzed. In particular, the functional definition of biochemical pathways and their role in the context of the whole cell is lacking. In this study, we show how the mass balance constraints that govern the function of biochemical reaction networks lead to the translation of this problem into the realm of linear algebra. The functional capabilities of biochemical reaction networks, and thus the choices that cells can make, are reflected in the null space of their stoichiometric matrix. The null space is spanned by a finite number of basis vectors. We present an algorithm for the synthesis of a set of basis vectors for spanning the null space of the stoichiometric matrix, in which these basis vectors represent the underlying biochemical pathways that are fundamental to the corresponding biochemical reaction network. In other words, all possible flux distributions achievable by a defined set of biochemical reactions are represented by a linear combination of these basis pathways. These basis pathways thus represent the underlying pathway structure of the defined biochemical reaction network. This development is significant from a fundamental and conceptual standpoint because it yields a holistic definition of biochemical pathways in contrast to definitions that have arisen from the historical development of our knowledge about biochemical processes. Additionally, this new conceptual framework will be important in defining, characterizing, and studying biochemical pathways from the rapidly growing information on cellular function. PMID:9539712

Cocoa butter-like lipid production ability of non-oleaginous and oleaginous yeasts under nitrogen-limited culture conditions.

PubMed

Wei, Yongjun; Siewers, Verena; Nielsen, Jens

2017-05-01

Cocoa butter (CB) extracted from cocoa beans is the main raw material for chocolate production. However, growing chocolate demands and limited CB production has resulted in a shortage of CB supply. CB is mainly composed of three different kinds of triacylglycerols (TAGs), POP (C16:0-C18:1-C16:0), POS (C16:0-C18:1-C18:0), and SOS (C18:0-C18:1-C18:0). The storage lipids of yeasts, mainly TAGs, also contain relative high-level of C16 and C18 fatty acids and might be used as CB-like lipids (CBL). In this study, we cultivated six different yeasts, including one non-oleaginous yeast strain, Saccharomyces cerevisiae CEN.PK113-7D, and five oleaginous yeast strains, Trichosporon oleaginosus DSM11815, Rhodotorula graminis DSM 27356, Lipomyces starkeyi DSM 70296, Rhodosporidium toruloides DSM 70398, and Yarrowia lipolytica CBS 6124, in nitrogen-limited medium and compared their CBL production ability. Under the same growth conditions, we found that TAGs were the main lipids in all six yeasts and that T. oleaginosus can produce more TAGs than the other five yeasts. Less than 3% of the total TAGs were identified as potential SOS in the six yeasts. However, T. oleaginosus produced 27.8% potential POP and POS at levels of 378 mg TAGs/g dry cell weight, hinting that this yeast may have potential as a CBL production host after further metabolic engineering in future.

Role of charge screening and delocalization for lipophilic cation permeability of model and mitochondrial membranes.

PubMed

Trendeleva, Tatiana A; Sukhanova, Evgenia I; Rogov, Anton G; Zvyagilskaya, Renata A; Seveina, Inna I; Ilyasova, Tatiana M; Cherepanov, Dmitry A; Skulachev, Vladimir P

2013-09-01

The effects of the mitochondria-targeted lipophilic cation dodecyltriphenylphosphonium (C12TPP, the charge is delocalized and screened by bulky hydrophobic residues) and those of lipophilic cations decyltriethylammonium bromide and cetyltrimethylammonium bromide (C10TEA and C16TMA, the charges are localized and screened by less bulky residues) on bilayer planar phospholipid membranes and tightly-coupled mitochondria from the yeast Yarrowia lipolytica have been compared. In planar membranes, C12TPP was found to generate a diffusion potential as if it easily penetrates these membranes. In the presence of palmitate, C12TPP induced H(+) permeability like plastoquinonyl decyltriphenilphosphonium that facilitates transfer of fatty acid anions (Severin et al., PNAS, 2010, 107, 663-668). C12TPP was shown to stimulate State 4 respiration of mitochondria and caused a mitochondrial membrane depolarization with a half-maximal effect at 6μM. Besides, C12TPP profoundly potentiated the uncoupling effect of endogenous or added fatty acids. C10TEA and C16TMA inhibited State 4 respiration and decreased the membrane potential, though at much higher concentrations than C12TPP, and they did not promote the uncoupling action of fatty acids. These relationships were modeled by molecular dynamics. They can be explained by different membrane permeabilities for studied cations, which in turn are due to different availabilities of the positive charge in these cations to water dipoles. Copyright © 2012 Elsevier B.V. All rights reserved.

Multi-wavelength dye concentration determination for enzymatic assays: evaluation of chromogenic para-nitrophenol over a wide pH range.

PubMed

Max, Jean-Joseph; Meddeb-Mouelhi, Fatma; Beauregard, Marc; Chapados, Camille

2012-12-01

Enzymatic assays need robust, rapid colorimetric methods that can follow ongoing reactions. For this, we developed a highly accurate, multi-wavelength detection method that could be used for several systems. Here, it was applied to the detection of para-nitrophenol (pNP) in basic and acidic solutions. First, we confirmed by factor analysis that pNP has two forms, with unique spectral characteristics in the 240 to 600 nm range: Phenol in acidic conditions absorbs in the lower range, whereas phenolate in basic conditions absorbs in the higher range. Thereafter, the method was used for the determination of species concentration. For this, the intensity measurements were made at only two wavelengths with a microtiter plate reader. This yielded total dye concentration, species relative abundance, and solution pH value. The method was applied to an enzymatic assay. For this, a chromogenic substrate that generates pNP after hydrolysis catalyzed by a lipase from the fungus Yarrowia lipolytica was used. Over the pH range of 3-11, accurate amounts of acidic and basic pNP were determined at 340 and 405 nm, respectively. This method surpasses the commonly used single-wavelength assay at 405 nm, which does not detect pNP acidic species, leading to activity underestimations. Moreover, alleviation of this pH-related problem by neutralization is not necessary. On the whole, the method developed is readily applicable to rapid high-throughput of enzymatic activity measurements over a wide pH range.

Gene cloning and characterization of a novel esterase from activated sludge metagenome

PubMed Central

2009-01-01

A metagenomic library was prepared using pCC2FOS vector containing about 3.0 Gbp of community DNA from the microbial assemblage of activated sludge. Screening of a part of the un-amplified library resulted in the finding of 1 unique lipolytic clone capable of hydrolyzing tributyrin, in which an esterase gene was identified. This esterase/lipase gene consists of 834 bp and encodes a polypeptide (designated EstAS) of 277 amino acid residuals with a molecular mass of 31 kDa. Sequence analysis indicated that it showed 33% and 31% amino acid identity to esterase/lipase from Gemmata obscuriglobus UQM 2246 (ZP_02733109) and Yarrowia lipolytica CLIB122 (XP_504639), respectively; and several conserved regions were identified, including the putative active site, HSMGG, a catalytic triad (Ser92, His125 and Asp216) and a LHYFRG conserved motif. The EstAS was overexpressed, purified and shown to hydrolyse p-nitrophenyl (NP) esters of fatty acids with short chain lengths (≤ C8). This EstAS had optimal temperature and pH at 35°C and 9.0, respectively, by hydrolysis of p-NP hexanoate. It also exhibited the same level of stability over wide temperature and pH ranges and in the presence of metal ions or detergents. The high level of stability of esterase EstAS with its unique substrate specificities make itself highly useful for biotechnological applications. PMID:20028524

Surface Microflora of Four Smear-Ripened Cheeses

PubMed Central

Mounier, Jérôme; Gelsomino, Roberto; Goerges, Stefanie; Vancanneyt, Marc; Vandemeulebroecke, Katrien; Hoste, Bart; Scherer, Siegfried; Swings, Jean; Fitzgerald, Gerald F.; Cogan, Timothy M.

2005-01-01

The microbial composition of smear-ripened cheeses is not very clear. A total of 194 bacterial isolates and 187 yeast isolates from the surfaces of four Irish farmhouse smear-ripened cheeses were identified at the midpoint of ripening using pulsed-field gel electrophoresis (PFGE), repetitive sequence-based PCR, and 16S rRNA gene sequencing for identifying and typing the bacteria and Fourier transform infrared spectroscopy and mitochondrial DNA restriction fragment length polymorphism (mtDNA RFLP) analysis for identifying and typing the yeast. The yeast microflora was very uniform, and Debaryomyces hansenii was the dominant species in the four cheeses. Yarrowia lipolytica was also isolated in low numbers from one cheese. The bacteria were highly diverse, and 14 different species, Corynebacterium casei, Corynebacterium variabile, Arthrobacter arilaitensis, Arthrobacter sp., Microbacterium gubbeenense, Agrococcus sp. nov., Brevibacterium linens, Staphylococcus epidermidis, Staphylococcus equorum, Staphylococcus saprophyticus, Micrococcus luteus, Halomonas venusta, Vibrio sp., and Bacillus sp., were identified on the four cheeses. Each cheese had a more or less unique microflora with four to nine species on its surface. However, two bacteria, C. casei and A. arilaitensis, were found on each cheese. Diversity at the strain level was also observed, based on the different PFGE patterns and mtDNA RFLP profiles of the dominant bacterial and yeast species. None of the ripening cultures deliberately inoculated onto the surface were reisolated from the cheeses. This study confirms the importance of the adventitious, resident microflora in the ripening of smear cheeses. PMID:16269673

Synthetic biology for manufacturing chemicals: constraints drive the use of non-conventional microbial platforms.

PubMed

Czajka, Jeffrey; Wang, Qinhong; Wang, Yechun; Tang, Yinjie J

2017-10-01

Genetically modified microbes have had much industrial success producing protein-based products (such as antibodies and enzymes). However, engineering microbial workhorses for biomanufacturing of commodity compounds remains challenging. First, microbes cannot afford burdens with both overexpression of multiple enzymes and metabolite drainage for product synthesis. Second, synthetic circuits and introduced heterologous pathways are not yet as "robust and reliable" as native pathways due to hosts' innate regulations, especially under suboptimal fermentation conditions. Third, engineered enzymes may lack channeling capabilities for cascade-like transport of metabolites to overcome diffusion barriers or to avoid intermediate toxicity in the cytoplasmic environment. Fourth, moving engineered hosts from laboratory to industry is unreliable because genetic mutations and non-genetic cell-to-cell variations impair the large-scale fermentation outcomes. Therefore, synthetic biology strains often have unsatisfactory industrial performance (titer/yield/productivity). To overcome these problems, many different species are being explored for their metabolic strengths that can be leveraged to synthesize specific compounds. Here, we provide examples of non-conventional and genetically amenable species for industrial manufacturing, including the following: Corynebacterium glutamicum for its TCA cycle-derived biosynthesis, Yarrowia lipolytica for its biosynthesis of fatty acids and carotenoids, cyanobacteria for photosynthetic production from its sugar phosphate pathways, and Rhodococcus for its ability to biotransform recalcitrant feedstock. Finally, we discuss emerging technologies (e.g., genome-to-phenome mapping, single cell methods, and knowledge engineering) that may facilitate the development of novel cell factories.

Serum Biochemical Phenotypes in the Domestic Dog

PubMed Central

Chang, Yu-Mei; Hadox, Erin; Szladovits, Balazs; Garden, Oliver A.

2016-01-01

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. PMID:26919479

Improving Marine Ecosystem Models with Biochemical Tracers

NASA Astrophysics Data System (ADS)

Pethybridge, Heidi R.; Choy, C. Anela; Polovina, Jeffrey J.; Fulton, Elizabeth A.

2018-01-01

Empirical data on food web dynamics and predator-prey interactions underpin ecosystem models, which are increasingly used to support strategic management of marine resources. These data have traditionally derived from stomach content analysis, but new and complementary forms of ecological data are increasingly available from biochemical tracer techniques. Extensive opportunities exist to improve the empirical robustness of ecosystem models through the incorporation of biochemical tracer data and derived indices, an area that is rapidly expanding because of advances in analytical developments and sophisticated statistical techniques. Here, we explore the trophic information required by ecosystem model frameworks (species, individual, and size based) and match them to the most commonly used biochemical tracers (bulk tissue and compound-specific stable isotopes, fatty acids, and trace elements). Key quantitative parameters derived from biochemical tracers include estimates of diet composition, niche width, and trophic position. Biochemical tracers also provide powerful insight into the spatial and temporal variability of food web structure and the characterization of dominant basal and microbial food web groups. A major challenge in incorporating biochemical tracer data into ecosystem models is scale and data type mismatches, which can be overcome with greater knowledge exchange and numerical approaches that transform, integrate, and visualize data.

Biochemical Engineering Fundamentals

ERIC Educational Resources Information Center

Bailey, J. E.; Ollis, D. F.

1976-01-01

Discusses a biochemical engineering course that is offered as part of a chemical engineering curriculum and includes topics that influence the behavior of man-made or natural microbial or enzyme reactors. (MLH)

Biochemical transformation of solid carbonaceous material

DOEpatents

Lin, Mow S.; Premuzic, Eugene T.

2001-09-25

A method of biochemically transforming macromolecular compounds found in solid carbonaceous materials, such as coal is provided. The preparation of new microorganisms, metabolically weaned through challenge growth processes to biochemically transform solid carbonaceous materials at extreme temperatures, pressures, pH, salt and toxic metal concentrations is also disclosed.

Waste Soybean Oil and Corn Steep Liquor as Economic Substrates for Bioemulsifier and Biodiesel Production by Candida lipolytica UCP 0998

PubMed Central

Souza, Adriana Ferreira; Rodriguez, Dayana M.; Ribeaux, Daylin R.; Luna, Marcos A. C.; Lima e Silva, Thayse A.; Andrade, Rosileide F. Silva; Gusmão, Norma B.; Campos-Takaki, Galba M.

2016-01-01

Almost all oleaginous microorganisms are available for biodiesel production, and for the mechanism of oil accumulation, which is what makes a microbial approach economically competitive. This study investigated the potential that the yeast Candida lipolytica UCP0988, in an anamorphous state, has to produce simultaneously a bioemulsifier and to accumulate lipids using inexpensive and alternative substrates. Cultivation was carried out using waste soybean oil and corn steep liquor in accordance with 22 experimental designs with 1% inoculums (107 cells/mL). The bioemulsifier was produced in the cell-free metabolic liquid in the late exponential phase (96 h), at Assay 4 (corn steep liquor 5% and waste soybean oil 8%), with 6.704 UEA, IE24 of 96.66%, and showed an anionic profile. The emulsion formed consisted of compact small and stable droplets (size 0.2–5 µm), stable at all temperatures, at pH 2 and 4, and 2% salinity, and showed an ability to remove 93.74% of diesel oil from sand. The displacement oil (ODA) showed 45.34 cm2 of dispersion (central point of the factorial design). The biomass obtained from Assay 4 was able to accumulate lipids of 0.425 g/g biomass (corresponding to 42.5%), which consisted of Palmitic acid (28.4%), Stearic acid (7.7%), Oleic acid (42.8%), Linoleic acid (19.0%), and γ-Linolenic acid (2.1%). The results showed the ability of C. lipopytica to produce both bioemulsifier and biodiesel using the metabolic conversion of waste soybean oil and corn steep liquor, which are economic renewable sources. PMID:27669227

Simulation studies in biochemical signaling and enzyme reactions

NASA Astrophysics Data System (ADS)

Nelatury, Sudarshan R.; Vagula, Mary C.

2014-06-01

Biochemical pathways characterize various biochemical reaction schemes that involve a set of species and the manner in which they are connected. Determination of schematics that represent these pathways is an important task in understanding metabolism and signal transduction. Examples of these Pathways are: DNA and protein synthesis, and production of several macro-molecules essential for cell survival. A sustained feedback mechanism arises in gene expression and production of mRNA that lead to protein synthesis if the protein so synthesized serves as a transcription factor and becomes a repressor of the gene expression. The cellular regulations are carried out through biochemical networks consisting of reactions and regulatory proteins. Systems biology is a relatively new area that attempts to describe the biochemical pathways analytically and develop reliable mathematical models for the pathways. A complete understanding of chemical reaction kinetics is prohibitively hard thanks to the nonlinear and highly complex mechanisms that regulate protein formation, but attempting to numerically solve some of the governing differential equations seems to offer significant insight about their biochemical picture. To validate these models, one can perform simple experiments in the lab. This paper introduces fundamental ideas in biochemical signaling and attempts to take first steps into the understanding of biochemical oscillations. Initially, the two-pool model of calcium is used to describe the dynamics behind the oscillations. Later we present some elementary results showing biochemical oscillations arising from solving differential equations of Elowitz and Leibler using MATLAB software.

Xeroderma pigmentosum: biochemical and genetic characteristics

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cleaver, J.E.; Bootsma, D.

1975-01-01

Biochemical and genetic studies on xeroderma pigmentosum are reviewed under the following headings: clinical features of xeroderma pigmentosum; karyotype; cell killing and host cell reactivation after irradiation or exposure to chemical carcinogens; SV40 transformation of xeroderma pigmentosum cells; biochemical defects in the common and de Sanctis-Cacchione forms of xeroderma pigmentosum; cell hybridization and complementation groups; biochemical defects in the xeroderma pigmentosum variant and the role of caffeine in DNA repair; DNA repair in xeroderma pigmentosum heterozygotes; response of xeroderma pigmentosum cells to various mutagens and chemical carcinogens; other high and low repair diseases; and possible significance of DNA repair inmore » theories of aging and carcinogenesis. (HLW)« less

Biochemical mechanisms of cisplatin cytotoxicity.

PubMed

Cepeda, Victoria; Fuertes, Miguel A; Castilla, Josefina; Alonso, Carlos; Quevedo, Celia; Pérez, Jose M

2007-01-01

Since the discovery by Rosenberg and collaborators of the antitumor activity of cisplatin 35 years ago, three platinum antitumor drugs (cisplatin, carboplatin and oxaliplatin) have enjoyed a huge clinical and commercial hit. Ever since the initial discovery of the anticancer activity of cisplatin, major efforts have been devoted to elucidate the biochemical mechanisms of antitumor activity of cisplatin in order to be able to rationally design novel platinum based drugs with superior pharmacological profiles. In this report we attempt to provide a current picture of the known facts pertaining to the mechanism of action of the drug, including those involved in drug uptake, DNA damage signals transduction, and cell death through apoptosis or necrosis. A deep knowledge of the biochemical mechanisms, which are triggered in the tumor cell in response to cisplatin injury not only may lead to the design of more efficient platinum antitumor drugs but also may provide new therapeutic strategies based on the biochemical modulation of cisplatin activity.

How to investigate neuro-biochemical relationships on a regional level in humans? Methodological considerations for combining functional with biochemical imaging.

PubMed

Duncan, Niall W; Wiebking, Christine; Muñoz-Torres, Zeidy; Northoff, Georg

2014-01-15

There is an increasing interest in combining different imaging modalities to investigate the relationship between neural and biochemical activity. More specifically, imaging techniques like MRS and PET that allow for biochemical measurement are combined with techniques like fMRI and EEG that measure neural activity in different states. Such combination of neural and biochemical measures raises not only technical issues, such as merging the different data sets, but also several methodological issues. These methodological issues – ranging from hypothesis generation and hypothesis-guided use of technical facilities to target measures and experimental measures – are the focus of this paper. We discuss the various methodological problems and issues raised by the combination of different imaging methodologies in order to investigate neuro-biochemical relationships on a regional level in humans. For example, the choice of transmitter and scan type is discussed, along with approaches to allow the establishment of particular specificities (such as regional or biochemical) to in turn make results fully interpretable. An algorithm that can be used as a form of checklist for designing such multimodal studies is presented. The paper concludes that while several methodological and technical caveats needs to be overcome and addressed, multimodal imaging of the neuro-biochemical relationship provides an important tool to better understand the physiological mechanisms of the human brain.

How to investigate neuro-biochemical relationships on a regional level in humans? Methodological considerations for combining functional with biochemical imaging.

PubMed

Duncan, Niall W; Wiebking, Christine; Munoz-Torres, Zeidy; Northoff, Georg

2013-10-25

There is an increasing interest in combining different imaging modalities to investigate the relationship between neural and biochemical activity. More specifically, imaging techniques like MRS and PET that allow for biochemical measurement are combined with techniques like fMRI and EEG that measure neural activity in different states. Such combination of neural and biochemical measures raises not only technical issues, such as merging the different data sets, but also several methodological issues. These methodological issues - ranging from hypothesis generation and hypothesis-guided use of technical facilities to target measures and experimental measures - are the focus of this paper. We discuss the various methodological problems and issues raised by the combination of different imaging methodologies in order to investigate neuro-biochemical relationships on a regional level in humans. For example, the choice of transmitter and scan type is discussed, along with approaches to allow the establishment of particular specificities (such as regional or biochemical) to in turn make results fully interpretable. An algorithm that can be used as a form of checklist for designing such multimodal studies is presented. The paper concludes that while several methodological and technical caveats needs to be overcome and addressed, multimodal imaging of the neuro-biochemical relationship provides an important tool to better understand the physiological mechanisms of the human brain. Copyright © 2013. Published by Elsevier B.V.

biochem4j: Integrated and extensible biochemical knowledge through graph databases.

PubMed

Swainston, Neil; Batista-Navarro, Riza; Carbonell, Pablo; Dobson, Paul D; Dunstan, Mark; Jervis, Adrian J; Vinaixa, Maria; Williams, Alan R; Ananiadou, Sophia; Faulon, Jean-Loup; Mendes, Pedro; Kell, Douglas B; Scrutton, Nigel S; Breitling, Rainer

2017-01-01

Biologists and biochemists have at their disposal a number of excellent, publicly available data resources such as UniProt, KEGG, and NCBI Taxonomy, which catalogue biological entities. Despite the usefulness of these resources, they remain fundamentally unconnected. While links may appear between entries across these databases, users are typically only able to follow such links by manual browsing or through specialised workflows. Although many of the resources provide web-service interfaces for computational access, performing federated queries across databases remains a non-trivial but essential activity in interdisciplinary systems and synthetic biology programmes. What is needed are integrated repositories to catalogue both biological entities and-crucially-the relationships between them. Such a resource should be extensible, such that newly discovered relationships-for example, those between novel, synthetic enzymes and non-natural products-can be added over time. With the introduction of graph databases, the barrier to the rapid generation, extension and querying of such a resource has been lowered considerably. With a particular focus on metabolic engineering as an illustrative application domain, biochem4j, freely available at http://biochem4j.org, is introduced to provide an integrated, queryable database that warehouses chemical, reaction, enzyme and taxonomic data from a range of reliable resources. The biochem4j framework establishes a starting point for the flexible integration and exploitation of an ever-wider range of biological data sources, from public databases to laboratory-specific experimental datasets, for the benefit of systems biologists, biosystems engineers and the wider community of molecular biologists and biological chemists.

biochem4j: Integrated and extensible biochemical knowledge through graph databases

PubMed Central

Batista-Navarro, Riza; Dunstan, Mark; Jervis, Adrian J.; Vinaixa, Maria; Ananiadou, Sophia; Faulon, Jean-Loup; Kell, Douglas B.

2017-01-01

Biologists and biochemists have at their disposal a number of excellent, publicly available data resources such as UniProt, KEGG, and NCBI Taxonomy, which catalogue biological entities. Despite the usefulness of these resources, they remain fundamentally unconnected. While links may appear between entries across these databases, users are typically only able to follow such links by manual browsing or through specialised workflows. Although many of the resources provide web-service interfaces for computational access, performing federated queries across databases remains a non-trivial but essential activity in interdisciplinary systems and synthetic biology programmes. What is needed are integrated repositories to catalogue both biological entities and–crucially–the relationships between them. Such a resource should be extensible, such that newly discovered relationships–for example, those between novel, synthetic enzymes and non-natural products–can be added over time. With the introduction of graph databases, the barrier to the rapid generation, extension and querying of such a resource has been lowered considerably. With a particular focus on metabolic engineering as an illustrative application domain, biochem4j, freely available at http://biochem4j.org, is introduced to provide an integrated, queryable database that warehouses chemical, reaction, enzyme and taxonomic data from a range of reliable resources. The biochem4j framework establishes a starting point for the flexible integration and exploitation of an ever-wider range of biological data sources, from public databases to laboratory-specific experimental datasets, for the benefit of systems biologists, biosystems engineers and the wider community of molecular biologists and biological chemists. PMID:28708831

An ultraviolet spectrophotometric assay for the screening of sn-2-specific lipases using 1,3-O-dioleoyl-2-O-α-eleostearoyl-sn-glycerol as substrate

PubMed Central

Mendoza, Lilia D.; Rodriguez, Jorge A.; Leclaire, Julien; Buono, Gerard; Fotiadu, Frédéric; Carrière, Frédéric; Abousalham, Abdelkarim

2012-01-01

In the present study, we propose a continuous assay for the screening of sn-2 lipases by using triacylglycerols (TAGs) from Aleurites fordii seed (tung oil) and a synthetic TAG containing the α-eleostearic acid at the sn-2 position and the oleic acid (OA) at the sn-1 and sn-3 positions [1,3-O-dioleoyl-2-O-α-eleostearoyl-sn-glycerol (sn-OEO)]. Each TAG was coated into a microplate well, and the lipase activity was measured by optical density increase at 272 nm due to transition of α-eleostearic acid from the adsorbed to the soluble state. The sn-1,3-regioselective lipases human pancreatic lipase (HPL), LIP2 lipase from Yarrowia lipolytica (YLLIP2), and a known sn-2 lipase, Candida antarctica lipase A (CALA) were used to validate this method. TLC analysis of lipolysis products showed that the lipases tested were able to hydrolyze the sn-OEO and the tung oil TAGs, but only CALA hydrolyzed the sn-2 position. The ratio of initial velocities on sn-OEO and tung oil TAGs was used to estimate the sn-2 preference of lipases. CALA was the enzyme with the highest ratio (0.22 ± 0.015), whereas HPL and YLLIP2 showed much lower ratios (0.072 ± 0.026 and 0.038 ± 0.016, respectively). This continuous sn-2 lipase assay is compatible with a high sample throughput and thus can be applied to the screening of sn-2 lipases. PMID:22114038

Complexity and performance of on-chip biochemical assays

NASA Astrophysics Data System (ADS)

Kopf-Sill, Anne R.; Nikiforov, Theo; Bousse, Luc J.; Nagle, Rob; Parce, J. W.

1997-03-01

The use of microchips for performing biochemical processes has the potential to reduce reagent use and thus assay costs, increase throughput, and automate complex processes. We are building a multifunctional platform that provides sensing and actuation functions for a variety of microchip- based biochemical and analytical processes. Here we describe recent experiments that include on-chip dilution, reagent mixing, reaction, separation, and detection for important classes of biochemical assays. Issues in chip design and control are discussed.

Biochemical Characterization of Prions.

PubMed

Fiorini, Michele; Bongianni, Matilde; Monaco, Salvatore; Zanusso, Gianluigi

2017-01-01

Prion disease or transmissible spongiform encephalopathies are characterized by the presence of the abnormal form of the prion protein (PrP Sc ). The pathological and transmissible properties of PrP Sc are enciphered in its secondary and tertiary structures. Since it's well established that different strains of prions are linked to different conformations of PrP Sc , biochemical characterization of prions seems a preliminary but reliable approach to detect, analyze, and compare prion strains. Experimental biochemical procedures might be helpful in distinguishing PrP Sc physicochemical properties and include resistance to proteinase K (PK) digestion, insolubility in nonionic detergents, PK-resistance under denaturing conditions and sedimentation properties in sucrose gradients. This biochemical approach has been extensively applied in human prion disorders and subsequently expanded for PrP Sc characterization in animals. In particular, in sporadic Creutzfedlt-Jakob disease (sCJD) PrP Sc is characterized by two main glycotypes conventionally named Type 1 and Type 2, based on the apparent gel migration at 21 and 19kDa of the PrP Sc PK-resistant fragment. An additional PrP Sc type was identified in sCJD characterized by an unglycosylated dominant glycoform pattern and in 2010 a variably protease-sensitive prionopathy (VPSPr) was reported showing a PrP Sc with an electrophoretic ladder like pattern. Additionally, the presence of PrP Sc truncated fragments completes the electrophoretic characterization of different prion strains. By two-dimensional (2D) electrophoretic analysis additional PrP Sc pattern was identified, since this procedure provides information about the isoelectric point and the different peptides length related to PK cleavage, as well as to glycosylation extent or GPI anchor presence. We here provide and extensive review on PrP Sc biochemical analysis in human and animal prion disorders. Further, we show that PrP Sc glycotypes observed in CJD share

Hypomagnesemia predicts postoperative biochemical hypocalcemia after thyroidectomy.

PubMed

Luo, Han; Yang, Hongliu; Zhao, Wanjun; Wei, Tao; Su, Anping; Wang, Bin; Zhu, Jingqiang

2017-05-25

To investigate the role of magnesium in biochemical and symptomatic hypocalcemia, a retrospective study was conducted. Less-than-total thyroidectomy patients were excluded from the final analysis. Identified the risk factors of biochemical and symptomatic hypocalcemia, and investigated the correlation by logistic regression and correlation test respectively. A total of 304 patients were included in the final analysis. General incidence of hypomagnesemia was 23.36%. Logistic regression showed that gender (female) (OR = 2.238, p = 0.015) and postoperative hypomagnesemia (OR = 2.010, p = 0.017) were independent risk factors for biochemical hypocalcemia. Both Pearson and partial correlation tests indicated there was indeed significant relation between calcium and magnesium. However, relative decreasing of iPTH (>70%) (6.691, p < 0.001) and hypocalcemia (2.222, p = 0.046) were identified as risk factors of symptomatic hypocalcemia. The difference remained significant even in normoparathyroidism patients. Postoperative hypomagnesemia was independent risk factor of biochemical hypocalcemia. Relative decline of iPTH was predominating in predicting symptomatic hypocalcemia.

Phaeochromocytoma: diagnostic challenges for biochemical screening and diagnosis.

PubMed

Barron, Jeffrey

2010-08-01

The aim of this article is to provide knowledge of the origin of catecholamines and metabolites so that there can be an informed approach to the methods for biochemical screening for a possible phaeochromocytoma; The article includes a review of catecholamine and metadrenaline metabolism, with methods used in biochemical screening. In the adrenal medulla and a phaeochromocytoma, catecholamines continuously leak from chromaffin granules into the cytoplasm and are converted to metadrenalines. For a phaeochromocytoma to become biochemically detectable, metnoradrenaline secretion needs to rise fourfold, whereas noradrenaline secretion needs to rise 15-fold. The prevalence of a sporadic phaeochromocytoma is low; therefore false-positive results exceed true-positive results. Assay sensitivity is high because it is important not to miss a possible phaeochromocytoma. The use of urine or plasma fractionated metadrenalines as the first-line test has been recommended due to improved sensitivity. A negative result excludes a phaeochromocytoma. Only after a sporadic phaeochromocytoma has been diagnosed biochemically is it cost effective to request imaging. Sensitivities and specificities of the assays differ according to pre-test probabilities of the presence of a phaeochromocytoma, with hereditary and incidentalomas having a higher pre-test probability than sporadic phaeochromocytoma. In conclusion, in screening for a possible phaeochromocytoma, biochemical investigations should be completed first to exclude or establish the diagnosis. The preferred biochemical screening test is fractionated metadrenalines, including methoxytyramine so as not to miss dopamine-secreting tumours.

eQuilibrator--the biochemical thermodynamics calculator.

PubMed

Flamholz, Avi; Noor, Elad; Bar-Even, Arren; Milo, Ron

2012-01-01

The laws of thermodynamics constrain the action of biochemical systems. However, thermodynamic data on biochemical compounds can be difficult to find and is cumbersome to perform calculations with manually. Even simple thermodynamic questions like 'how much Gibbs energy is released by ATP hydrolysis at pH 5?' are complicated excessively by the search for accurate data. To address this problem, eQuilibrator couples a comprehensive and accurate database of thermodynamic properties of biochemical compounds and reactions with a simple and powerful online search and calculation interface. The web interface to eQuilibrator (http://equilibrator.weizmann.ac.il) enables easy calculation of Gibbs energies of compounds and reactions given arbitrary pH, ionic strength and metabolite concentrations. The eQuilibrator code is open-source and all thermodynamic source data are freely downloadable in standard formats. Here we describe the database characteristics and implementation and demonstrate its use.

eQuilibrator—the biochemical thermodynamics calculator

PubMed Central

Flamholz, Avi; Noor, Elad; Bar-Even, Arren; Milo, Ron

2012-01-01

The laws of thermodynamics constrain the action of biochemical systems. However, thermodynamic data on biochemical compounds can be difficult to find and is cumbersome to perform calculations with manually. Even simple thermodynamic questions like ‘how much Gibbs energy is released by ATP hydrolysis at pH 5?’ are complicated excessively by the search for accurate data. To address this problem, eQuilibrator couples a comprehensive and accurate database of thermodynamic properties of biochemical compounds and reactions with a simple and powerful online search and calculation interface. The web interface to eQuilibrator (http://equilibrator.weizmann.ac.il) enables easy calculation of Gibbs energies of compounds and reactions given arbitrary pH, ionic strength and metabolite concentrations. The eQuilibrator code is open-source and all thermodynamic source data are freely downloadable in standard formats. Here we describe the database characteristics and implementation and demonstrate its use. PMID:22064852

Biochemical adaptation to ocean acidification.

PubMed

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

Biochemical Testing in Thyroid Disorders.

PubMed

Esfandiari, Nazanene H; Papaleontiou, Maria

2017-09-01

This article summarizes the main principles for the appropriate use of laboratory testing in the diagnosis and management of thyroid disorders, as well as controversies that have arisen in association with some of these biochemical tests. To place a test in perspective, its sensitivity and accuracy should be taken into account. Ordering the correct laboratory tests facilitates the early diagnosis of a thyroid disorder and allows for timely and appropriate treatment. This article focuses on a comprehensive update regarding thyroid-stimulating hormone, thyroxine/triiodothyronine, thyroid autoantibodies, thyroglobulin, and calcitonin. Clinical uses of these biochemical tests are outlined. Copyright © 2017 Elsevier Inc. All rights reserved.

Biochemical phenotypes to discriminate microbial subpopulations and improve outbreak detection.

PubMed

Galar, Alicia; Kulldorff, Martin; Rudnick, Wallis; O'Brien, Thomas F; Stelling, John

2013-01-01

Clinical microbiology laboratories worldwide constitute an invaluable resource for monitoring emerging threats and the spread of antimicrobial resistance. We studied the growing number of biochemical tests routinely performed on clinical isolates to explore their value as epidemiological markers. Microbiology laboratory results from January 2009 through December 2011 from a 793-bed hospital stored in WHONET were examined. Variables included patient location, collection date, organism, and 47 biochemical and 17 antimicrobial susceptibility test results reported by Vitek 2. To identify biochemical tests that were particularly valuable (stable with repeat testing, but good variability across the species) or problematic (inconsistent results with repeat testing), three types of variance analyses were performed on isolates of K. pneumonia: descriptive analysis of discordant biochemical results in same-day isolates, an average within-patient variance index, and generalized linear mixed model variance component analysis. 4,200 isolates of K. pneumoniae were identified from 2,485 patients, 32% of whom had multiple isolates. The first two variance analyses highlighted SUCT, TyrA, GlyA, and GGT as "nuisance" biochemicals for which discordant within-patient test results impacted a high proportion of patient results, while dTAG had relatively good within-patient stability with good heterogeneity across the species. Variance component analyses confirmed the relative stability of dTAG, and identified additional biochemicals such as PHOS with a large between patient to within patient variance ratio. A reduced subset of biochemicals improved the robustness of strain definition for carbapenem-resistant K. pneumoniae. Surveillance analyses suggest that the reduced biochemical profile could improve the timeliness and specificity of outbreak detection algorithms. The statistical approaches explored can improve the robust recognition of microbial subpopulations with routinely available

Biochemical Phenotypes to Discriminate Microbial Subpopulations and Improve Outbreak Detection

PubMed Central

Galar, Alicia; Kulldorff, Martin; Rudnick, Wallis; O'Brien, Thomas F.; Stelling, John

2013-01-01

Background Clinical microbiology laboratories worldwide constitute an invaluable resource for monitoring emerging threats and the spread of antimicrobial resistance. We studied the growing number of biochemical tests routinely performed on clinical isolates to explore their value as epidemiological markers. Methodology/Principal Findings Microbiology laboratory results from January 2009 through December 2011 from a 793-bed hospital stored in WHONET were examined. Variables included patient location, collection date, organism, and 47 biochemical and 17 antimicrobial susceptibility test results reported by Vitek 2. To identify biochemical tests that were particularly valuable (stable with repeat testing, but good variability across the species) or problematic (inconsistent results with repeat testing), three types of variance analyses were performed on isolates of K. pneumonia: descriptive analysis of discordant biochemical results in same-day isolates, an average within-patient variance index, and generalized linear mixed model variance component analysis. Results: 4,200 isolates of K. pneumoniae were identified from 2,485 patients, 32% of whom had multiple isolates. The first two variance analyses highlighted SUCT, TyrA, GlyA, and GGT as “nuisance” biochemicals for which discordant within-patient test results impacted a high proportion of patient results, while dTAG had relatively good within-patient stability with good heterogeneity across the species. Variance component analyses confirmed the relative stability of dTAG, and identified additional biochemicals such as PHOS with a large between patient to within patient variance ratio. A reduced subset of biochemicals improved the robustness of strain definition for carbapenem-resistant K. pneumoniae. Surveillance analyses suggest that the reduced biochemical profile could improve the timeliness and specificity of outbreak detection algorithms. Conclusions The statistical approaches explored can improve the

Correlations between female breast density and biochemical markers.

PubMed

Kim, Ji-Hye; Lee, Hae-Kag; Cho, Jae-Hwan; Park, Hyong-Keun; Yang, Han-Jun

2015-07-01

[Purpose] The aim of this study was to identify biochemical markers related to breast density. The study was performed with 200 patients who received mammography and biochemical marker testing between March 1, 2014 to October 1, 2014. [Subjects and Methods] Following the American College of Radiology, Breast Imaging Reporting and Data System (ACR BI-RADS), breast parenchymal pattern density from mammography was categorized into four grades: grade 1, almost entirely fat; grade 2, fibroglandular densities; grade 3, heterogeneously dense; and grade 4, extremely dense. Regarding biochemical markers, subjects underwent blood and urine tests after a 12-h fast. We analyzed correlations among breast density, general characteristics, and biochemical markers. [Results] Breast density-related factors were age, height, weight, body mass index (BMI), hematocrit, MCH, RDW, AST, ALT, ALP, uric acid, γGT, triglycerides, total cholesterol, HDL-cholesterol, and LDL-cholesterol. [Conclusion] The results can be used as basic and comparative data for the prevention and early control of breast cancer.

Stochastic hybrid systems for studying biochemical processes.

PubMed

Singh, Abhyudai; Hespanha, João P

2010-11-13

Many protein and mRNA species occur at low molecular counts within cells, and hence are subject to large stochastic fluctuations in copy numbers over time. Development of computationally tractable frameworks for modelling stochastic fluctuations in population counts is essential to understand how noise at the cellular level affects biological function and phenotype. We show that stochastic hybrid systems (SHSs) provide a convenient framework for modelling the time evolution of population counts of different chemical species involved in a set of biochemical reactions. We illustrate recently developed techniques that allow fast computations of the statistical moments of the population count, without having to run computationally expensive Monte Carlo simulations of the biochemical reactions. Finally, we review different examples from the literature that illustrate the benefits of using SHSs for modelling biochemical processes.

40 CFR 158.2080 - Experimental use permit data requirements-biochemical pesticides.

Code of Federal Regulations, 2011 CFR

2011-07-01

... requirements-biochemical pesticides. 158.2080 Section 158.2080 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) PESTICIDE PROGRAMS DATA REQUIREMENTS FOR PESTICIDES Biochemical Pesticides § 158.2080 Experimental use permit data requirements—biochemical pesticides. (a) Sections 158.2081...

40 CFR 158.2080 - Experimental use permit data requirements-biochemical pesticides.

Code of Federal Regulations, 2012 CFR

2012-07-01

... requirements-biochemical pesticides. 158.2080 Section 158.2080 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) PESTICIDE PROGRAMS DATA REQUIREMENTS FOR PESTICIDES Biochemical Pesticides § 158.2080 Experimental use permit data requirements—biochemical pesticides. (a) Sections 158.2081...

40 CFR 158.2080 - Experimental use permit data requirements-biochemical pesticides.

Code of Federal Regulations, 2014 CFR

2014-07-01

... requirements-biochemical pesticides. 158.2080 Section 158.2080 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) PESTICIDE PROGRAMS DATA REQUIREMENTS FOR PESTICIDES Biochemical Pesticides § 158.2080 Experimental use permit data requirements—biochemical pesticides. (a) Sections 158.2081...

40 CFR 158.2080 - Experimental use permit data requirements-biochemical pesticides.

Code of Federal Regulations, 2010 CFR

2010-07-01

... requirements-biochemical pesticides. 158.2080 Section 158.2080 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) PESTICIDE PROGRAMS DATA REQUIREMENTS FOR PESTICIDES Biochemical Pesticides § 158.2080 Experimental use permit data requirements—biochemical pesticides. (a) Sections 158.2081...

40 CFR 158.2080 - Experimental use permit data requirements-biochemical pesticides.

Code of Federal Regulations, 2013 CFR

2013-07-01

... requirements-biochemical pesticides. 158.2080 Section 158.2080 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) PESTICIDE PROGRAMS DATA REQUIREMENTS FOR PESTICIDES Biochemical Pesticides § 158.2080 Experimental use permit data requirements—biochemical pesticides. (a) Sections 158.2081...

Incidence of Abnormal Liver Biochemical Tests in Hyperthyroidism

PubMed Central

Lin, Tiffany Y.; Shekar, Anshula O.; Li, Ning; Yeh, Michael W.; Saab, Sammy; Wilson, Mark; Leung, Angela M.

2017-01-01

Objective Abnormal serum liver function tests are common in patients with untreated thyrotoxicosis, even prior to the initiation of antithyroidal medications that may worsen their severity. There is a wide range of the incidence of these abnormalities in the published literature. The aim of this study was to assess the risks factors and threshold of thyrotoxicosis severity for developing an abnormal liver biochemical test upon the diagnosis of new thyrotoxicosis. Design Single-institution retrospective cohort study. Patients Patients ≥18 years old receiving medical care at a large, academic, urban U.S. medical center between 2002–2016. Measurements Inclusion criteria were a serum thyroid stimulating hormone [TSH] concentration < 0.3 mIU/L or ICD-9 code for thyrotoxicosis, with thyrotoxicosis confirmed by either a concurrent elevated serum triiodothyronine (T3) and/or thyroxine (T4) concentration [total or free] within 3 months), and an available liver biochemical test(s) within 6 months of thyrotoxicosis. The biochemical liver tests assessed were serum aspartate transaminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (AP), gamma-glutamyltransferase (GGT), total bilirubin, and conjugated bilirubin concentrations. Results In this cohort of 1,514 subjects, the overall incidence of any biochemical liver test abnormality within 6 months of thyrotoxicosis was 39%. An initial serum TSH concentration <0.02 mIU/L, male gender, and African-American race were significant predictors of an abnormal serum liver biochemical test within 6 months of the diagnosis of new-onset untreated thyrotoxicosis. Conclusions This study identifies risk factors for patients who develop an abnormal serum liver biochemical test result within 6 months of a diagnosis of untreated thyrotoxicosis. PMID:28199740

Incidence of abnormal liver biochemical tests in hyperthyroidism.

PubMed

Lin, Tiffany Y; Shekar, Anshula O; Li, Ning; Yeh, Michael W; Saab, Sammy; Wilson, Mark; Leung, Angela M

2017-05-01

Abnormal serum liver function tests are common in patients with untreated thyrotoxicosis, even prior to the initiation of antithyroidal medications that may worsen the severity of the abnormal serum liver biochemistries. There is a wide range of the incidence of these abnormalities in the published literature. The aim of this study was to assess the risks factors and threshold of thyrotoxicosis severity for developing an abnormal liver biochemical test upon the diagnosis of new thyrotoxicosis. Single-institution retrospective cohort study. Patients of ≥18 years old receiving medical care at a large, academic, urban US medical centre between 2002-2016. Inclusion criteria were a serum thyroid stimulating hormone (TSH) concentration of <0·3 mIU/l or ICD-9 code for thyrotoxicosis, with thyrotoxicosis confirmed by either a concurrent elevated serum triiodothyronine (T3) or thyroxine (T4) concentration ([total or free] within 3 months), and an available liver biochemical test(s) within 6 months of thyrotoxicosis. The biochemical liver tests assessed were serum aspartate transaminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (AP), gamma-glutamyltransferase (GGT), total bilirubin, and conjugated bilirubin concentrations. In this cohort of 1514 subjects, the overall incidence of any biochemical liver test abnormality within 6 months of thyrotoxicosis was 39%. An initial serum TSH concentration <0·02 mIU/l, male gender, and African-American race were significant predictors of an abnormal serum liver biochemical test within 6 months of the diagnosis of new-onset untreated thyrotoxicosis. This study identifies risk factors for patients who develop an abnormal serum liver biochemical test result within 6 months of a diagnosis of untreated thyrotoxicosis. © 2017 John Wiley & Sons Ltd.

Biotinidase deficiency: Genotype-biochemical phenotype association in Brazilian patients

PubMed Central

Borsatto, Taciane; Sperb-Ludwig, Fernanda; Lima, Samyra E.; S. Carvalho, Maria R.; S. Fonseca, Pablo A.; S. Camelo, José; M. Ribeiro, Erlane; F. V. de Medeiros, Paula; M. Lourenço, Charles; F. M. de Souza, Carolina; Boy, Raquel; Félix, Têmis M.; M. Bittar, Camila; L. C. Pinto, Louise; C. Neto, Eurico; J. Blom, Henk; D. Schwartz, Ida V.

2017-01-01

Introduction The association between the BTD genotype and biochemical phenotype [profound biotinidase deficiency (BD), partial BD or heterozygous activity] is not always consistent. This study aimed to investigate the genotype-biochemical phenotype association in patients with low biotinidase activity. Methods All exons, the 5'UTR and the promoter of the BTD gene were sequenced in 72 Brazilian individuals who exhibited low biotinidase activity. For each patient, the expected biochemical phenotype based on the known genotype was compared with the observed biochemical phenotype. Additional non-genetic factors that could affect the biotinidase activity were also analysed. Results Most individuals were identified by neonatal screening (n = 66/72). When consecutive results for the same patient were compared, age, prematurity and neonatal jaundice appeared to affect the level of biotinidase activity. The biochemical phenotype at the time of the second blood collection changed in 11/22 patients compared to results from the first sample. Three novel variants were found: c.1337T>C (p.L446P), c.1466A>G (p.N489S) and c.962G>A (p.W321*). Some patients with the same genotype presented different biochemical phenotypes. The expected and observed biochemical phenotypes agreed in 68.5% of cases (concordant patients). The non-coding variants c.-183G>A, c.-315A>G and c.-514C>T were present in heterozygosis in 5/17 discordant patients. In addition, c.-183G>A and c.-514C>T were also present in 10/37 concordant patients. Conclusions The variants found in the promoter region do not appear to have a strong impact on biotinidase activity. Since there is a disparity between the BTD genotype and biochemical phenotype, and biotinidase activity may be affected by both genetic and non-genetic factors, we suggest that the diagnosis of BD should be based on more than one measurement of plasma biotinidase activity. DNA analysis can be of additional relevance to differentiate between partial BD and

Accurate atom-mapping computation for biochemical reactions.

PubMed

Latendresse, Mario; Malerich, Jeremiah P; Travers, Mike; Karp, Peter D

2012-11-26

The complete atom mapping of a chemical reaction is a bijection of the reactant atoms to the product atoms that specifies the terminus of each reactant atom. Atom mapping of biochemical reactions is useful for many applications of systems biology, in particular for metabolic engineering where synthesizing new biochemical pathways has to take into account for the number of carbon atoms from a source compound that are conserved in the synthesis of a target compound. Rapid, accurate computation of the atom mapping(s) of a biochemical reaction remains elusive despite significant work on this topic. In particular, past researchers did not validate the accuracy of mapping algorithms. We introduce a new method for computing atom mappings called the minimum weighted edit-distance (MWED) metric. The metric is based on bond propensity to react and computes biochemically valid atom mappings for a large percentage of biochemical reactions. MWED models can be formulated efficiently as Mixed-Integer Linear Programs (MILPs). We have demonstrated this approach on 7501 reactions of the MetaCyc database for which 87% of the models could be solved in less than 10 s. For 2.1% of the reactions, we found multiple optimal atom mappings. We show that the error rate is 0.9% (22 reactions) by comparing these atom mappings to 2446 atom mappings of the manually curated Kyoto Encyclopedia of Genes and Genomes (KEGG) RPAIR database. To our knowledge, our computational atom-mapping approach is the most accurate and among the fastest published to date. The atom-mapping data will be available in the MetaCyc database later in 2012; the atom-mapping software will be available within the Pathway Tools software later in 2012.

Biochemical studies in patients with hyperinsulinaemic hypoglycaemia.

PubMed

Al-Otaibi, Hessah; Senniappan, Senthil; Alam, Syeda; Hussain, Khalid

2013-11-01

Hyperinsulinaemic hypoglycaemia (HH) is characterised by the dysregulated secretion of insulin from the pancreatic β-cell. It is a major cause of severe and persistent hypoglycaemia in the newborn period. There have been no previous studies assessing the various biochemical alterations at the time of hypoglycaemia in relation to the severity of the hypoglycaemia. Biochemical and clinical data were collected on 90 neonates (gestational age range, 32-42 weeks) with a diagnosis of HH [(based on glucose requirement  > 8 mg/kg/min) and the biochemical profile of insulin action (low beta-hydroxybutyrate and fatty acid concentrations)] who had undergone fasting studies. The results showed that (a) the serum insulin level measured at the time of hypoglycaemia had no correlation with the severity of hypoglycaemia, (b) the serum insulin level was undetectable despite severe hypoglycaemia in a significant proportion of patients, (c) there was no correlation between the birth weight and the insulin level at the time of hypoglycaemia, (d) the suppression of ketogenesis was more marked than that of the non-esterified fatty acids. This study suggests that the diagnosis of HH should not rely solely on a raised serum insulin level at the time of hypoglycaemia but on the constellation of clinical and biochemical findings.

Biochemical characteristics among Mycobacterium bovis BCG substrains.

PubMed

Hayashi, Daisuke; Takii, Takemasa; Mukai, Tetsu; Makino, Masahiko; Yasuda, Emi; Horita, Yasuhiro; Yamamoto, Ryuji; Fujiwara, Akiko; Kanai, Keita; Kondo, Maki; Kawarazaki, Aya; Yano, Ikuya; Yamamoto, Saburo; Onozaki, Kikuo

2010-05-01

In order to evaluate the biochemical characteristics of 14 substrains of Mycobacterium bovis bacillus Calmette Guérin (BCG) - Russia, Moreau, Japan, Sweden, Birkhaug, Danish, Glaxo, Mexico, Tice, Connaught, Montreal, Phipps, Australia and Pasteur - we performed eight different biochemical tests, including those for nitrate reduction, catalase, niacin accumulation, urease, Tween 80 hydrolysis, pyrazinamidase, p-amino salicylate degradation and resistance to thiophene 2-carboxylic acid hydrazide. Catalase activities of the substrains were all low. Data for nitrate reduction, niacin accumulation, Tween 80 hydrolysis, susceptibility to hydrogen peroxide and nitrate, and optimal pH for growth were all variable among these substrains. These findings suggest that the heterogeneities of biochemical characteristics are relevant to the differences in resistance of BCG substrains to environmental stress. The study also contributes to the re-evaluation of BCG substrains for use as vaccines.

Biochemical Characterization of Prion Strains in Bank Voles

PubMed Central

Pirisinu, Laura; Marcon, Stefano; Di Bari, Michele Angelo; D’Agostino, Claudia; Agrimi, Umberto; Nonno, Romolo

2013-01-01

Prions exist as different strains exhibiting distinct disease phenotypes. Currently, the identification of prion strains is still based on biological strain typing in rodents. However, it has been shown that prion strains may be associated with distinct PrPSc biochemical types. Taking advantage of the availability of several prion strains adapted to a novel rodent model, the bank vole, we investigated if any prion strain was actually associated with distinctive PrPSc biochemical characteristics and if it was possible to univocally identify strains through PrPSc biochemical phenotypes. We selected six different vole-adapted strains (three human-derived and three animal-derived) and analyzed PrPSc from individual voles by epitope mapping of protease resistant core of PrPSc (PrPres) and by conformational stability and solubility assay. Overall, we discriminated five out of six prion strains, while two different scrapie strains showed identical PrPSc types. Our results suggest that the biochemical strain typing approach here proposed was highly discriminative, although by itself it did not allow us to identify all prion strains analyzed. PMID:25437201

Craniometaphyseal dysplasia with obvious biochemical abnormality and rickets-like features.

PubMed

Wu, Bo; Jiang, Yan; Wang, Ou; Li, Mei; Xing, Xiao-Ping; Xia, Wei-Bo

2016-05-01

Craniometaphyseal dysplasia (CMD) is a rare genetic disorder that is characterized by progressive sclerosis of the craniofacial bones and metaphyseal widening of long bones, and biochemical indexes were mostly normal. To further the understanding of the disease from a biochemical perspective, we reported a CMD case with obviously abnormal biochemical indexes. A 1-year-old boy was referred to our clinic. Biochemical test showed obviously increased alkaline phosphatase (ALP) and parathyroid hormone (PTH), mild hypocalcemia and hypophosphatemia. Moreover, significant elevated receptor activator of nuclear factor kappa-B ligand (RANKL) level, but normal β-C-terminal telopeptide of type I collagen (β-CTX) concentration were revealed. He was initially suspected of rickets, because the radiological examination also showed broadened epiphysis in his long bones. Supplementation with calcium and calcitriol alleviated biochemical abnormality. However, the patient gradually developed osteosclerosis which was inconformity with rickets. Considering that he was also presented with facial paralysis and nasal obstruction symptom, the diagnosis of craniometaphyseal dysplasia was suspected, and then was confirmed by the mutation analysis of ANKH of the proband and his family, which showed a de novo heterozygous mutation (C1124-1126delCCT) on exon 9. Our study revealed that obvious biochemical abnormality and rickets-like features might present as uncommon characteristics in CMD patients, and the calcium and calcitriol supplementation could alleviate biochemical abnormalities. Furthermore, although early osteoclast differentiation factor was excited in CMD patient, activity of osteoclast was still inert. Copyright © 2016. Published by Elsevier B.V.

A Biochemical Approach to the Problem of Dyslexia.

ERIC Educational Resources Information Center

Baker, Sidney McDonald

1985-01-01

The paper presents the case of a sixth-grade boy, labeled dyslexic, who responded positively to a biochemical approach. Remedy of iron, zinc, and Vitamin B-6 deficiencies as well as an imbalance of fatty acids resulted in improvements in hair and skin and also in reading. A biochemical approach to behavior problems is proposed. (Author/CL)

Developments in commercially produced microbials at Biochem Products

Treesearch

John Lublinkhof; Douglas H. Ross

1985-01-01

Biochem Products is part of a large industrial and scientific family - the Solvay Group. Solvay, headquartered in Brussels, Belgium is a multinational company with 46,000 employees worldwide. In the U.S., our working partners include a large polymer manufacturer, a peroxygen producer and a leading poultry and animal health products company. Biochem Products is a...

Management Options for Biochemically Recurrent Prostate Cancer.

PubMed

Fakhrejahani, Farhad; Madan, Ravi A; Dahut, William L

2017-05-01

Prostate cancer is the most common solid tumor malignancy in men worldwide. Treatment with surgery and radiation can be curative in organ-confined disease. Unfortunately, about one third of men develop biochemically recurrent disease based only on rising prostate-specific antigen (PSA) in the absence of visible disease on conventional imaging. For these patients with biochemical recurrent prostate cancer, there is no uniform guideline for subsequent management. Based on available data, it seems prudent that biochemical recurrent prostate cancer should initially be evaluated for salvage radiation or prostatectomy, with curative intent. In selected cases, high-intensity focused ultrasound and cryotherapy may be considered in patients that meet very narrow criteria as defined by non-randomized trials. If salvage options are not practical or unsuccessful, androgen deprivation therapy (ADT) is a standard option for disease control. While some patients prefer ADT to manage the disease immediately, others defer treatment because of the associated toxicity. In the absence of definitive randomized data, patients may be followed using PSA doubling time as a trigger to initiate ADT. Based on retrospective data, a PSA doubling time of less than 3-6 months has been associated with near-term development of metastasis and thus could be used signal to initiate ADT. Once treatment is begun, patients and their providers can choose between an intermittent and continuous ADT strategy. The intermittent approach may limit side effects but in patients with metastatic disease studies could not exclude a 20% greater risk of death. In men with biochemical recurrence, large studies have shown that intermittent therapy is non-inferior to continuous therapy, thus making this a reasonable option. Since biochemically recurrent prostate cancer is defined by technological limitations of radiographic detection, as new imaging (i.e., PSMA) strategies are developed, it may alter how the disease is

Kombucha tea fermentation: Microbial and biochemical dynamics.

PubMed

Chakravorty, Somnath; Bhattacharya, Semantee; Chatzinotas, Antonis; Chakraborty, Writachit; Bhattacharya, Debanjana; Gachhui, Ratan

2016-03-02

Kombucha tea, a non-alcoholic beverage, is acquiring significant interest due to its claimed beneficial properties. The microbial community of Kombucha tea consists of bacteria and yeast which thrive in two mutually non-exclusive compartments: the soup or the beverage and the biofilm floating on it. The microbial community and the biochemical properties of the beverage have so far mostly been described in separate studies. This, however, may prevent understanding the causal links between the microbial communities and the beneficial properties of Kombucha tea. Moreover, an extensive study into the microbial and biochemical dynamics has also been missing. In this study, we thus explored the structure and dynamics of the microbial community along with the biochemical properties of Kombucha tea at different time points up to 21 days of fermentation. We hypothesized that several biochemical properties will change during the course of fermentation along with the shifts in the yeast and bacterial communities. The yeast community of the biofilm did not show much variation over time and was dominated by Candida sp. (73.5-83%). The soup however, showed a significant shift in dominance from Candida sp. to Lachancea sp. on the 7th day of fermentation. This is the first report showing Candida as the most dominating yeast genus during Kombucha fermentation. Komagateibacter was identified as the single largest bacterial genus present in both the biofilm and the soup (~50%). The bacterial diversity was higher in the soup than in the biofilm with a peak on the seventh day of fermentation. The biochemical properties changed with the progression of the fermentation, i.e., beneficial properties of the beverage such as the radical scavenging ability increased significantly with a maximum increase at day 7. We further observed a significantly higher D-saccharic acid-1,4-lactone content and caffeine degradation property compared to previously described Kombucha tea fermentations. Our

Maximizing the Biochemical Resolving Power of Fluorescence Microscopy

PubMed Central

Esposito, Alessandro; Popleteeva, Marina; Venkitaraman, Ashok R.

2013-01-01

Most recent advances in fluorescence microscopy have focused on achieving spatial resolutions below the diffraction limit. However, the inherent capability of fluorescence microscopy to non-invasively resolve different biochemical or physical environments in biological samples has not yet been formally described, because an adequate and general theoretical framework is lacking. Here, we develop a mathematical characterization of the biochemical resolution in fluorescence detection with Fisher information analysis. To improve the precision and the resolution of quantitative imaging methods, we demonstrate strategies for the optimization of fluorescence lifetime, fluorescence anisotropy and hyperspectral detection, as well as different multi-dimensional techniques. We describe optimized imaging protocols, provide optimization algorithms and describe precision and resolving power in biochemical imaging thanks to the analysis of the general properties of Fisher information in fluorescence detection. These strategies enable the optimal use of the information content available within the limited photon-budget typically available in fluorescence microscopy. This theoretical foundation leads to a generalized strategy for the optimization of multi-dimensional optical detection, and demonstrates how the parallel detection of all properties of fluorescence can maximize the biochemical resolving power of fluorescence microscopy, an approach we term Hyper Dimensional Imaging Microscopy (HDIM). Our work provides a theoretical framework for the description of the biochemical resolution in fluorescence microscopy, irrespective of spatial resolution, and for the development of a new class of microscopes that exploit multi-parametric detection systems. PMID:24204821

Measures of Biochemical Sociology

ERIC Educational Resources Information Center

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…

Biochemical Education in Brazil.

ERIC Educational Resources Information Center

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)

Weak Perturbations of Biochemical Oscillators

NASA Astrophysics Data System (ADS)

Gailey, Paul

2001-03-01

Biochemical oscillators may play important roles in gene regulation, circadian rhythms, physiological signaling, and sensory processes. These oscillations typically occur inside cells where the small numbers of reacting molecules result in fluctuations in the oscillation period. Some oscillation mechanisms have been reported that resist fluctuations and produce more stable oscillations. In this paper, we consider the use of biochemical oscillators as sensors by comparing inherent fluctuations with the effects of weak perturbations to one of the reactants. Such systems could be used to produce graded responses to weak stimuli. For example, a leading hypothesis to explain geomagnetic navigation in migrating birds and other animals is based on magnetochemical reactions. Because the magnitude of magnetochemical effects is small at geomagnetic field strengths, a sensitive, noise resistant detection scheme would be required.

Discovering Reliable Sources of Biochemical Thermodynamic Data to Aid Students' Understanding

ERIC Educational Resources Information Center

Me´ndez, Eduardo; Cerda´, María F.

2016-01-01

Students of physical chemistry in biochemical disciplines need biochemical examples to capture the need, not always understood, of a difficult area in their studies. The use of thermodynamic data in the chemical reference state may lead to incorrect interpretations in the analysis of biochemical examples when the analysis does not include relevant…

The purification and characterization of ATP synthase complexes from the mitochondria of four fungal species.

PubMed

Liu, Sidong; Charlesworth, Thomas J; Bason, John V; Montgomery, Martin G; Harbour, Michael E; Fearnley, Ian M; Walker, John E

2015-05-15

The ATP synthases have been isolated by affinity chromatography from the mitochondria of the fungal species Yarrowia lipolytica, Pichia pastoris, Pichia angusta and Saccharomyces cerevisiae. The subunit compositions of the purified enzyme complexes depended on the detergent used to solubilize and purify the complex, and the presence or absence of exogenous phospholipids. All four enzymes purified in the presence of n-dodecyl-β-D-maltoside had a complete complement of core subunits involved directly in the synthesis of ATP, but they were deficient to different extents in their supernumerary membrane subunits. In contrast, the enzymes from P. angusta and S. cerevisiae purified in the presence of n-decyl-β-maltose neopentyl glycol and the phospholipids 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine, 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine, cardiolipin (diphosphatidylglycerol) and 1-palmitoyl-2-oleoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] had a complete complement of core subunits and also contained all of the known supernumerary membrane subunits, e, f, g, j, k and ATP8 (or Aap1), plus an additional new membrane component named subunit l, related in sequence to subunit k. The catalytic domain of the enzyme from P. angusta was more resistant to thermal denaturation than the enzyme from S. cerevisiae, but less stable than the catalytic domain of the bovine enzyme, but the stator and the integrity of the transmembrane proton pathway were most stable in the enzyme from P. angusta. The P. angusta enzyme provides a suitable source of enzyme for studying the structure of the membrane domain and properties associated with that sector of the enzyme complex.

The purification and characterization of ATP synthase complexes from the mitochondria of four fungal species

PubMed Central

Liu, Sidong; Charlesworth, Thomas J.; Bason, John V.; Montgomery, Martin G.; Harbour, Michael E.; Fearnley, Ian M.; Walker, John E.

2015-01-01

The ATP synthases have been isolated by affinity chromatography from the mitochondria of the fungal species Yarrowia lipolytica, Pichia pastoris, Pichia angusta and Saccharomyces cerevisiae. The subunit compositions of the purified enzyme complexes depended on the detergent used to solubilize and purify the complex, and the presence or absence of exogenous phospholipids. All four enzymes purified in the presence of n-dodecyl-β-D-maltoside had a complete complement of core subunits involved directly in the synthesis of ATP, but they were deficient to different extents in their supernumerary membrane subunits. In contrast, the enzymes from P. angusta and S. cerevisiae purified in the presence of n-decyl-β-maltose neopentyl glycol and the phospholipids 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine, 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine, cardiolipin (diphosphatidylglycerol) and 1-palmitoyl-2-oleoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] had a complete complement of core subunits and also contained all of the known supernumerary membrane subunits, e, f, g, j, k and ATP8 (or Aap1), plus an additional new membrane component named subunit l, related in sequence to subunit k. The catalytic domain of the enzyme from P. angusta was more resistant to thermal denaturation than the enzyme from S. cerevisiae, but less stable than the catalytic domain of the bovine enzyme, but the stator and the integrity of the transmembrane proton pathway were most stable in the enzyme from P. angusta. The P. angusta enzyme provides a suitable source of enzyme for studying the structure of the membrane domain and properties associated with that sector of the enzyme complex. PMID:25759169

Novel Strategy of Using Methyl Esters as Slow Release Methanol Source during Lipase Expression by mut+ Pichia pastoris X33

PubMed Central

Kumari, Arti; Gupta, Rani

2014-01-01

One of the major issues with heterologous production of proteins in Pichia pastoris X33 under AOX1 promoter is repeated methanol induction. To obviate repeated methanol induction, methyl esters were used as a slow release source of methanol in lipase expressing mut+ recombinant. Experimental design was based on the strategy that in presence of lipase, methyl esters can be hydrolysed to release their products as methanol and fatty acid. Hence, upon break down of methyl esters by lipase, first methanol will be used as a carbon source and inducer. Then P. pastoris can switch over to fatty acid as a carbon source for multiplication and biomass maintenance till further induction by methyl esters. We validated this strategy using recombinant P. pastoris expressing Lip A, Lip C from Trichosporon asahii and Lip11 from Yarrowia lipolytica. We found that the optimum lipase yield under repeated methanol induction after 120 h was 32866 U/L, 28271 U/L and 21978 U/L for Lip C, Lip A and Lip 11 respectively. In addition, we found that a single dose of methyl ester supported higher production than repeated methanol induction. Among various methyl esters tested, methyl oleate (0.5%) caused 1.2 fold higher yield for LipA and LipC and 1.4 fold for Lip11 after 120 h of induction. Sequential utilization of methanol and oleic acid by P. pastoris was observed and was supported by differential peroxisome proliferation studies by transmission electron microscopy. Our study identifies a novel strategy of using methyl esters as slow release methanol source during lipase expression. PMID:25170843

Integrated bioprocess for conversion of gaseous substrates to liquids

PubMed Central

Hu, Peng; Chakraborty, Sagar; Kumar, Amit; Woolston, Benjamin; Liu, Hongjuan; Emerson, David; Stephanopoulos, Gregory

2016-01-01

In the quest for inexpensive feedstocks for the cost-effective production of liquid fuels, we have examined gaseous substrates that could be made available at low cost and sufficiently large scale for industrial fuel production. Here we introduce a new bioconversion scheme that effectively converts syngas, generated from gasification of coal, natural gas, or biomass, into lipids that can be used for biodiesel production. We present an integrated conversion method comprising a two-stage system. In the first stage, an anaerobic bioreactor converts mixtures of gases of CO2 and CO or H2 to acetic acid, using the anaerobic acetogen Moorella thermoacetica. The acetic acid product is fed as a substrate to a second bioreactor, where it is converted aerobically into lipids by an engineered oleaginous yeast, Yarrowia lipolytica. We first describe the process carried out in each reactor and then present an integrated system that produces microbial oil, using synthesis gas as input. The integrated continuous bench-scale reactor system produced 18 g/L of C16-C18 triacylglycerides directly from synthesis gas, with an overall productivity of 0.19 g⋅L−1⋅h−1 and a lipid content of 36%. Although suboptimal relative to the performance of the individual reactor components, the presented integrated system demonstrates the feasibility of substantial net fixation of carbon dioxide and conversion of gaseous feedstocks to lipids for biodiesel production. The system can be further optimized to approach the performance of its individual units so that it can be used for the economical conversion of waste gases from steel mills to valuable liquid fuels for transportation. PMID:26951649

[Biochemical failure after curative treatment for localized prostate cancer].

PubMed

Zouhair, Abderrahim; Jichlinski, Patrice; Mirimanoff, René-Olivier

2005-12-07

Biochemical failure after curative treatment for localized prostate cancer is frequent. The diagnosis of biochemical failure is clear when PSA levels rise after radical prostatectomy, but may be more difficult after external beam radiation therapy. The main difficulty once biochemical failure is diagnosed is to distinguish between local and distant failure, given the low sensitivity of standard work-up exams. Metabolic imaging techniques currently under evaluation may in the future help us to localize the site of failures. There are several therapeutic options depending on the initial curative treatment, each with morbidity risks that should be considered in multidisciplinary decision-making.

Know-how and know-why in biochemical engineering.

PubMed

von Stockar, U; Valentinotti, S; Marison, I; Cannizzaro, C; Herwig, C

2003-08-01

This contribution analyzes the position of biochemical engineering in general and bioprocess engineering particularly in the force fields between fundamental science and applications, and between academia and industry. By using culture technology as an example, it can be shown that bioprocess engineering has moved slowly but steadily from an empirical art concerned with mainly know-how to a science elucidating the know-why of culture behavior. Highly powerful monitoring tools enable biochemical engineers to understand and explain quantitatively the activity of cellular culture on a metabolic basis. Among these monitoring tools are not just semi-online analyses of culture broth by HPLC, GC and FIA, but, increasingly, also noninvasive methods such as midrange IR, Raman and capacitance spectroscopy, as well as online calorimetry. The detailed and quantitative insight into the metabolome and the fluxome that bioprocess engineers are establishing offers an unprecedented opportunity for building bridges between molecular biology and engineering biosciences. Thus, one of the major tasks of biochemical engineering sciences is not developing new know-how for industrial applications, but elucidating the know-why in biochemical engineering by conducting research on the underlying scientific fundamentals.

Recommendations for terminology and databases for biochemical thermodynamics.

PubMed

Alberty, Robert A; Cornish-Bowden, Athel; Goldberg, Robert N; Hammes, Gordon G; Tipton, Keith; Westerhoff, Hans V

2011-05-01

Chemical equations are normally written in terms of specific ionic and elemental species and balance atoms of elements and electric charge. However, in a biochemical context it is usually better to write them with ionic reactants expressed as totals of species in equilibrium with each other. This implies that atoms of elements assumed to be at fixed concentrations, such as hydrogen at a specified pH, should not be balanced in a biochemical equation used for thermodynamic analysis. However, both kinds of equations are needed in biochemistry. The apparent equilibrium constant K' for a biochemical reaction is written in terms of such sums of species and can be used to calculate standard transformed Gibbs energies of reaction Δ(r)G'°. This property for a biochemical reaction can be calculated from the standard transformed Gibbs energies of formation Δ(f)G(i)'° of reactants, which can be calculated from the standard Gibbs energies of formation of species Δ(f)G(j)° and measured apparent equilibrium constants of enzyme-catalyzed reactions. Tables of Δ(r)G'° of reactions and Δ(f)G(i)'° of reactants as functions of pH and temperature are available on the web, as are functions for calculating these properties. Biochemical thermodynamics is also important in enzyme kinetics because apparent equilibrium constant K' can be calculated from experimentally determined kinetic parameters when initial velocities have been determined for both forward and reverse reactions. Specific recommendations are made for reporting experimental results in the literature. Copyright © 2011 Elsevier B.V. All rights reserved.

Conditions for duality between fluxes and concentrations in biochemical networks

PubMed Central

Fleming, Ronan M.T.; Vlassis, Nikos; Thiele, Ines; Saunders, Michael A.

2016-01-01

Mathematical and computational modelling of biochemical networks is often done in terms of either the concentrations of molecular species or the fluxes of biochemical reactions. When is mathematical modelling from either perspective equivalent to the other? Mathematical duality translates concepts, theorems or mathematical structures into other concepts, theorems or structures, in a one-to-one manner. We present a novel stoichiometric condition that is necessary and sufficient for duality between unidirectional fluxes and concentrations. Our numerical experiments, with computational models derived from a range of genome-scale biochemical networks, suggest that this flux-concentration duality is a pervasive property of biochemical networks. We also provide a combinatorial characterisation that is sufficient to ensure flux-concentration duality. The condition prescribes that, for every two disjoint sets of molecular species, there is at least one reaction complex that involves species from only one of the two sets. When unidirectional fluxes and molecular species concentrations are dual vectors, this implies that the behaviour of the corresponding biochemical network can be described entirely in terms of either concentrations or unidirectional fluxes. PMID:27345817

Conditions for duality between fluxes and concentrations in biochemical networks

DOE PAGES

Fleming, Ronan M. T.; Vlassis, Nikos; Thiele, Ines; ...

2016-06-23

Mathematical and computational modelling of biochemical networks is often done in terms of either the concentrations of molecular species or the fluxes of biochemical reactions. When is mathematical modelling from either perspective equivalent to the other? Mathematical duality translates concepts, theorems or mathematical structures into other concepts, theorems or structures, in a one-to-one manner. We present a novel stoichiometric condition that is necessary and sufficient for duality between unidirectional fluxes and concentrations. Our numerical experiments, with computational models derived from a range of genome-scale biochemical networks, suggest that this flux-concentration duality is a pervasive property of biochemical networks. We alsomore » provide a combinatorial characterisation that is sufficient to ensure flux-concentration duality.The condition prescribes that, for every two disjoint sets of molecular species, there is at least one reaction complex that involves species from only one of the two sets. When unidirectional fluxes and molecular species concentrations are dual vectors, this implies that the behaviour of the corresponding biochemical network can be described entirely in terms of either concentrations or unidirectional fluxes« less

Conditions for duality between fluxes and concentrations in biochemical networks

DOE Office of Scientific and Technical Information (OSTI.GOV)

Fleming, Ronan M. T.; Vlassis, Nikos; Thiele, Ines

Mathematical and computational modelling of biochemical networks is often done in terms of either the concentrations of molecular species or the fluxes of biochemical reactions. When is mathematical modelling from either perspective equivalent to the other? Mathematical duality translates concepts, theorems or mathematical structures into other concepts, theorems or structures, in a one-to-one manner. We present a novel stoichiometric condition that is necessary and sufficient for duality between unidirectional fluxes and concentrations. Our numerical experiments, with computational models derived from a range of genome-scale biochemical networks, suggest that this flux-concentration duality is a pervasive property of biochemical networks. We alsomore » provide a combinatorial characterisation that is sufficient to ensure flux-concentration duality.The condition prescribes that, for every two disjoint sets of molecular species, there is at least one reaction complex that involves species from only one of the two sets. When unidirectional fluxes and molecular species concentrations are dual vectors, this implies that the behaviour of the corresponding biochemical network can be described entirely in terms of either concentrations or unidirectional fluxes« less

A Course in... Biochemical Engineering.

ERIC Educational Resources Information Center

Ng, Terry K-L.; And Others

1988-01-01

Describes a chemical engineering course for senior undergraduates and first year graduate students in biochemical engineering. Discusses five experiments used in the course: aseptic techniques, dissolved oxygen measurement, oxygen uptake by yeast, continuous sterilization, and cultivation of microorganisms. (MVL)

40 CFR 158.2050 - Biochemical pesticides human health assessment data requirements table.

Code of Federal Regulations, 2011 CFR

2011-07-01

... 40 Protection of Environment 24 2011-07-01 2011-07-01 false Biochemical pesticides human health... § 158.2050 Biochemical pesticides human health assessment data requirements table. (a) General. (1... table shows the data requirements for biochemical pesticides human health assessment. The test notes are...

40 CFR 158.2050 - Biochemical pesticides human health assessment data requirements table.

Code of Federal Regulations, 2010 CFR

2010-07-01

... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Biochemical pesticides human health... § 158.2050 Biochemical pesticides human health assessment data requirements table. (a) General. (1... table shows the data requirements for biochemical pesticides human health assessment. The test notes are...

40 CFR 158.2050 - Biochemical pesticides human health assessment data requirements table.

Code of Federal Regulations, 2013 CFR

2013-07-01

... 40 Protection of Environment 25 2013-07-01 2013-07-01 false Biochemical pesticides human health... § 158.2050 Biochemical pesticides human health assessment data requirements table. (a) General. (1... table shows the data requirements for biochemical pesticides human health assessment. The test notes are...

40 CFR 158.2050 - Biochemical pesticides human health assessment data requirements table.

Code of Federal Regulations, 2012 CFR

2012-07-01

... 40 Protection of Environment 25 2012-07-01 2012-07-01 false Biochemical pesticides human health... § 158.2050 Biochemical pesticides human health assessment data requirements table. (a) General. (1... table shows the data requirements for biochemical pesticides human health assessment. The test notes are...

40 CFR 158.2050 - Biochemical pesticides human health assessment data requirements table.

Code of Federal Regulations, 2014 CFR

2014-07-01

... 40 Protection of Environment 24 2014-07-01 2014-07-01 false Biochemical pesticides human health... § 158.2050 Biochemical pesticides human health assessment data requirements table. (a) General. (1... table shows the data requirements for biochemical pesticides human health assessment. The test notes are...

Development of a new first-aid biochemical detector

NASA Astrophysics Data System (ADS)

Hu, Jingfei; Liao, Haiyang; Su, Shilin; Ding, Hao; Liu, Suquan

2016-10-01

The traditional biochemical detector exhibits poor adaptability, inconvenient carrying and slow detection, which can't meet the needs of first-aid under field condition like natural or man-made disasters etc. Therefore a scheme of first-aid biochemical detector based on MOMES Micro Spectrometer, UV LED and Photodiode was proposed. An optical detection structure combined continuous spectrum sweep with fixed wavelength measurement was designed, which adopted mobile detection optical path consisting of Micro Spectrometer and Halogen Lamp to detect Chloride (Cl-), Creatinine (Cre), Glucose (Glu), Hemoglobin (Hb). The UV LED and Photodiode were designed to detect Potassium (K-), Carbon dioxide (CO2), Sodium (Na+). According to the field diagnosis and treatment requirements, we designed the embedded control hardware circuit and software system, the prototype of first-aid biochemical detector was developed and the clinical trials were conducted. Experimental results show that the sample's absorbance repeatability is less than 2%, the max coefficient of variation (CV) in the batch repeatability test of all 7 biochemical parameters in blood samples is 4.68%, less than the clinical requirements 10%, the correlation coefficient (R2) in the clinical contrast test with AU5800 is almost greater than 0.97. To sum up, the prototype meets the requirements of clinical application.

Multidimensional biochemical information processing of dynamical patterns

NASA Astrophysics Data System (ADS)

Hasegawa, Yoshihiko

2018-02-01

Cells receive signaling molecules by receptors and relay information via sensory networks so that they can respond properly depending on the type of signal. Recent studies have shown that cells can extract multidimensional information from dynamical concentration patterns of signaling molecules. We herein study how biochemical systems can process multidimensional information embedded in dynamical patterns. We model the decoding networks by linear response functions, and optimize the functions with the calculus of variations to maximize the mutual information between patterns and output. We find that, when the noise intensity is lower, decoders with different linear response functions, i.e., distinct decoders, can extract much information. However, when the noise intensity is higher, distinct decoders do not provide the maximum amount of information. This indicates that, when transmitting information by dynamical patterns, embedding information in multiple patterns is not optimal when the noise intensity is very large. Furthermore, we explore the biochemical implementations of these decoders using control theory and demonstrate that these decoders can be implemented biochemically through the modification of cascade-type networks, which are prevalent in actual signaling pathways.

Stoichiometric network theory for nonequilibrium biochemical systems.

PubMed

Qian, Hong; Beard, Daniel A; Liang, Shou-dan

2003-02-01

We introduce the basic concepts and develop a theory for nonequilibrium steady-state biochemical systems applicable to analyzing large-scale complex isothermal reaction networks. In terms of the stoichiometric matrix, we demonstrate both Kirchhoff's flux law sigma(l)J(l)=0 over a biochemical species, and potential law sigma(l) mu(l)=0 over a reaction loop. They reflect mass and energy conservation, respectively. For each reaction, its steady-state flux J can be decomposed into forward and backward one-way fluxes J = J+ - J-, with chemical potential difference deltamu = RT ln(J-/J+). The product -Jdeltamu gives the isothermal heat dissipation rate, which is necessarily non-negative according to the second law of thermodynamics. The stoichiometric network theory (SNT) embodies all of the relevant fundamental physics. Knowing J and deltamu of a biochemical reaction, a conductance can be computed which directly reflects the level of gene expression for the particular enzyme. For sufficiently small flux a linear relationship between J and deltamu can be established as the linear flux-force relation in irreversible thermodynamics, analogous to Ohm's law in electrical circuits.

Multidimensional biochemical information processing of dynamical patterns.

PubMed

Hasegawa, Yoshihiko

2018-02-01

Cells receive signaling molecules by receptors and relay information via sensory networks so that they can respond properly depending on the type of signal. Recent studies have shown that cells can extract multidimensional information from dynamical concentration patterns of signaling molecules. We herein study how biochemical systems can process multidimensional information embedded in dynamical patterns. We model the decoding networks by linear response functions, and optimize the functions with the calculus of variations to maximize the mutual information between patterns and output. We find that, when the noise intensity is lower, decoders with different linear response functions, i.e., distinct decoders, can extract much information. However, when the noise intensity is higher, distinct decoders do not provide the maximum amount of information. This indicates that, when transmitting information by dynamical patterns, embedding information in multiple patterns is not optimal when the noise intensity is very large. Furthermore, we explore the biochemical implementations of these decoders using control theory and demonstrate that these decoders can be implemented biochemically through the modification of cascade-type networks, which are prevalent in actual signaling pathways.

Thermodynamics of stoichiometric biochemical networks in living systems far from equilibrium.

PubMed

Qian, Hong; Beard, Daniel A

2005-04-22

The principles of thermodynamics apply to both equilibrium and nonequilibrium biochemical systems. The mathematical machinery of the classic thermodynamics, however, mainly applies to systems in equilibrium. We introduce a thermodynamic formalism for the study of metabolic biochemical reaction (open, nonlinear) networks in both time-dependent and time-independent nonequilibrium states. Classical concepts in equilibrium thermodynamics-enthalpy, entropy, and Gibbs free energy of biochemical reaction systems-are generalized to nonequilibrium settings. Chemical motive force, heat dissipation rate, and entropy production (creation) rate, key concepts in nonequilibrium systems, are introduced. Dynamic equations for the thermodynamic quantities are presented in terms of the key observables of a biochemical network: stoichiometric matrix Q, reaction fluxes J, and chemical potentials of species mu without evoking empirical rate laws. Energy conservation and the Second Law are established for steady-state and dynamic biochemical networks. The theory provides the physiochemical basis for analyzing large-scale metabolic networks in living organisms.

Biochemical upgrading of oils

DOEpatents

Premuzic, Eugene T.; Lin, Mow S.

1999-01-12

A process for biochemical conversion of heavy crude oils is provided. The process includes contacting heavy crude oils with adapted biocatalysts. The resulting upgraded oil shows, a relative increase in saturated hydrocarbons, emulsions and oxygenates and a decrease in compounds containing in organic sulfur, organic nitrogen and trace metals. Adapted microorganisms which have been modified under challenged growth processes are also disclosed.

Biochemical upgrading of oils

DOEpatents

Premuzic, E.T.; Lin, M.S.

1999-01-12

A process for biochemical conversion of heavy crude oils is provided. The process includes contacting heavy crude oils with adapted biocatalysts. The resulting upgraded oil shows, a relative increase in saturated hydrocarbons, emulsions and oxygenates and a decrease in compounds containing organic sulfur, organic nitrogen and trace metals. Adapted microorganisms which have been modified under challenged growth processes are also disclosed. 121 figs.

40 CFR 158.2070 - Biochemical pesticides product performance data requirements.

Code of Federal Regulations, 2012 CFR

2012-07-01

... efficacy data unless the pesticide product bears a claim to control public health pests, such as pest... 40 Protection of Environment 25 2012-07-01 2012-07-01 false Biochemical pesticides product... AGENCY (CONTINUED) PESTICIDE PROGRAMS DATA REQUIREMENTS FOR PESTICIDES Biochemical Pesticides § 158.2070...

40 CFR 158.2070 - Biochemical pesticides product performance data requirements.

Code of Federal Regulations, 2011 CFR

2011-07-01

... efficacy data unless the pesticide product bears a claim to control public health pests, such as pest... 40 Protection of Environment 24 2011-07-01 2011-07-01 false Biochemical pesticides product... AGENCY (CONTINUED) PESTICIDE PROGRAMS DATA REQUIREMENTS FOR PESTICIDES Biochemical Pesticides § 158.2070...

40 CFR 158.2070 - Biochemical pesticides product performance data requirements.

Code of Federal Regulations, 2014 CFR

2014-07-01

... efficacy data unless the pesticide product bears a claim to control public health pests, such as pest... 40 Protection of Environment 24 2014-07-01 2014-07-01 false Biochemical pesticides product... AGENCY (CONTINUED) PESTICIDE PROGRAMS DATA REQUIREMENTS FOR PESTICIDES Biochemical Pesticides § 158.2070...

40 CFR 158.2070 - Biochemical pesticides product performance data requirements.

Code of Federal Regulations, 2013 CFR

2013-07-01

... efficacy data unless the pesticide product bears a claim to control public health pests, such as pest... 40 Protection of Environment 25 2013-07-01 2013-07-01 false Biochemical pesticides product... AGENCY (CONTINUED) PESTICIDE PROGRAMS DATA REQUIREMENTS FOR PESTICIDES Biochemical Pesticides § 158.2070...

40 CFR 158.2070 - Biochemical pesticides product performance data requirements.

Code of Federal Regulations, 2010 CFR

2010-07-01

... efficacy data unless the pesticide product bears a claim to control public health pests, such as pest... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Biochemical pesticides product... AGENCY (CONTINUED) PESTICIDE PROGRAMS DATA REQUIREMENTS FOR PESTICIDES Biochemical Pesticides § 158.2070...

Biochemical correlates in an animal model of depression

DOE Office of Scientific and Technical Information (OSTI.GOV)

Johnson, J.O.

1986-01-01

A valid animal model of depression was used to explore specific adrenergic receptor differences between rats exhibiting aberrant behavior and control groups. Preliminary experiments revealed a distinct upregulation of hippocampal beta-receptors (as compared to other brain regions) in those animals acquiring a response deficit as a result of exposure to inescapable footshock. Concurrent studies using standard receptor binding techniques showed no large changes in the density of alpha-adrenergic, serotonergic, or dopaminergic receptor densities. This led to the hypothesis that the hippocampal beta-receptor in responses deficient animals could be correlated with the behavioral changes seen after exposure to the aversive stimulus.more » Normalization of the behavior through the administration of antidepressants could be expected to reverse the biochemical changes if these are related to the mechanism of action of antidepressant drugs. This study makes three important points: (1) there is a relevant biochemical change in the hippocampus of response deficient rats which occurs in parallel to a well-defined behavior, (2) the biochemical and behavioral changes are normalized by antidepressant treatments exhibiting both serotonergic and adrenergic mechanisms of action, and (3) the mode of action of antidepressants in this model is probably a combination of serotonergic and adrenergic influences modulating the hippocampal beta-receptor. These results are discussed in relation to anatomical and biochemical aspects of antidepressant action.« less

BNDB - the Biochemical Network Database.

PubMed

Küntzer, Jan; Backes, Christina; Blum, Torsten; Gerasch, Andreas; Kaufmann, Michael; Kohlbacher, Oliver; Lenhof, Hans-Peter

2007-10-02

Technological advances in high-throughput techniques and efficient data acquisition methods have resulted in a massive amount of life science data. The data is stored in numerous databases that have been established over the last decades and are essential resources for scientists nowadays. However, the diversity of the databases and the underlying data models make it difficult to combine this information for solving complex problems in systems biology. Currently, researchers typically have to browse several, often highly focused, databases to obtain the required information. Hence, there is a pressing need for more efficient systems for integrating, analyzing, and interpreting these data. The standardization and virtual consolidation of the databases is a major challenge resulting in a unified access to a variety of data sources. We present the Biochemical Network Database (BNDB), a powerful relational database platform, allowing a complete semantic integration of an extensive collection of external databases. BNDB is built upon a comprehensive and extensible object model called BioCore, which is powerful enough to model most known biochemical processes and at the same time easily extensible to be adapted to new biological concepts. Besides a web interface for the search and curation of the data, a Java-based viewer (BiNA) provides a powerful platform-independent visualization and navigation of the data. BiNA uses sophisticated graph layout algorithms for an interactive visualization and navigation of BNDB. BNDB allows a simple, unified access to a variety of external data sources. Its tight integration with the biochemical network library BN++ offers the possibility for import, integration, analysis, and visualization of the data. BNDB is freely accessible at http://www.bndb.org.

Measurement of biochemical oxygen demand of the leachates.

PubMed

Fulazzaky, Mohamad Ali

2013-06-01

Biochemical oxygen demand (BOD) of the leachates originally from the different types of landfill sites was studied based on the data measured using the two manometric methods. The measurements of BOD using the dilution method were carried out to assess the typical physicochemical and biological characteristics of the leachates together with some other parameters. The linear regression analysis was used to predict rate constants for biochemical reactions and ultimate BOD values of the different leachates. The rate of a biochemical reaction implicated in microbial biodegradation of pollutants depends on the leachate characteristics, mass of contaminant in the leachate, and nature of the leachate. Character of leachate samples for BOD analysis of using the different methods may differ significantly during the experimental period, resulting in different BOD values. This work intends to verify effect of the different dilutions for the manometric method tests on the BOD concentrations of the leachate samples to contribute to the assessment of reaction rate and microbial consumption of oxygen.

40 CFR 158.2040 - Biochemical pesticides residue data requirements table.

Code of Federal Regulations, 2014 CFR

2014-07-01

... for biochemical agents in the biochemical human health assessment data requirements, § 158.2050. 2... be available for human or livestock consumption. 13. Data on fish are required for all pesticides... for human consumption. 14. Data are required when a pesticide is to be applied directly to water that...

40 CFR 158.2040 - Biochemical pesticides residue data requirements table.

Code of Federal Regulations, 2011 CFR

2011-07-01

... for biochemical agents in the biochemical human health assessment data requirements, § 158.2050. 2... be available for human or livestock consumption. 13. Data on fish are required for all pesticides... for human consumption. 14. Data are required when a pesticide is to be applied directly to water that...

40 CFR 158.2040 - Biochemical pesticides residue data requirements table.

Code of Federal Regulations, 2012 CFR

2012-07-01

... for biochemical agents in the biochemical human health assessment data requirements, § 158.2050. 2... be available for human or livestock consumption. 13. Data on fish are required for all pesticides... for human consumption. 14. Data are required when a pesticide is to be applied directly to water that...

40 CFR 158.2040 - Biochemical pesticides residue data requirements table.

Code of Federal Regulations, 2013 CFR

2013-07-01

... for biochemical agents in the biochemical human health assessment data requirements, § 158.2050. 2... be available for human or livestock consumption. 13. Data on fish are required for all pesticides... for human consumption. 14. Data are required when a pesticide is to be applied directly to water that...

40 CFR 158.2040 - Biochemical pesticides residue data requirements table.

Code of Federal Regulations, 2010 CFR

2010-07-01

... for biochemical agents in the biochemical human health assessment data requirements, § 158.2050. 2... be available for human or livestock consumption. 13. Data on fish are required for all pesticides... for human consumption. 14. Data are required when a pesticide is to be applied directly to water that...

40 CFR 158.2084 - Experimental use permit biochemical pesticides nontarget organisms and environmental fate data...

Code of Federal Regulations, 2013 CFR

2013-07-01

... test notes are shown in paragraph (e) of this section. Table—EUP Biochemical Pesticides Nontarget... 40 Protection of Environment 25 2013-07-01 2013-07-01 false Experimental use permit biochemical... FOR PESTICIDES Biochemical Pesticides § 158.2084 Experimental use permit biochemical pesticides...

40 CFR 158.2084 - Experimental use permit biochemical pesticides nontarget organisms and environmental fate data...

Code of Federal Regulations, 2011 CFR

2011-07-01

... test notes are shown in paragraph (e) of this section. Table—EUP Biochemical Pesticides Nontarget... 40 Protection of Environment 24 2011-07-01 2011-07-01 false Experimental use permit biochemical... FOR PESTICIDES Biochemical Pesticides § 158.2084 Experimental use permit biochemical pesticides...

40 CFR 158.2084 - Experimental use permit biochemical pesticides nontarget organisms and environmental fate data...

Code of Federal Regulations, 2012 CFR

2012-07-01

... test notes are shown in paragraph (e) of this section. Table—EUP Biochemical Pesticides Nontarget... 40 Protection of Environment 25 2012-07-01 2012-07-01 false Experimental use permit biochemical... FOR PESTICIDES Biochemical Pesticides § 158.2084 Experimental use permit biochemical pesticides...

40 CFR 158.2084 - Experimental use permit biochemical pesticides nontarget organisms and environmental fate data...

Code of Federal Regulations, 2014 CFR

2014-07-01

... test notes are shown in paragraph (e) of this section. Table—EUP Biochemical Pesticides Nontarget... 40 Protection of Environment 24 2014-07-01 2014-07-01 false Experimental use permit biochemical... FOR PESTICIDES Biochemical Pesticides § 158.2084 Experimental use permit biochemical pesticides...

[Biochemical indicators of anaphylactic shock and the application in forensic medicine].

PubMed

Mi, Li; Chen, Jie; Gao, Wei-Min; Du, Zhong-Bo; Cao, Zhi-Peng; Zhang, Yuan; Zhu, Bao-Li

2014-04-01

Fatal anaphylactic shock is common in forensic practice. However, it is difficult to diagnose for lacking specific pathological and morphologic changes in forensic autopsy. The application of some biochemical indicators is of great significance. This paper reviews the biological characteristics of some biochemical indicators and detection methods. The forensic application, problems and prospects of these indicators are also introduced in details. The stable biochemical indicators, IgE, tryptase and chymase, show great potential and advantages in the identification of fatal anaphylactic shock in forensic medicine.

Causal correlation of foliar biochemical concentrations with AVIRIS spectra using forced entry linear regression

NASA Technical Reports Server (NTRS)

Dawson, Terence P.; Curran, Paul J.; Kupiec, John A.

1995-01-01

A major goal of airborne imaging spectrometry is to estimate the biochemical composition of vegetation canopies from reflectance spectra. Remotely-sensed estimates of foliar biochemical concentrations of forests would provide valuable indicators of ecosystem function at regional and eventually global scales. Empirical research has shown a relationship exists between the amount of radiation reflected from absorption features and the concentration of given biochemicals in leaves and canopies (Matson et al., 1994, Johnson et al., 1994). A technique commonly used to determine which wavelengths have the strongest correlation with the biochemical of interest is unguided (stepwise) multiple regression. Wavelengths are entered into a multivariate regression equation, in their order of importance, each contributing to the reduction of the variance in the measured biochemical concentration. A significant problem with the use of stepwise regression for determining the correlation between biochemical concentration and spectra is that of 'overfitting' as there are significantly more wavebands than biochemical measurements. This could result in the selection of wavebands which may be more accurately attributable to noise or canopy effects. In addition, there is a real problem of collinearity in that the individual biochemical concentrations may covary. A strong correlation between the reflectance at a given wavelength and the concentration of a biochemical of interest, therefore, may be due to the effect of another biochemical which is closely related. Furthermore, it is not always possible to account for potentially suitable waveband omissions in the stepwise selection procedure. This concern about the suitability of stepwise regression has been identified and acknowledged in a number of recent studies (Wessman et al., 1988, Curran, 1989, Curran et al., 1992, Peterson and Hubbard, 1992, Martine and Aber, 1994, Kupiec, 1994). These studies have pointed to the lack of a physical

Physiological and molecular biochemical mechanisms of bile formation

PubMed Central

Reshetnyak, Vasiliy Ivanovich

2013-01-01

This review considers the physiological and molecular biochemical mechanisms of bile formation. The composition of bile and structure of a bile canaliculus, biosynthesis and conjugation of bile acids, bile phospholipids, formation of bile micellar structures, and enterohepatic circulation of bile acids are described. In general, the review focuses on the molecular physiology of the transporting systems of the hepatocyte sinusoidal and apical membranes. Knowledge of physiological and biochemical basis of bile formation has implications for understanding the mechanisms of development of pathological processes, associated with diseases of the liver and biliary tract. PMID:24259965

New biochemical markers: from bench to bedside.

PubMed

Zaninotto, Martina; Mion, Monica Maria; Novello, Enrica; Altinier, Sara; Plebani, Mario

2007-05-01

Evaluation of patients presenting to hospital with chest pain or other signs or symptoms suggesting acute coronary syndrome (ACS) is problematic, time-consuming and sometimes expensive, even if new biochemical markers, such as troponins, have improved the ability to detect cardiac injury. However, patients with normal troponin values are not necessarily risk-free for major cardiac events. Recent investigations indicate that the overall patient risk may be assessed earlier than before, thanks to new knowledge acquired concerning the pathobiology of atherosclerosis and molecular events involved in the progression of disease, thus allowing the development of new biochemical markers. Some selected markers are released during the different phases of development of cardiovascular disease and may be useful for the diagnosis of patients with cardiovascular disease. In particular, the identification of emerging markers that provide relevant information on the inflammatory process, and the development of biomarkers whose circulating concentrations suggest the status of plaque instability and rupture, seems to be of particular value in prognosis and risk stratification. The overall expectations for a cardiovascular biochemical marker are not only its biological plausibility but also the availability at a reasonable cost of rapid, high quality assays, and their correct interpretation by clinicians using optimal cut-offs. The crossing from bench to bedside for each new marker discovered, must be associated with concurrent advances in the characterization of analytical features and the development of routine assay, in the assessment of analytical performance and in interpretative reporting of test results as well as in the training of physicians to use the array of biomarkers available appropriately and to interpret them correctly. This approach calls for the coordinated support of clinicians, technology experts, statisticians and the industry so that new biochemical

Optimal Information Processing in Biochemical Networks

NASA Astrophysics Data System (ADS)

Wiggins, Chris

2012-02-01

A variety of experimental results over the past decades provide examples of near-optimal information processing in biological networks, including in biochemical and transcriptional regulatory networks. Computing information-theoretic quantities requires first choosing or computing the joint probability distribution describing multiple nodes in such a network --- for example, representing the probability distribution of finding an integer copy number of each of two interacting reactants or gene products while respecting the `intrinsic' small copy number noise constraining information transmission at the scale of the cell. I'll given an overview of some recent analytic and numerical work facilitating calculation of such joint distributions and the associated information, which in turn makes possible numerical optimization of information flow in models of noisy regulatory and biochemical networks. Illustrating cases include quantification of form-function relations, ideal design of regulatory cascades, and response to oscillatory driving.

Improved biochemical preservation of lung slices during cold storage.

PubMed

Bull, D A; Connors, R C; Reid, B B; Albanil, A; Stringham, J C; Karwande, S V

2000-05-15

Development of lung preservation solutions typically requires whole-organ models which are animal and labor intensive. These models rely on physiologic rather than biochemical endpoints, making accurate comparison of the relative efficacy of individual solution components difficult. We hypothesized that lung slices could be used to assess preservation of biochemical function during cold storage. Whole rat lungs were precision cut into slices with a thickness of 500 microm and preserved at 4 degrees C in the following solutions: University of Wisconsin (UW), Euro-Collins (EC), low-potassium-dextran (LPD), Kyoto (K), normal saline (NS), or a novel lung preservation solution (NPS) developed using this model. Lung biochemical function was assessed by ATP content (etamol ATP/mg wet wt) and capacity for protein synthesis (cpm/mg protein) immediately following slicing (0 h) and at 6, 12, 18, and 24 h of cold storage. Six slices were assayed at each time point for each solution. The data were analyzed using analysis of variance and are presented as means +/- SD. ATP content was significantly higher in the lung slices stored in NPS compared with all other solutions at each time point (P < 0.0001). Protein synthesis was significantly higher in the lung slices stored in NPS compared with all other solutions at 6, 12, and 18 h of preservation (P < 0.05). This lung slice model allows the rapid and efficient screening of lung preservation solutions and their components using quantifiable biochemical endpoints. Using this model, we have developed a novel solution that improves the biochemical preservation of lung slices during cold storage. Copyright 2000 Academic Press.

Biosensors and bioelectronics on smartphone for portable biochemical detection.

PubMed

Zhang, Diming; Liu, Qingjun

2016-01-15

Smartphone has been widely integrated with sensors, such as test strips, sensor chips, and hand-held detectors, for biochemical detections due to its portability and ubiquitous availability. Utilizing built-in function modules, smartphone is often employed as controller, analyzer, and displayer for rapid, real-time, and point-of-care monitoring, which can significantly simplify design and reduce cost of the detecting systems. This paper presents a review of biosensors and bioelectronics on smartphone for portable biochemical detections. The biosensors and bioelectronics based on smartphone can mainly be classified into biosensors using optics, surface plasmon resonance, electrochemistry, and near-field communication. The developments of these biosensors and bioelectronics on smartphone are reviewed along with typical biochemical detecting cases. Sensor strategies, detector attachments, and coupling methods are highlighted to show designs of the compact, lightweight, and low-cost sensor systems. The performances and advantages of these designs are introduced with their applications in healthcare diagnosis, environment monitoring, and food evaluation. With advances in micro-manufacture, sensor technology, and miniaturized electronics, biosensor and bioelectronic devices on smartphone can be used to perform biochemical detections as common and convenient as electronic tag readout in foreseeable future. Copyright © 2015 Elsevier B.V. All rights reserved.

Designing Epigenome Editors: Considerations of Biochemical and Locus Specificities.

PubMed

Sen, Dilara; Keung, Albert J

2018-01-01

The advent of locus-specific protein recruitment technologies has enabled a new class of studies in chromatin biology. Epigenome editors enable biochemical modifications of chromatin at almost any specific endogenous locus. Their locus specificity unlocks unique information including the functional roles of distinct modifications at specific genomic loci. Given the growing interest in using these tools for biological and translational studies, there are many specific design considerations depending on the scientific question or clinical need. Here we present and discuss important design considerations and challenges regarding the biochemical and locus specificities of epigenome editors. These include how to account for the complex biochemical diversity of chromatin; control for potential interdependency of epigenome editors and their resultant modifications; avoid sequestration effects; quantify the locus specificity of epigenome editors; and improve locus specificity by considering concentration, affinity, avidity, and sequestration effects.

[Interpretation of false positive results of biochemical prenatal tests].

PubMed

Sieroszewski, Piotr; Słowakiewicz, Katarzyna; Perenc, Małgorzata

2010-03-01

Modern, non-invasive prenatal diagnostics based on biochemical and ultrasonographic markers of fetal defects allows us to calculate the risk of fetal chromosomal aneuploidies with high sensitivity and specificity An introduction of biochemical, non-invasive prenatal tests turned out to result in frequent false positive results of these tests in cases when invasive diagnostics does not confirm fetal defects. However prospective analysis of these cases showed numerous complications in the third trimester of the pregnancies.

Advanced Nanoporous Materials for Micro-Gravimetric Sensing to Trace-Level Bio/Chemical Molecules

PubMed Central

Xu, Pengcheng; Li, Xinxin; Yu, Haitao; Xu, Tiegang

2014-01-01

Functionalized nanoporous materials have been developed recently as bio/chemical sensing materials. Due to the huge specific surface of the nano-materials for molecular adsorption, high hopes have been placed on gravimetric detection with micro/nano resonant cantilevers for ultra-sensitive sensing of low-concentration bio/chemical substances. In order to enhance selectivity of the gravimetric resonant sensors to the target molecules, it is crucial to modify specific groups onto the pore-surface of the nano-materials. By loading the nanoporous sensing material onto the desired region of the mass-type transducers like resonant cantilevers, the micro-gravimetric bio/chemical sensors can be formed. Recently, such micro-gravimetric bio/chemical sensors have been successfully applied for rapid or on-the-spot detection of various bio/chemical molecules at the trace-concentration level. The applicable nanoporous sensing materials include mesoporous silica, zeolite, nanoporous graphene oxide (GO) and so on. This review article focuses on the recent achievements in design, preparation, functionalization and characterization of advanced nanoporous sensing materials for micro-gravimetric bio/chemical sensing. PMID:25313499

Complete integrability of information processing by biochemical reactions

NASA Astrophysics Data System (ADS)

Agliari, Elena; Barra, Adriano; Dello Schiavo, Lorenzo; Moro, Antonio

2016-11-01

Statistical mechanics provides an effective framework to investigate information processing in biochemical reactions. Within such framework far-reaching analogies are established among (anti-) cooperative collective behaviors in chemical kinetics, (anti-)ferromagnetic spin models in statistical mechanics and operational amplifiers/flip-flops in cybernetics. The underlying modeling - based on spin systems - has been proved to be accurate for a wide class of systems matching classical (e.g. Michaelis-Menten, Hill, Adair) scenarios in the infinite-size approximation. However, the current research in biochemical information processing has been focusing on systems involving a relatively small number of units, where this approximation is no longer valid. Here we show that the whole statistical mechanical description of reaction kinetics can be re-formulated via a mechanical analogy - based on completely integrable hydrodynamic-type systems of PDEs - which provides explicit finite-size solutions, matching recently investigated phenomena (e.g. noise-induced cooperativity, stochastic bi-stability, quorum sensing). The resulting picture, successfully tested against a broad spectrum of data, constitutes a neat rationale for a numerically effective and theoretically consistent description of collective behaviors in biochemical reactions.

Biochemical differences in ethnic groups in Durango, Mexico.

PubMed

Lares-Asseff, Ismael; Lujín-García, Azalia; Sosa-Macías, Martha; Lazalde-Ramos, Blanca; Loera-Castañeda, Veronica; Galaviz-Hernández, Carlos; Villanueva-Fierro, Ignacio

2012-01-01

The aim of this study was to assess biochemical differences between Tepehuano indigenous people, and Mennonite and Mestizo populations of Durango, Mexico. Our study involved 334 volunteers aged 15 to 80 years; 132 Mennonite and 130 Mestizo individuals from Nuevo Ideal Municipality and 72 Tepehuano indigenous people from Mezquital Durango were evaluated. A clinical history and fast determination of aspartate aminotransferase (AST), alanine aminotransferase (ALT), uric acid, urea and creatinine were performed on each studied case. Statistically significant differences between the three studied groups were found for age, weight and height (P < .05), with higher values observed in men. The highest plasma urea levels were found in Mennonite compared to Mestizo people, followed by the Tepehuano indigenous. Higher biochemical parameters were found in men (vs women) in the studied groups. The percentage of individuals with abnormal levels for AST, ALT and uric acid were higher in Tepehuano indigenous people than in Mestizo, whereas the urea and creatinine percentages were higher in Mestizo people. The differences found on biochemical tests, could be explained by differences in lifestyle such as diet and sanitary habits.

Complete integrability of information processing by biochemical reactions.

PubMed

Agliari, Elena; Barra, Adriano; Dello Schiavo, Lorenzo; Moro, Antonio

2016-11-04

Statistical mechanics provides an effective framework to investigate information processing in biochemical reactions. Within such framework far-reaching analogies are established among (anti-) cooperative collective behaviors in chemical kinetics, (anti-)ferromagnetic spin models in statistical mechanics and operational amplifiers/flip-flops in cybernetics. The underlying modeling - based on spin systems - has been proved to be accurate for a wide class of systems matching classical (e.g. Michaelis-Menten, Hill, Adair) scenarios in the infinite-size approximation. However, the current research in biochemical information processing has been focusing on systems involving a relatively small number of units, where this approximation is no longer valid. Here we show that the whole statistical mechanical description of reaction kinetics can be re-formulated via a mechanical analogy - based on completely integrable hydrodynamic-type systems of PDEs - which provides explicit finite-size solutions, matching recently investigated phenomena (e.g. noise-induced cooperativity, stochastic bi-stability, quorum sensing). The resulting picture, successfully tested against a broad spectrum of data, constitutes a neat rationale for a numerically effective and theoretically consistent description of collective behaviors in biochemical reactions.

Circadian Clocks: Unexpected Biochemical Cogs.

PubMed

Mori, Tetsuya; Mchaourab, Hassane; Johnson, Carl Hirschie

2015-10-05

A circadian oscillation can be reconstituted in vitro from three proteins that cycles with a period of ∼ 24 h. Two recent studies provide surprising biochemical answers to why this remarkable oscillator has such a long time constant and how it can switch effortlessly between alternating enzymatic modes. Copyright © 2015 Elsevier Ltd. All rights reserved.

Autonomous bio-chemical decontaminator (ABCD) against weapons of mass destruction

NASA Astrophysics Data System (ADS)

Hyacinthe, Berg P.

2006-05-01

The proliferation of weapons of mass destruction (WMD) and the use of such elements pose an eminent asymmetric threat with disastrous consequences to the national security of any nation. In particular, the use of biochemical warfare agents against civilians and unprotected troops in international conflicts or by terrorists against civilians is considered as a very peculiar threat. Accordingly, taking a quarantine-before-inhalation approach to biochemical warfare, the author introduces the notion of autonomous biochemical decontamination against WMD. In the unfortunate event of a biochemical attack, the apparatus proposed herein is intended to automatically detect, identify, and more importantly neutralize a biochemical threat. Along with warnings concerning a cyber-WMD nexus, various sections cover discussions on human senses and computer sensors, corroborating evidence related to detection and neutralization of chemical toxins, and cyber-assisted olfaction in stand alone, peer-to-peer, and network settings. In essence, the apparatus can be used in aviation and mass transit security to initiate mass decontamination by dispersing a decontaminant aerosol or to protect the public water supply against a potential bioterrorist attack. Future effort may involve a system-on-chip (SoC) embodiment of this apparatus that allows a safer environment for the emerging phenomenon of cyber-assisted olfaction and morph cell phones into ubiquitous sensors/decontaminators. Although this paper covers mechanisms and protocols to avail a neutralizing substance, further research will need to explore the substance's various pharmacological profiles and potential side effects.

Screening for antimicrobial and proteolytic activities of lactic acid bacteria isolated from cow, buffalo and goat milk and cheeses marketed in the southeast region of Brazil.

PubMed

Tulini, Fabricio L; Hymery, Nolwenn; Haertlé, Thomas; Le Blay, Gwenaelle; De Martinis, Elaine C P

2016-02-01

Lactic acid bacteria (LAB) can be isolated from different sources such as milk and cheese, and the lipolytic, proteolytic and glycolytic enzymes of LAB are important in cheese preservation and in flavour production. Moreover, LAB produce several antimicrobial compounds which make these bacteria interesting for food biopreservation. These characteristics stimulate the search of new strains with technological potential. From 156 milk and cheese samples from cow, buffalo and goat, 815 isolates were obtained on selective agars for LAB. Pure cultures were evaluated for antimicrobial activities by agar antagonism tests and for proteolytic activity on milk proteins by cultivation on agar plates. The most proteolytic isolates were also tested by cultivation in skim milk followed by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) analysis of the fermented milk. Among the 815 tested isolates, three of them identified as Streptococcus uberis (strains FT86, FT126 and FT190) were bacteriocin producers, whereas four other ones identified as Weissella confusa FT424, W. hellenica FT476, Leuconostoc citreum FT671 and Lactobacillus plantarum FT723 showed high antifungal activity in preliminary assays. Complementary analyses showed that the most antifungal strain was L. plantarum FT723 that inhibited Penicillium expansum in modified MRS agar (De Man, Rogosa, Sharpe, without acetate) and fermented milk model, however no inhibition was observed against Yarrowia lipolytica. The proteolytic capacities of three highly proteolytic isolates identified as Enterococcus faecalis (strains FT132 and FT522) and Lactobacillus paracasei FT700 were confirmed by SDS-PAGE, as visualized by the digestion of caseins and whey proteins (β-lactoglobulin and α-lactalbumin). These results suggest potential applications of these isolates or their activities (proteolytic activity or production of antimicrobials) in dairy foods production.

A novel junctional adhesion molecule A (CgJAM-A-L) from oyster (Crassostrea gigas) functions as pattern recognition receptor and opsonin.

PubMed

Liu, Conghui; Wang, Mengqiang; Jiang, Shuai; Wang, Lingling; Chen, Hao; Liu, Zhaoqun; Qiu, Limei; Song, Linsheng

2016-02-01

Junctional adhesion molecule (JAM), a subfamily of immunoglobulin superfamily (IgSF) with a couple of immunoglobulin domains, can act as regulator in homeostasis and inflammation of vertebrates. In the present study, a structural homolog of JAM-A (designated CgJAM-A-L) was screened out from oyster, Crassostrea gigas, through a search of JAM-A D1 domain (N-terminal Ig domain in JAM-A). The cDNA of CgJAM-A-L was of 1188 bp encoding a predicted polypeptide of 395 amino acids. The immunoreactive area of CgJAM-A-L mainly distributed over the plasma membrane of hemocytes. After Vibro splendidus or tumor necrosis factor (CgTNF-1) stimulation, the mRNA transcripts of CgJAM-A-L in hemocytes increased significantly by 4.46-fold and 9.00-fold (p < 0.01) of those in control group, respectively. The recombinant CgJAM-A-L protein (rCgJAM-A-L) could bind multiple PAMPs including lipopolysaccharides (LPS), peptidoglycan (PGN), lipoteichoic acid (LTA), mannose (MAN), β-glucan (GLU) and poly(I:C), and various microorganisms including Micrococcus luteus, Staphylococcus aureus, Escherichia coli, Vibro anguillarum, V. splendidus, Pastoris pastoris and Yarrowia lipolytica. The phagocytic rates of oyster hemocytes towards Gram-negative bacteria V. anguillarum and yeast P. pastoris were significantly enhanced after the incubation of rCgJAM-A-L, and even increased more significantly after the pre-incubation of rCgJAM-A-L with microbes (p < 0.01). The results collectively indicated that CgJAM-A-L functioned as an important pattern recognition receptor (PRR) and opsonin in the immune defense against invading pathogen in oyster. Moreover, as the most primitive specie with homolog of JAMs, the information of CgJAM-A-L in oyster would provide useful clues for the evolutionary study of JAMs and immunoglobulins. Copyright © 2015 Elsevier Ltd. All rights reserved.

Diversity of spoilage fungi associated with various French dairy products.

PubMed

Garnier, Lucille; Valence, Florence; Pawtowski, Audrey; Auhustsinava-Galerne, Lizaveta; Frotté, Nicolas; Baroncelli, Riccardo; Deniel, Franck; Coton, Emmanuel; Mounier, Jérôme

2017-01-16

Yeasts and molds are responsible for dairy product spoilage, resulting in significant food waste and economic losses. Yet, few studies have investigated the diversity of spoilage fungi encountered in dairy products. In the present study, 175 isolates corresponding to 105 from various spoiled dairy products and 70 originating from dairy production environments, were identified using sequencing of the ITS region, the partial β-tubulin, calmodulin and/or EFα genes, and the D1-D2 domain of the 26S rRNA gene for filamentous fungi and yeasts, respectively. Among the 41 species found in spoiled products, Penicillium commune and Penicillium bialowiezense were the most common filamentous fungi, representing around 10% each of total isolates while Meyerozyma guilliermondii and Trichosporon asahii were the most common yeasts (4.8% each of total isolates). Several species (e.g. Penicillium antarcticum, Penicillium salamii and Cladosporium phyllophilum) were identified for the first time in dairy products or their environment. In addition, numerous species were identified in both spoiled products and their corresponding dairy production environment suggesting that the latter acts as a primary source of contamination. Secondly, the resistance to chemical preservatives (sodium benzoate, calcium propionate, potassium sorbate and natamycin) of 10 fungal isolates representative of the observed biodiversity was also evaluated. Independently of the fungal species, natamycin had the lowest minimum inhibitory concentration (expressed in gram of preservative/l), followed by potassium sorbate, sodium benzoate and calcium propionate. In the tested conditions, Cladosporium halotolerans and Didymella pinodella were the most sensitive fungi while Yarrowia lipolytica and Candida parapsilosis were the most resistant towards the tested preservatives. This study provides interesting information on the occurrence of fungal contaminants in dairy products and environments that may help developing

Application of experimental design and derivative spectrophotometry methods in optimization and analysis of biosorption of binary mixtures of basic dyes from aqueous solutions.

PubMed

Asfaram, Arash; Ghaedi, Mehrorang; Ghezelbash, Gholam Reza; Pepe, Francesco

2017-05-01

Simultaneous biosorption of malachite green (MG) and crystal violet (CV) on biosorbent Yarrowia lipolytica ISF7 was studied. An appropriate derivative spectrophotometry technique was used to evaluate the concentration of each dye in binary solutions, despite significant interferences in visible light absorbances. The effects of pH, temperature, growth time, initial MG and CV concentration in batch experiments were assessed using Design of Experiment (DOE) according to central composite second order response surface methodology (RSM). The analysis showed that the greatest biosorption efficiency (>99% for both dyes) can be obtained at pH 7.0, T=28°C, 24h mixing and 20mgL -1 initial concentrations for both MG and CV dyes. The quadratic constructed equation ability for fitting experimental data is judged based on criterions like R 2 values, significant p and lack-of-fit value strongly confirm its high adequacy and applicability for prediction of revel behavior of the system under study. The proposed model showed very high correlation coefficients (R 2 =0.9997 for CV and R 2 =0.9989 for MG), while supported by closeness of predicted and experimental value. A kinetic analysis was carried out, showing that for both dyes a pseudo-second order kinetic model adequately describes the available data. The Langmuir isotherm model in single and binary components has better performance for description of dyes biosorption with maximum monolayer biosorption capacity of 59.4 and 62.7mgg -1 in single component and 46.4 and 50.0mgg -1 for CV and MB in binary components, respectively. The surface structure of biosorbents and the possible biosorbents-dyes interactions between were also evaluated by Fourier transform infrared (FT-IR) spectroscopy and scanning electron microscopy (SEM). The values of thermodynamic parameters including ΔG° and ΔH° strongly confirm which method is spontaneous and endothermic. Copyright © 2017. Published by Elsevier Inc.

Process Analysis of Variables for Standardization of Antifungal Susceptibility Testing of Nonfermentative Yeasts ▿

PubMed Central

Zaragoza, Oscar; Mesa-Arango, Ana C.; Gómez-López, Alicia; Bernal-Martínez, Leticia; Rodríguez-Tudela, Juan Luis; Cuenca-Estrella, Manuel

2011-01-01

Nonfermentative yeasts, such as Cryptococcus spp., have emerged as fungal pathogens during the last few years. However, standard methods to measure their antifungal susceptibility (antifungal susceptibility testing [AST]) are not completely reliable due to the impaired growth of these yeasts in standard media. In this work, we have compared the growth kinetics and the antifungal susceptibilities of representative species of nonfermentative yeasts such as Cryptococcus neoformans, Cryptococcus gattii, Cryptococcus albidus, Rhodotorula spp., Yarrowia lipolytica, Geotrichum spp., and Trichosporon spp. The effect of the growth medium (RPMI medium versus yeast nitrogen base [YNB]), glucose concentration (0.2% versus 2%), nitrogen source (ammonium sulfate), temperature (30°C versus 35°C), shaking, and inoculum size (103, 104, and 105 cells) were analyzed. The growth rate, lag phase, and maximum optical density were obtained from each growth experiment, and after multivariate analysis, YNB-based media demonstrated a significant improvement in the growth of yeasts. Shaking, an inoculum size of 105 CFU/ml, and incubation at 30°C also improved the growth kinetics of organisms. Supplementation with ammonium sulfate and with 2% glucose did not have any effect on growth. We also tested the antifungal susceptibilities of all the isolates by the reference methods of the CLSI and EUCAST, the EUCAST method with shaking, YNB under static conditions, and YNB with shaking. MIC values obtained under different conditions showed high percentages of agreement and significant correlation coefficient values between them. MIC value determinations according to CLSI and EUCAST standards were rather complicated, since more than half of isolates tested showed a limited growth index, hampering endpoint determinations. We conclude that AST conditions including YNB as an assay medium, agitation of the plates, reading after 48 h of incubation, an inoculum size of 105 CFU/ml, and incubation at 30�

Synthesis, structural characterization and photoluminescence properties of a novel La(III) complex

NASA Astrophysics Data System (ADS)

Köse, Muhammet; Ceyhan, Gökhan; Atcı, Emine; McKee, Vickie; Tümer, Mehmet

2015-05-01

In this study, a novel La(III) complex [La(H2L)2(NO3)3(MeOH)] of a Schiff base ligand was synthesized and characterized by spectroscopic and analytical methods. Single crystals of the complex suitable for X-ray diffraction study were obtained by slow diffusion of diethyl ether into a MeOH solution of the complex which was found to crystallise as [La(H2L)2(NO3)3(MeOH)]ṡ2MeOHṡH2O. The structure was solved in monoclinic crystal system, P21/n space group with unit cell parameters a = 10.5641(11), b = 12.6661(16), c = 16.0022(17) Å, α = 67.364(2), β = 83.794(2)°, γ = 70.541(2)°, V = 1862.9(4) Å3 and Z = 2 with R final value of 0.526. In the complex, the La(III) ion is ten-coordinated by O atoms, five of which come from three nitrate ions, four from the two Schiff base ligands and one from MeOH oxygen atom. The Schiff base ligands in the structure are in a zwitter ion form with the phenolic H transferred to the imine N atom. Thermal properties of the La(III) complex were examined by thermogravimetric analysis and the complex was found to be thermally stable up to 310 °C. The Schiff base ligand and its La(II) complex were screened for their in vitro antimicrobial activity against Bacillus megaterium, Staphylococcus aureus, Bacillus subtilis, Micrococcus luteus (Gram positive bacteria), Klebsiella pneumonia, Escherichia coli, Enterobacter aerogenes, Pseudomonas aeruginosa (Gram negative bacteria), Candida albicans,Yarrowia lipolytica (fungus) and Saccharomyces cerevisiae (yeast). The complex shows more antimicrobial activity than the free ligand.

Raman spectroscopy of oral tissues: correlation of spectral and biochemical markers

NASA Astrophysics Data System (ADS)

Singh, S. P.; Krishna, C. Murali

2014-03-01

Introduction Optical spectroscopic methods are being explored as novel tools for early and non-invasive cancer diagnosis. Both ex vivo and in vivo Raman spectroscopic studies carried out in oral cancer over the past decade have demonstrated that spectra of normal tissues are rich in lipids while tumor spectra show predominance of proteins. An accurate understanding of spectral features with respect to the biochemical composition is a pre-requisite before transferring these technologies for routine clinical usage. Therefore, in the present study, we have carried out Raman and biochemical studies on same tissues to correlate spectral markers and biochemical composition of normal and tumor oral tissues. Materials and Methods Spectra of 20 pairs of normal and tumor oral tissues were acquired using fiber-optic probe coupled HE-785 Raman spectrometer. Intensity associated with lipid (1440 cm-1) and protein (1450 and 1660 cm-1) bands were computed using curve-deconvolution method. Same tissues were then subjected to biochemical estimations of major biomolecules i.e., protein, lipid and phospholipids. Results and Discussion The intensity of the lipid band was found to be higher in normal tissues with respect to tumors, and the protein band was higher in tumors compared to normal tissues. Biochemical estimation yielded similar results i.e. high protein to lipid or phospholipid ratio in tumors with-respect to normal tissues. These differences were found to be statistically significant. Conclusion Findings of curve-deconvolution and biochemical estimation correlate very well and corroborate the spectral profile noted in earlier studies.

40 CFR 158.2081 - Experimental use permit biochemical pesticides product chemistry data requirements table.

Code of Federal Regulations, 2014 CFR

2014-07-01

... 40 Protection of Environment 24 2014-07-01 2014-07-01 false Experimental use permit biochemical... Pesticides § 158.2081 Experimental use permit biochemical pesticides product chemistry data requirements...: (d) Table. The following table shows the data requirements for experimental use permit biochemical...

40 CFR 158.2081 - Experimental use permit biochemical pesticides product chemistry data requirements table.

Code of Federal Regulations, 2013 CFR

2013-07-01

... 40 Protection of Environment 25 2013-07-01 2013-07-01 false Experimental use permit biochemical... Pesticides § 158.2081 Experimental use permit biochemical pesticides product chemistry data requirements...: (d) Table. The following table shows the data requirements for experimental use permit biochemical...

40 CFR 158.2081 - Experimental use permit biochemical pesticides product chemistry data requirements table.

Code of Federal Regulations, 2012 CFR

2012-07-01

... 40 Protection of Environment 25 2012-07-01 2012-07-01 false Experimental use permit biochemical... Pesticides § 158.2081 Experimental use permit biochemical pesticides product chemistry data requirements...: (d) Table. The following table shows the data requirements for experimental use permit biochemical...

40 CFR 158.2081 - Experimental use permit biochemical pesticides product chemistry data requirements table.

Code of Federal Regulations, 2010 CFR

2010-07-01

... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Experimental use permit biochemical... Pesticides § 158.2081 Experimental use permit biochemical pesticides product chemistry data requirements...: (d) Table. The following table shows the data requirements for experimental use permit biochemical...

40 CFR 158.2081 - Experimental use permit biochemical pesticides product chemistry data requirements table.

Code of Federal Regulations, 2011 CFR

2011-07-01

... 40 Protection of Environment 24 2011-07-01 2011-07-01 false Experimental use permit biochemical... Pesticides § 158.2081 Experimental use permit biochemical pesticides product chemistry data requirements...: (d) Table. The following table shows the data requirements for experimental use permit biochemical...

2009 Biochemical Conversion Platform Review Report

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ferrell, John

2009-12-01

This document summarizes the recommendations and evaluations provided by an independent external panel of experts at the U.S. Department of Energy Biomass Program’s Biochemical Conversion platform review meeting, held on April 14-16, 2009, at the Sheraton Denver Downtown, Denver, Colorado.

Length of positive surgical margin after radical prostatectomy as a predictor of biochemical recurrence.

PubMed

Shikanov, Sergey; Song, Jie; Royce, Cassandra; Al-Ahmadie, Hikmat; Zorn, Kevin; Steinberg, Gary; Zagaja, Gregory; Shalhav, Arieh; Eggener, Scott

2009-07-01

Length and location of positive surgical margins are independent predictors of biochemical recurrence after open radical prostatectomy. We assessed their impact on biochemical recurrence in a large robotic prostatectomy series. Data were collected prospectively from 1,398 men undergoing robotic radical prostatectomy for clinically localized prostate cancer from 2003 to 2008 at a single institution. The associations of preoperative prostate specific antigen, pathological Gleason score, pathological stage and positive surgical margin parameters (location, length and focality) with biochemical recurrence rate were evaluated. Margin status and length were measured by a single uropathologist. Biochemical recurrence was defined as serum prostate specific antigen greater than 0.1 ng/ml on 2 consecutive tests. Cox regression models were constructed to evaluate predictors of biochemical recurrence. Of 1,398 consecutive patients who underwent robotic prostatectomy positive margins were present in 243 (17%) (11% of pathological T2 and 41% of T3). Preoperative prostate specific antigen, pathological stage, Gleason score, margin status, and margin length as a continuous and categorical variable (less than 1, 1 to 3, more than 3 mm) were independent predictors of biochemical recurrence. Patients with negative margins and those with a positive margin less than 1 mm had similar rates of biochemical recurrence (log rank test p = 0.18). Surgical margin location was not independently associated with biochemical recurrence. Margin status and length are independent predictors of biochemical recurrence following robotic radical prostatectomy. Although longer followup and validation studies are necessary for confirmation, patients with a positive margin less than 1 mm appear to have similar recurrence rates as those with negative margins.

Improved biochemical preservation of heart slices during cold storage.

PubMed

Bull, D A; Reid, B B; Connors, R C; Albanil, A; Stringham, J C; Karwande, S V

2000-01-01

Development of myocardial preservation solutions requires the use of whole organ models which are animal and labor intensive. These models rely on physiologic rather than biochemical endpoints, making accurate comparison of the relative efficacy of individual solution components difficult. We hypothesized that myocardial slices could be used to assess preservation of biochemical function during cold storage. Whole rat hearts were precision cut into slices with a thickness of 200 microm and preserved at 4 degrees C in one of the following solutions: Columbia University (CU), University of Wisconsin (UW), D5 0.2% normal saline with 20 meq/l KCL (QNS), normal saline (NS), or a novel cardiac preservation solution (NPS) developed using this model. Myocardial biochemical function was assessed by ATP content (etamoles ATP/mg wet weight) and capacity for protein synthesis (counts per minute (cpm)/mg protein) immediately following slicing (0 hours), and at 6, 12, 18, and 24 hours of cold storage. Six slices were assayed at each time point for each solution. The data were analyzed using analysis of variance and are presented as the mean +/- standard deviation. ATP content was higher in the heart slices stored in the NPS compared to all other solutions at 6, 12, 18 and 24 hours of cold storage (p < 0.05). Capacity for protein synthesis was higher in the heart slices stored in the NPS compared to all other solutions at 6, 12, and 18 hours of cold storage (p < 0.05). CONCLUSIONS This myocardial slice model allows the rapid and efficient screening of cardiac preservation solutions and their components using quantifiable biochemical endpoints. Using this model, we have developed a novel preservation solution which improves the biochemical function of myocardial slices during cold storage.

Rapid Methods for Biochemical Testing of Anaerobic Bacteria

PubMed Central

Schreckenberger, Paul C.; Blazevic, Donna J.

1974-01-01

Rapid biochemical tests for nitrate, indole, gelatin, starch, esculin, and o-nitrophenyl-β-D-galactopyranoside were performed on 112 strains of anaerobic bacteria. All tests were incubated under aerobic conditions, and results were recorded within 4 h. The tests for nitrate, indole, and starch showed a 95% or greater correlation when compared to the standard biochemical tests. Tests for esculin and gelatin showed an agreement of 86 and 77%, respectively. PathoTec test strips for nitrate, indole, esculin, o-nitrophenyl-β-D-galactopyranoside, Voges-Proskauer, and urease were also tested and showed encouraging results. PMID:4613268

Changes in Biochemical Properties of the Blood in Winter Swimmers.

PubMed

Teleglow, Aneta; Marchewka, Jakub; Marchewka, Anna; Kulpa, Jan

The aim of the study was to investigate the effects of winter swimming on biochemical indicators of the blood. The subjects - winter swimmers - belonged to the Krakow Walrus Club "Kaloryfer" - "The Heater". The study group consisted of 11 men, aged 30-50 years, 'walrusing' throughout the whole season from November to March. Statistically significant changes throughout the 'walrusing' season were observed for the following biochemical parameters: a decrease in sodium (mmol/1), chloride (mmol/1), alpha-2 globulin(g/1), gamma globulin (g/1), IgG (g/1), and an increase in albumin (g/1), indicator A/G, IgA (g/l ), Herpes simplex virus IgM. Seasonal effort of winter swimmers has a positive influence on biochemical blood parameters.

A Program on Biochemical and Biomedical Engineering.

ERIC Educational Resources Information Center

San, Ka-Yiu; McIntire, Larry V.

1989-01-01

Presents an introduction to the Biochemical and Biomedical Engineering program at Rice University. Describes the development of the academic and enhancement programs, including organizational structure and research project titles. (YP)

CLINICAL FACTORS ASSOCIATED WITH BIOCHEMICAL ADRENAL-CORTISOL INSUFFICIENCY IN HOSPITALIZED PATIENTS

PubMed Central

Ben-Shlomo, Anat; Mirocha, James; Liu, Ning-Ai; Sheinin, Renee C.; Melmed, Shlomo

2014-01-01

Background Diagnosis of adrenal-cortisol insufficiency is often misleading in hospitalized patients as clinical and biochemical features overlap with co-morbidities. We analyzed clinical determinants associated with a biochemical diagnosis of adrenal-cortisol insufficiency in non-ICU hospitalized patients. Methods In a retrospective cohort study we reviewed 4668 inpatients with random morning cortisol levels ≤15 μg/dL hospitalized in our center between 2003 and 2010. Using serum cortisol threshold level of 18 μg/dL 30 and/or 60 minutes after cortrosyn (250 μg) injection to define biochemical adrenal-cortisol status, we characterized and compared insufficient (n=108, serum cortisol ≤18 μg/dL) and sufficient ( n=394; serum cortisol >18 μg/dL) non-ICU hospitalized patients. Results Commonly reported clinical and routine biochemical adrenal-cortisol insufficiency features were similar between insufficient and sufficient inpatients. Biochemical adrenal-cortisol insufficiency was associated with increased frequency of liver disease, specifically hepatitis C (p=0.01) and prior orthotopic liver transplantation (p<0.001), HIV (p=0.005) and reported preexisting male hypogonadism (p<0.001) as compared to biochemical adrenal-cortisol sufficiency group. Forty percent of insufficient inpatients were not treated with glucocorticoids after diagnosis. Multivariable logistic analysis demonstrated that inpatients with higher cortisol levels (p=0.0001), higher diastolic blood pressure (p=0.05) and females (p=0.009) were more likely not to be treated, while those with previous short-term glucocorticoid treatment (p=0.002), had other co-existing endocrine diseases (p=0.005) or received an inhospital endocrinology consultation (p<0.0001) were more likely to be replaced with glucocorticoids. Conclusions Commonly reported adrenal-cortisol insufficiency features do not reliably identify hospitalized patients biochemically confirmed to have this disorder. Co-morbidities including

Multiple Ion Binding Equilibria, Reaction Kinetics, and Thermodynamics in Dynamic Models of Biochemical Pathways

PubMed Central

Vinnakota, Kalyan C.; Wu, Fan; Kushmerick, Martin J.; Beard, Daniel A.

2009-01-01

The operation of biochemical systems in vivo and in vitro is strongly influenced by complex interactions between biochemical reactants and ions such as H+, Mg2+, K+, and Ca2+. These are important second messengers in metabolic and signaling pathways that directly influence the kinetics and thermodynamics of biochemical systems. Herein we describe the biophysical theory and computational methods to account for multiple ion binding to biochemical reactants and demonstrate the crucial effects of ion binding on biochemical reaction kinetics and thermodynamics. In simulations of realistic systems, the concentrations of these ions change with time due to dynamic buffering and competitive binding. In turn, the effective thermodynamic properties vary as functions of cation concentrations and important environmental variables such as temperature and overall ionic strength. Physically realistic simulations of biochemical systems require incorporating all of these phenomena into a coherent mathematical description. Several applications to physiological systems are demonstrated based on this coherent simulation framework. PMID:19216922

An Integrated Qualitative and Quantitative Biochemical Model Learning Framework Using Evolutionary Strategy and Simulated Annealing.

PubMed

Wu, Zujian; Pang, Wei; Coghill, George M

2015-01-01

Both qualitative and quantitative model learning frameworks for biochemical systems have been studied in computational systems biology. In this research, after introducing two forms of pre-defined component patterns to represent biochemical models, we propose an integrative qualitative and quantitative modelling framework for inferring biochemical systems. In the proposed framework, interactions between reactants in the candidate models for a target biochemical system are evolved and eventually identified by the application of a qualitative model learning approach with an evolution strategy. Kinetic rates of the models generated from qualitative model learning are then further optimised by employing a quantitative approach with simulated annealing. Experimental results indicate that our proposed integrative framework is feasible to learn the relationships between biochemical reactants qualitatively and to make the model replicate the behaviours of the target system by optimising the kinetic rates quantitatively. Moreover, potential reactants of a target biochemical system can be discovered by hypothesising complex reactants in the synthetic models. Based on the biochemical models learned from the proposed framework, biologists can further perform experimental study in wet laboratory. In this way, natural biochemical systems can be better understood.

Chemical and biochemical thermodynamics: Is it time for a reunification?

PubMed

Iotti, Stefano; Raff, Lionel; Sabatini, Antonio

2017-02-01

The thermodynamics of chemical reactions in which all species are explicitly considered with atoms and charge balanced is compared with the transformed thermodynamics generally used to treat biochemical reactions where atoms and charges are not balanced. The transformed thermodynamic quantities suggested by Alberty are obtained by execution of Legendre transformation of the usual thermodynamic potentials. The present analysis demonstrates that the transformed values for Δ r G' 0 and Δ r H' 0 can be obtained directly without performing Legendre transformations by simply writing the chemical reactions with all the pseudoisomers explicitly included and charges balanced. The appropriate procedures for computing the stoichiometric coefficients for the pseudoisomers are fully explained by means of an example calculation for the biochemical ATP hydrolysis reaction. It is concluded that the analysis has reunited the "two separate worlds" of conventional thermodynamics and transformed thermodynamics. In addition, it is also shown that the value of the conditional Gibbs energy of reaction, Δ r G', for a biochemical reaction is the same of the value of Δ r G for any chemical reaction involving pseudoisomers of the biochemical reagents. Copyright © 2016 Elsevier B.V. All rights reserved.

Explorations into Chemical Reactions and Biochemical Pathways.

PubMed

Gasteiger, Johann

2016-12-01

A brief overview of the work in the research group of the present author on extracting knowledge from chemical reaction data is presented. Methods have been developed to calculate physicochemical effects at the reaction site. It is shown that these physicochemical effects can quite favourably be used to derive equations for the calculation of data on gas phase reactions and on reactions in solution such as aqueous acidity of alcohols or carboxylic acids or the hydrolysis of amides. Furthermore, it is shown that these physicochemical effects are quite effective for assigning reactions into reaction classes that correspond to chemical knowledge. Biochemical reactions constitute a particularly interesting and challenging task for increasing our understanding of living species. The BioPath.Database is a rich source of information on biochemical reactions and has been used for a variety of applications of chemical, biological, or medicinal interests. Thus, it was shown that biochemical reactions can be assigned by the physicochemical effects into classes that correspond to the classification of enzymes by the EC numbers. Furthermore, 3D models of reaction intermediates can be used for searching for novel enzyme inhibitors. It was shown in a combined application of chemoinformatics and bioinformatics that essential pathways of diseases can be uncovered. Furthermore, a study showed that bacterial flavor-forming pathways can be discovered. © 2016 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Biochemical Thermodynamics under near Physiological Conditions

ERIC Educational Resources Information Center

Mendez, Eduardo

2008-01-01

The recommendations for nomenclature and tables in Biochemical Thermodynamics approved by IUBMB and IUPAC in 1994 can be easily introduced after the chemical thermodynamic formalism. Substitution of the usual standard thermodynamic properties by the transformed ones in the thermodynamic equations, and the use of appropriate thermodynamic tables…

Biochemical Approaches to Improved Nitrogen Fixation

USDA-ARS?s Scientific Manuscript database

Improving symbiotic nitrogen fixation by legumes has emerged again as an important topic on the world scene due to the energy crisis and lack of access to nitrogen fertilizer in developing countries. We have taken a biochemical genomics approach to improving symbiotic nitrogen fixation in legumes. L...

Biochemical responses of the Skylab crewman

NASA Technical Reports Server (NTRS)

Leach, C. S.; Rambaut, P. C.

1974-01-01

The biochemical investigations of the Skylab crewmen were designed to study the physiological changes that were observed on flight crews returning from previous space flight missions as well as to study those changes expected to result from prolonged weightless exposure. These studies can be divided into two broad categories. One category included routine blood studies similar to those used in clinical medical practice. The second included research-type endocrine analyses used to investigate more thoroughly the metabolic/endocrine responses to the space flight environment. The premission control values indicated that all Skylab crewmen were healthy and were free from biochemical abnormalities. The routine results during and after flight showed slight but significant changes in electrolytes, glucose, total protein, osmolality, uric acid, cholesterol, and creatinine. Plasma hormal changes included adrenocorticotrophic hormone, cortisol, angiotensin I, aldosterone, insulin, and thyroxine. The 24-hour urine analyses results revealed increased excretion of cortisol, catecholamines, antidiuretic hormone, and aldosterone as well as excretion of significant electrolyte and uric acid during the Skylab flights.

Complete integrability of information processing by biochemical reactions

PubMed Central

Agliari, Elena; Barra, Adriano; Dello Schiavo, Lorenzo; Moro, Antonio

2016-01-01

Statistical mechanics provides an effective framework to investigate information processing in biochemical reactions. Within such framework far-reaching analogies are established among (anti-) cooperative collective behaviors in chemical kinetics, (anti-)ferromagnetic spin models in statistical mechanics and operational amplifiers/flip-flops in cybernetics. The underlying modeling – based on spin systems – has been proved to be accurate for a wide class of systems matching classical (e.g. Michaelis–Menten, Hill, Adair) scenarios in the infinite-size approximation. However, the current research in biochemical information processing has been focusing on systems involving a relatively small number of units, where this approximation is no longer valid. Here we show that the whole statistical mechanical description of reaction kinetics can be re-formulated via a mechanical analogy – based on completely integrable hydrodynamic-type systems of PDEs – which provides explicit finite-size solutions, matching recently investigated phenomena (e.g. noise-induced cooperativity, stochastic bi-stability, quorum sensing). The resulting picture, successfully tested against a broad spectrum of data, constitutes a neat rationale for a numerically effective and theoretically consistent description of collective behaviors in biochemical reactions. PMID:27812018

Reconstructing biochemical pathways from time course data.

PubMed

Srividhya, Jeyaraman; Crampin, Edmund J; McSharry, Patrick E; Schnell, Santiago

2007-03-01

Time series data on biochemical reactions reveal transient behavior, away from chemical equilibrium, and contain information on the dynamic interactions among reacting components. However, this information can be difficult to extract using conventional analysis techniques. We present a new method to infer biochemical pathway mechanisms from time course data using a global nonlinear modeling technique to identify the elementary reaction steps which constitute the pathway. The method involves the generation of a complete dictionary of polynomial basis functions based on the law of mass action. Using these basis functions, there are two approaches to model construction, namely the general to specific and the specific to general approach. We demonstrate that our new methodology reconstructs the chemical reaction steps and connectivity of the glycolytic pathway of Lactococcus lactis from time course experimental data.

The use of biochemical methods in extraterrestrial life detection

NASA Astrophysics Data System (ADS)

McDonald, Gene

2006-08-01

Instrument development for in situ extraterrestrial life detection focuses primarily on the ability to distinguish between biological and non-biological material, mostly through chemical analysis for potential biosignatures (e.g., biogenic minerals, enantiomeric excesses). In constrast, biochemical analysis techniques commonly applied to Earth life focus primarily on the exploration of cellular and molecular processes, not on the classification of a given system as biological or non-biological. This focus has developed because of the relatively large functional gap between life and non-life on Earth today. Life on Earth is very diverse from an environmental and physiological point of view, but is highly conserved from a molecular point of view. Biochemical analysis techniques take advantage of this similarity of all terrestrial life at the molecular level, particularly through the use of biologically-derived reagents (e.g., DNA polymerases, antibodies), to enable analytical methods with enormous sensitivity and selectivity. These capabilities encourage consideration of such reagents and methods for use in extraterrestrial life detection instruments. The utility of this approach depends in large part on the (unknown at this time) degree of molecular compositional differences between extraterrestrial and terrestrial life. The greater these differences, the less useful laboratory biochemical techniques will be without significant modification. Biochemistry and molecular biology methods may need to be "de-focused" in order to produce instruments capable of unambiguously detecting a sufficiently wide range of extraterrestrial biochemical systems. Modern biotechnology tools may make that possible in some cases.

Biochemical tests of placental function for assessment in pregnancy.

PubMed

Neilson, James P

2012-08-15

Biochemical tests of placental or feto-placental function were widely used in the 1960s and 1970s in high-risk pregnancies to try to predict, and thus try to avoid, adverse fetal outcome. To assess the effects of performing biochemical tests of placental function in high-risk, low-risk, or unselected pregnancies. We searched the Cochrane Pregnancy and Childbirth Group's Trials Register (10 May 2012). Controlled trials (randomized or 'quasi-randomized') that compare the use of biochemical tests of placental function in pregnancy with non-use. Trial quality was assessed and data were extracted by the review author. A single eligible trial of poor quality was identified. It involved 622 women with high-risk pregnancies who had had plasma (o)estriol estimations. Women were allocated to have their (o)estriol results revealed or concealed on the basis of hospital record number (with attendant risk of selection bias). There were no obvious differences in perinatal mortality (relative risk (RR) 0.88, 95% confidence interval (CI) 0.36 to 2.13) or planned delivery (RR 0.97, 95% CI 0.81 to 1.15) between the two groups. The available trial data do not support the use of (o)estriol estimation in high-risk pregnancies. The single small trial available does not have the power to exclude a beneficial effect but this is probably of historical interest since biochemical testing has been superseded by biophysical testing in antepartum fetal assessment.

40 CFR 158.2083 - Experimental use permit biochemical pesticides human health assessment data requirements table.

Code of Federal Regulations, 2013 CFR

2013-07-01

... pesticides human health assessment data requirements table. 158.2083 Section 158.2083 Protection of... Biochemical Pesticides § 158.2083 Experimental use permit biochemical pesticides human health assessment data... determine the human health assessment data requirements for a particular biochemical pesticide product. (2...

40 CFR 158.2083 - Experimental use permit biochemical pesticides human health assessment data requirements table.

Code of Federal Regulations, 2014 CFR

2014-07-01

... pesticides human health assessment data requirements table. 158.2083 Section 158.2083 Protection of... Biochemical Pesticides § 158.2083 Experimental use permit biochemical pesticides human health assessment data... determine the human health assessment data requirements for a particular biochemical pesticide product. (2...

40 CFR 158.2083 - Experimental use permit biochemical pesticides human health assessment data requirements table.

Code of Federal Regulations, 2012 CFR

2012-07-01

... pesticides human health assessment data requirements table. 158.2083 Section 158.2083 Protection of... Biochemical Pesticides § 158.2083 Experimental use permit biochemical pesticides human health assessment data... determine the human health assessment data requirements for a particular biochemical pesticide product. (2...

40 CFR 158.2083 - Experimental use permit biochemical pesticides human health assessment data requirements table.

Code of Federal Regulations, 2010 CFR

2010-07-01

... pesticides human health assessment data requirements table. 158.2083 Section 158.2083 Protection of... Biochemical Pesticides § 158.2083 Experimental use permit biochemical pesticides human health assessment data... determine the human health assessment data requirements for a particular biochemical pesticide product. (2...

40 CFR 158.2083 - Experimental use permit biochemical pesticides human health assessment data requirements table.

Code of Federal Regulations, 2011 CFR

2011-07-01

... pesticides human health assessment data requirements table. 158.2083 Section 158.2083 Protection of... Biochemical Pesticides § 158.2083 Experimental use permit biochemical pesticides human health assessment data... determine the human health assessment data requirements for a particular biochemical pesticide product. (2...

Biochemical reactions of ozone in plants

Treesearch

J. Brian Mudd

1998-01-01

Plants react biochemically to ozone in three phases: with constitutive chemicals in the apoplastic fluid and cell membranes; by forming messenger molecules by the affected constitutive materials (ethylene); and by responding to the messenger molecules with pathogenic RNAs and proteins. For instance, plant reactions with ozone result in constitutive molecules such as...

Overview of the DOE/SERI Biochemical Conversion Program

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wright, J D

1986-09-01

The Solar Energy Research Institute manages a program of research and development on the biochemical conversion of renewable lignocellulosic materials to liquid fuels for the Department of Energy's Biofuels and Municipal Waste Technology Division. The Biochemical Conversion Program is mission oriented so effort is concentrated on technologies which appear to have the greatest potential for being adopted by the private sector to economically convert lignocellulosic materials into high value liquid transportation fuels such as ethanol. The program is structured to supply the technology for such fuels to compete economically first as an octane booster or fuel additive, and, with additionalmore » improvements, as a neat fuel. 18 refs., 3 figs., 1 tab.« less

Biochemical Reconstitution of the WAVE Regulatory Complex

PubMed Central

Chen, Baoyu; Padrick, Shae B.; Henry, Lisa; Rosen, Michael K.

2014-01-01

The WAVE regulatory complex (WRC) is a 400-KDa heteropentameric protein assembly that plays a central role in controlling actin cytoskeletal dynamics in many cellular processes. The WRC acts by integrating diverse cellular cues and stimulating the actin nucleating activity of the Arp2/3 complex at membranes. Biochemical and biophysical studies of the underlying mechanisms of these processes require large amounts of purified WRC. Recent success in recombinant expression, reconstitution, purification and crystallization of the WRC has greatly advanced our understanding of the inhibition, activation and membrane recruitment mechanisms of this complex. But many important questions remain to be answered. Here we summarize and update the methods developed in our laboratory, which allow reliable and flexible production of tens of milligrams of recombinant WRC of crystallographic quality, sufficient for many biochemical and structural studies. PMID:24630101

A systematic petri net approach for multiple-scale modeling and simulation of biochemical processes.

PubMed

Chen, Ming; Hu, Minjie; Hofestädt, Ralf

2011-06-01

A method to exploit hybrid Petri nets for modeling and simulating biochemical processes in a systematic way was introduced. Both molecular biology and biochemical engineering aspects are manipulated. With discrete and continuous elements, the hybrid Petri nets can easily handle biochemical factors such as metabolites concentration and kinetic behaviors. It is possible to translate both molecular biological behavior and biochemical processes workflow into hybrid Petri nets in a natural manner. As an example, penicillin production bioprocess is modeled to illustrate the concepts of the methodology. Results of the dynamic of production parameters in the bioprocess were simulated and observed diagrammatically. Current problems and post-genomic perspectives were also discussed.

Aspects on the Physiological and Biochemical Foundations of Neurocritical Care

PubMed Central

Nordström, Carl-Henrik; Koskinen, Lars-Owe; Olivecrona, Magnus

2017-01-01

Neurocritical care (NCC) is a branch of intensive care medicine characterized by specific physiological and biochemical monitoring techniques necessary for identifying cerebral adverse events and for evaluating specific therapies. Information is primarily obtained from physiological variables related to intracranial pressure (ICP) and cerebral blood flow (CBF) and from physiological and biochemical variables related to cerebral energy metabolism. Non-surgical therapies developed for treating increased ICP are based on knowledge regarding transport of water across the intact and injured blood–brain barrier (BBB) and the regulation of CBF. Brain volume is strictly controlled as the BBB permeability to crystalloids is very low restricting net transport of water across the capillary wall. Cerebral pressure autoregulation prevents changes in intracranial blood volume and intracapillary hydrostatic pressure at variations in arterial blood pressure. Information regarding cerebral oxidative metabolism is obtained from measurements of brain tissue oxygen tension (PbtO2) and biochemical data obtained from intracerebral microdialysis. As interstitial lactate/pyruvate (LP) ratio instantaneously reflects shifts in intracellular cytoplasmatic redox state, it is an important indicator of compromised cerebral oxidative metabolism. The combined information obtained from PbtO2, LP ratio, and the pattern of biochemical variables reveals whether impaired oxidative metabolism is due to insufficient perfusion (ischemia) or mitochondrial dysfunction. Intracerebral microdialysis and PbtO2 give information from a very small volume of tissue. Accordingly, clinical interpretation of the data must be based on information of the probe location in relation to focal brain damage. Attempts to evaluate global cerebral energy state from microdialysis of intraventricular fluid and from the LP ratio of the draining venous blood have recently been presented. To be of clinical relevance, the

Biochemical basis for the biological clock

NASA Technical Reports Server (NTRS)

Morre, D. James; Chueh, Pin-Ju; Pletcher, Jake; Tang, Xiaoyu; Wu, Lian-Ying; Morre, Dorothy M.

2002-01-01

NADH oxidases at the external surface of plant and animal cells (ECTO-NOX proteins) exhibit stable and recurring patterns of oscillations with potentially clock-related, entrainable, and temperature-compensated period lengths of 24 min. To determine if ECTO-NOX proteins might represent the ultradian time keepers (pacemakers) of the biological clock, COS cells were transfected with cDNAs encoding tNOX proteins having a period length of 22 min or with C575A or C558A cysteine to alanine replacements having period lengths of 36 or 42 min. Here we demonstrate that such transfectants exhibited 22, 36, or 40 to 42 h circadian patterns in the activity of glyceraldehyde-3-phosphate dehydrogenase, a common clock-regulated protein, in addition to the endogenous 24 h circadian period length. The fact that the expression of a single oscillatory ECTO-NOX protein determines the period length of a circadian biochemical marker (60 X the ECTO-NOX period length) provides compelling evidence that ECTO-NOX proteins are the biochemical ultradian drivers of the cellular biological clock.

A Randomized Phase 2 Trial of 177Lu Radiolabeled Anti-PSMA Biochemically Monoclonal Antibody J591 in Patients with High-Risk Castrate, Biochemically Relapsed Prostate Cancer

DTIC Science & Technology

2010-09-01

relapsed prostate cancer (PC) after local therapy. J Clin Oncol 28: 15s, 2010 (suppl; Abstr TPS248) Presentation: Poster presentation, 2010 ASCO...Annual Meeting V. Conclusions Biochemical relapse is common after local therapy for prostate cancer. Based on the physical properties of 177Lu...ketoconazole in patients (pts) with high-risk castrate biochemically relapsed prostate cancer (PC) after local therapy. S. T. Tagawa, J. Osborne, P. J

ReactPRED: a tool to predict and analyze biochemical reactions.

PubMed

Sivakumar, Tadi Venkata; Giri, Varun; Park, Jin Hwan; Kim, Tae Yong; Bhaduri, Anirban

2016-11-15

Biochemical pathways engineering is often used to synthesize or degrade target chemicals. In silico screening of the biochemical transformation space allows predicting feasible reactions, constituting these pathways. Current enabling tools are customized to predict reactions based on pre-defined biochemical transformations or reaction rule sets. Reaction rule sets are usually curated manually and tailored to specific applications. They are not exhaustive. In addition, current systems are incapable of regulating and refining data with an aim to tune specificity and sensitivity. A robust and flexible tool that allows automated reaction rule set creation along with regulated pathway prediction and analyses is a need. ReactPRED aims to address the same. ReactPRED is an open source flexible and customizable tool enabling users to predict biochemical reactions and pathways. The tool allows automated reaction rule creation from a user defined reaction set. Additionally, reaction rule degree and rule tolerance features allow refinement of predicted data. It is available as a flexible graphical user interface and a console application. ReactPRED is available at: https://sourceforge.net/projects/reactpred/ CONTACT: anirban.b@samsung.com or ty76.kim@samsung.comSupplementary information: Supplementary data are available at Bioinformatics online. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Thermosyntropha lipolytica gen. nov., sp. nov., a lipolytic, anaerobic, alkalitolerant, thermophilic bacterium utilizing short- and long-chain fatty acids in syntrophic coculture with a methanogenic archaeum.

PubMed

Svetlitshnyi, V; Rainey, F; Wiegel, J

1996-10-01

Three strains of an anaerobic thermophilic organoheterotrophic lipolytic alkalitolerant bacterium, Thermosyntropha lipolytica gen. nov., sp. nov. (type strain JW/VS-265T; DSM 11003), were isolated from alkaline hot springs of Lake Bogoria (Kenya). The cells were nonmotile, non-spore forming, straight or slightly curved rods. At 60 degrees C the pH range for growth determined at 25 degrees C [pH25 degrees C] was 7.15 to 9.5, with an optimum between 8.1 and 8.9 (pH60 degrees C of 7.6 and 8.1). At a pH25 degrees C of 8.5 the temperature range for growth was from 52 to 70 degrees C, with an optimum between 60 and 66 degrees C. The shortest doubling time was around 1 h. In pure culture the bacterium grew in a mineral base medium supplemented with yeast extract, tryptone, Casamino Acids, betaine, and crotonate as carbon sources, producing acetate as a major product and constitutively a lipase. During growth in the presence of olive oil, free long-chain fatty acids were accumulated in the medium but the pure culture could not utilize olive oil, triacylglycerols, short- and long-chain fatty acids, and glycerol for growth. In syntrophic coculture (Methanobacterium strain JW/VS-M29) the lipolytic bacteria grew on triacylglycerols and linear saturated and unsaturated fatty acids with 4 to 18 carbon atoms, but glycerol was not utilized. Fatty acids with even numbers of carbon atoms were degraded to acetate and methane, while from odd-numbered fatty acids 1 mol of propionate per mol of fatty acid was additionally formed. 16S rDNA sequence analysis identified Syntrophospora and Syntrophomonas spp. as closest phylogenetic neighbors.

Variations in biochemical values for common laboratory tests: a comparison among multi-ethnic Israeli women cohort.

PubMed

Birk, Ruth; Heifetz, Eliyahu M

2018-04-28

Biochemical laboratory values are an essential tool in medical diagnosis, treatment, and follow-up; however, they are known to vary between populations. Establishment of ethnicity-adjusted reference values is recommended by health organizations. To investigate the ethnicity element in biochemical lab values studying women of different ethnic groups. Biochemical lab values (n = 27) of 503 adult Israeli women of three ethnicities (Jewish Ashkenazi, Jewish Sephardic, and Bedouin Arab) attending a single medical center were analyzed. Biochemical data were extracted from medical center records. Ethnic differences of laboratory biochemicals were studied using ANCOVA to analyze the center of the distribution as well as quartile regression analysis to analyze the upper and lower limits, both done with an adjustment for age. Significant ethnic differences were found in almost half (n = 12) of the biochemical laboratory tests. Ashkenazi Jews exhibited significantly higher mean values compared to Bedouins in most of the biochemical tests, including albumin, alkaline phosphatase, calcium, cholesterol, cholesterol LDL and HDL, cholesterol LDL calc., folic acid, globulin, and iron saturation, while the Bedouins exhibited the highest mean values in the creatinine and triglycerides. For most of these tests, Sephardic Jews exhibited biochemical mean levels in between the two other groups. Compared to Ashkenazi Jews, Sephardic Jews had a significant shift to lower values in cholesterol LDL. Ethnic subpopulations have distinct distributions in biochemical laboratory test values, which should be taken into consideration in medical practice enabling precision medicine.

[Circulating miR-152 helps early prediction of postoperative biochemical recurrence of prostate cancer].

PubMed

Chen, Jun-Feng; Liao, Yu-Feng; Ma, Jian-Bo; Mao, Qi-Feng; Jia, Guang-Cheng; Dong, Xue-Jun

2017-07-01

To investigate the value of circulating miR-152 in the early prediction of postoperative biochemical recurrence of prostate cancer. Sixty-six cases of prostate cancer were included in this study, 35 with and 31 without biochemical recurrence within two years postoperatively, and another 31 healthy individuals were enrolled as normal controls. The relative expression levels of circulating miR-152 in the serum of the subjects were detected by qRT-PCR, its value in the early diagnosis of postoperative biochemical recurrence of prostate cancer was assessed by ROC curve analysis, and the correlation of its expression level with the clinicopathological parameters of the patients were analyzed. The expression of circulating miR-152 was significantly lower in the serum of the prostate cancer patients than in the normal controls (t = －5.212, P = 0.001), and so was it in the patients with than in those without postoperative biochemical recurrence (t = －5.727, P = 0.001). The ROC curve for the value of miR-152 in the early prediction of postoperative biochemical recurrence of prostate cancer showed the area under the curve (AUC) to be 0.906 (95% CI: 0.809－0.964), with a sensitivity of 91.4% and a specificity of 80.6%. The expression level of miR-152 was correlated with the Gleason score, clinical stage of prostate cancer, biochemical recurrence, and bone metastasis (P <0.05), decreasing with increased Gleason scores and elevated clinical stage of the malignancy. No correlation, however, was found between the miR-152 expression and the patients' age or preoperative PSA level (P >0.05). The expression level of circulating miR-152 is significantly reduced in prostate cancer patients with biochemical recurrence after prostatectomy and could be a biomarker in the early prediction of postoperative biochemical recurrence of the malignancy.

Real-time biochemical sensor based on Raman scattering with CMOS contact imaging.

PubMed

Muyun Cao; Yuhua Li; Yadid-Pecht, Orly

2015-08-01

This work presents a biochemical sensor based on Raman scattering with Complementary metal-oxide-semiconductor (CMOS) contact imaging. This biochemical optical sensor is designed for detecting the concentration of solutions. The system is built with a laser diode, an optical filter, a sample holder and a commercial CMOS sensor. The output of the system is analyzed by an image processing program. The system provides instant measurements with a resolution of 0.2 to 0.4 Mol. This low cost and easy-operated small scale system is useful in chemical, biomedical and environmental labs for quantitative bio-chemical concentration detection with results reported comparable to a highly cost commercial spectrometer.

Thermodynamic considerations on Ca2+-induced biochemical reactions in living cells

NASA Astrophysics Data System (ADS)

Lucia, Umberto; Ponzetto, Antonio

2016-02-01

Cells can be regarded as complex engines that execute a series of chemical reactions. Energy transformations, thermo-electro-chemical processes and transport phenomena can occur across cell membranes. Different, related thermo-electro-biochemical behaviour can occur between health and disease states. Analysis of the irreversibility related to ion fluxes can represent a new approach to study and control the biochemical behaviour of living cells.

Thermodynamics of Computational Copying in Biochemical Systems

NASA Astrophysics Data System (ADS)

Ouldridge, Thomas E.; Govern, Christopher C.; ten Wolde, Pieter Rein

2017-04-01

Living cells use readout molecules to record the state of receptor proteins, similar to measurements or copies in typical computational devices. But is this analogy rigorous? Can cells be optimally efficient, and if not, why? We show that, as in computation, a canonical biochemical readout network generates correlations; extracting no work from these correlations sets a lower bound on dissipation. For general input, the biochemical network cannot reach this bound, even with arbitrarily slow reactions or weak thermodynamic driving. It faces an accuracy-dissipation trade-off that is qualitatively distinct from and worse than implied by the bound, and more complex steady-state copy processes cannot perform better. Nonetheless, the cost remains close to the thermodynamic bound unless accuracy is extremely high. Additionally, we show that biomolecular reactions could be used in thermodynamically optimal devices under exogenous manipulation of chemical fuels, suggesting an experimental system for testing computational thermodynamics.

Biochemical Diagnosis in Substance and Non-substance Addiction.

PubMed

Shen, Wenwen; Liu, Huifeng; Xie, Xiaohu; Liu, Haixiong; Zhou, Wenhua

2017-01-01

An optimal biochemical marker for addiction would be some easily traced molecules in body specimens, which indicates indulgent addictive behaviors, or susceptibility to certain addictive stimuli. In this chapter, we discussed existing literature about possible biomarkers, and classified them into three categories: origin forms and metabolites of substances, markers from biochemical responses to certain addiction, and genetic and epigenetic biomarkers suggesting susceptibility to addiction. In every category, we examined studies concerning certain type of addiction one by one, with focuses mainly on opiates, psychostimulants, and pathological gambling. Several promising molecules were highlighted, including those of neurotrophic factors, inflammatory factors, and indicators of vascular injury, and genetic and epigenetic biomarkers such as serum miRNAs. DNA methylation signatures and signal nucleotide polymorphism of candidate gene underlying the addiction.

Integration of electrochemistry in micro-total analysis systems for biochemical assays: recent developments.

PubMed

Xu, Xiaoli; Zhang, Song; Chen, Hui; Kong, Jilie

2009-11-15

Micro-total analysis systems (microTAS) integrate different analytical operations like sample preparation, separation and detection into a single microfabricated device. With the outstanding advantages of low cost, satisfactory analytical efficiency and flexibility in design, highly integrated and miniaturized devices from the concept of microTAS have gained widespread applications, especially in biochemical assays. Electrochemistry is shown to be quite compatible with microanalytical systems for biochemical assays, because of its attractive merits such as simplicity, rapidity, high sensitivity, reduced power consumption, and sample/reagent economy. This review presents recent developments in the integration of electrochemistry in microdevices for biochemical assays. Ingenious microelectrode design and fabrication methods, and versatility of electrochemical techniques are involved. Practical applications of such integrated microsystem in biochemical assays are focused on in situ analysis, point-of-care testing and portable devices. Electrochemical techniques are apparently suited to microsystems, since easy microfabrication of electrochemical elements and a high degree of integration with multi-analytical functions can be achieved at low cost. Such integrated microsystems will play an increasingly important role for analysis of small volume biochemical samples. Work is in progress toward new microdevice design and applications.

Performance limits and trade-offs in entropy-driven biochemical computers.

PubMed

Chu, Dominique

2018-04-14

It is now widely accepted that biochemical reaction networks can perform computations. Examples are kinetic proof reading, gene regulation, or signalling networks. For many of these systems it was found that their computational performance is limited by a trade-off between the metabolic cost, the speed and the accuracy of the computation. In order to gain insight into the origins of these trade-offs, we consider entropy-driven computers as a model of biochemical computation. Using tools from stochastic thermodynamics, we show that entropy-driven computation is subject to a trade-off between accuracy and metabolic cost, but does not involve time-trade-offs. Time trade-offs appear when it is taken into account that the result of the computation needs to be measured in order to be known. We argue that this measurement process, although usually ignored, is a major contributor to the cost of biochemical computation. Copyright © 2018 Elsevier Ltd. All rights reserved.

Identification of biochemical features of defective Coffea arabica L. beans.

PubMed

Casas, María I; Vaughan, Michael J; Bonello, Pierluigi; McSpadden Gardener, Brian; Grotewold, Erich; Alonso, Ana P

2017-05-01

Coffee organoleptic properties are based in part on the quality and chemical composition of coffee beans. The presence of defective beans during processing and roasting contribute to off flavors and reduce overall cup quality. A multipronged approach was undertaken to identify specific biochemical markers for defective beans. To this end, beans were split into defective and non-defective fractions and biochemically profiled in both green and roasted states. A set of 17 compounds in green beans, including organic acids, amino acids and reducing sugars; and 35 compounds in roasted beans, dominated by volatile compounds, organic acids, sugars and sugar alcohols, were sufficient to separate the defective and non-defective fractions. Unsorted coffee was examined for the presence of the biochemical markers to test their utility in detecting defective beans. Although the green coffee marker compounds were found in all fractions, three of the roasted coffee marker compounds (1-methylpyrrole, 5-methyl- 2-furfurylfuran, and 2-methylfuran) were uniquely present in defective fractions. Published by Elsevier Ltd.

Label-Free Imaging and Biochemical Characterization of Bovine Sperm Cells

PubMed Central

Ferrara, Maria Antonietta; Di Caprio, Giuseppe; Managò, Stefano; De Angelis, Annalisa; Sirleto, Luigi; Coppola, Giuseppe; De Luca, Anna Chiara

2015-01-01

A full label-free morphological and biochemical characterization is desirable to select spermatozoa during preparation for artificial insemination. In order to study these fundamental parameters, we take advantage of two attractive techniques: digital holography (DH) and Raman spectroscopy (RS). DH presents new opportunities for studying morphological aspect of cells and tissues non-invasively, quantitatively and without the need for staining or tagging, while RS is a very specific technique allowing the biochemical analysis of cellular components with a spatial resolution in the sub-micrometer range. In this paper, morphological and biochemical bovine sperm cell alterations were studied using these techniques. In addition, a complementary DH and RS study was performed to identify X- and Y-chromosome-bearing sperm cells. We demonstrate that the two techniques together are a powerful and highly efficient tool elucidating some important criterions for sperm morphological selection and sex-identification, overcoming many of the limitations associated with existing protocols. PMID:25836358

Slot-waveguide biochemical sensor.

PubMed

Barrios, Carlos A; Gylfason, Kristinn B; Sánchez, Benito; Griol, Amadeu; Sohlström, H; Holgado, M; Casquel, R

2007-11-01

We report an experimental demonstration of an integrated biochemical sensor based on a slot-waveguide microring resonator. The microresonator is fabricated on a Si3N4-SiO2 platform and operates at a wavelength of 1.3 microm. The transmission spectrum of the sensor is measured with different ambient refractive indices ranging from n=1.33 to 1.42. A linear shift of the resonant wavelength with increasing ambient refractive index of 212 nm/refractive index units (RIU) is observed. The sensor detects a minimal refractive index variation of 2x10(-4) RIU.

40 CFR 180.1127 - Biochemical pesticide plant floral volatile attractant compounds: cinnamaldehyde, cinnamyl...

Code of Federal Regulations, 2010 CFR

2010-07-01

... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Biochemical pesticide plant floral... (CONTINUED) PESTICIDE PROGRAMS TOLERANCES AND EXEMPTIONS FOR PESTICIDE CHEMICAL RESIDUES IN FOOD Exemptions From Tolerances § 180.1127 Biochemical pesticide plant floral volatile attractant compounds...

40 CFR 180.1127 - Biochemical pesticide plant floral volatile attractant compounds: cinnamaldehyde, cinnamyl...

Code of Federal Regulations, 2011 CFR

2011-07-01

... 40 Protection of Environment 24 2011-07-01 2011-07-01 false Biochemical pesticide plant floral... (CONTINUED) PESTICIDE PROGRAMS TOLERANCES AND EXEMPTIONS FOR PESTICIDE CHEMICAL RESIDUES IN FOOD Exemptions From Tolerances § 180.1127 Biochemical pesticide plant floral volatile attractant compounds...

Biochemical recurrence after radical prostatectomy: what does it mean?

PubMed Central

Tourinho-Barbosa, Rafael; Srougi, Victor; Nunes-Silva, Igor; Baghdadi, Mohammed; Rembeyo, Gregory; Eiffel, Sophie S.; Barret, Eric; Rozet, Francois; Galiano, Marc; Cathelineau, Xavier; Sanchez-Salas, Rafael

2018-01-01

ABSTRACT Background Radical prostatectomy (RP) has been used as the main primary treatment for prostate cancer (PCa) for many years with excellent oncologic results. However, approximately 20-40% of those patients has failed to RP and presented biochemical recurrence (BCR). Prostatic specific antigen (PSA) has been the pivotal tool for recurrence diagnosis, but there is no consensus about the best PSA threshold to define BCR until this moment. The natural history of BCR after surgical procedure is highly variable, but it is important to distinguish biochemical and clinical recurrence and to find the correct timing to start multimodal treatment strategy. Also, it is important to understand the role of each clinical and pathological feature of prostate cancer in BCR, progression to metastatic disease and cancer specific mortality (CSM). Review design A simple review was made in Medline for articles written in English language about biochemical recurrence after radical prostatectomy. Objective To provide an updated assessment of BCR definition, its meaning, PCa natural history after BCR and the weight of each clinical/pathological feature and risk group classifications in BCR, metastatic disease and CSM. PMID:29039897

Biochemical and nutritional components of selected honey samples.

PubMed

Chua, Lee Suan; Adnan, Nur Ardawati

2014-01-01

The purpose of this study was to investigate the relationship of biochemical (enzymes) and nutritional components in the selected honey samples from Malaysia. The relationship is important to estimate the quality of honey based on the concentration of these nutritious components. Such a study is limited for honey samples from tropical countries with heavy rainfall throughout the year. A number of six honey samples that commonly consumed by local people were collected for the study. Both the biochemical and nutritional components were analysed by using standard methods from Association of Official Analytical Chemists (AOAC). Individual monosaccharides, disaccharides and 17 amino acids in honey were determined by using liquid chromatographic method. The results showed that the peroxide activity was positively correlated with moisture content (r = 0.8264), but negatively correlated with carbohydrate content (r = 0.7755) in honey. The chromatographic sugar and free amino acid profiles showed that the honey samples could be clustered based on the type and maturity of honey. Proline explained for 64.9% of the total variance in principle component analysis (PCA). The correlation between honey components and honey quality has been established for the selected honey samples based on their biochemical and nutritional concentrations. PCA results revealed that the ratio of sucrose to maltose could be used to measure honey maturity, whereas proline was the marker compound used to distinguish honey either as floral or honeydew.

Biochemical and Cellular Assessment of Acetabular Chondral Flaps Identified During Hip Arthroscopy.

PubMed

Hariri, Sanaz; Truntzer, Jeremy; Smith, Robert Lane; Safran, Marc R

2015-06-01

To analyze chondral flaps debrided during hip arthroscopy to determine their biochemical and cellular composition. Thirty-one full-thickness acetabular chondral flaps were collected during hip arthroscopy. Biochemical analysis was undertaken in 21 flaps from 20 patients, and cellular viability was determined in 10 flaps from 10 patients. Biochemical analysis included concentrations of (1) DNA (an indicator of chondrocyte content), (2) hydroxyproline (an indicator of collagen content), and (3) glycosaminoglycan (an indicator of chondrocyte biosynthesis). Higher values for these parameters indicated more healthy tissue. The flaps were examined to determine the percentage of viable chondrocytes. The percentage of acetabular chondral flap specimens that had concentrations within 1 SD of the mean values reported in previous normal cartilage studies was 38% for DNA, 0% for glycosaminoglycan, and 43% for hydroxyproline. The average cellular viability of our acetabular chondral flap specimens was 39% (SD, 14%). Only 2 of the 10 specimens had more than half the cells still viable. There was no correlation between (1) the gross examination of the joint or knowledge of the patient's demographic characteristics and symptoms and (2) biochemical properties and cell viability of the flap, with one exception: a degenerative appearance of the surrounding cartilage correlated with a higher hydroxyproline concentration. Although full-thickness acetabular chondral flaps can appear normal grossly, the biochemical properties and percentage of live chondrocytes in full-thickness chondral flaps encountered in hip arthroscopy show that this tissue is not normal. There has been recent interest in repairing chondral flaps encountered during hip arthroscopy. These data suggest that acetabular chondral flaps are not biochemically and cellularly normal. Although these flaps may still be valuable mechanically and/or as a scaffold in some conductive or inductive capacity, further study is

Metabonomics and medicine: the Biochemical Oracle.

PubMed

Mitchell, Steve; Holmes, Elaine; Carmichael, Paul

2002-10-01

Occasionally, a new idea emerges that has the potential to revolutionize an entire field of scientific endeavour. It is now within our grasp to be able to detect subtle perturbations within the phenomenally complex biochemical matrix of living organisms. The discipline of metabonomics promises an all-encompassing approach to understanding total, yet fundamental, changes occurring in disease processes, drug toxicity and cell function.

Feasibility of biochemical verification in a web-based smoking cessation study.

PubMed

Cha, Sarah; Ganz, Ollie; Cohn, Amy M; Ehlke, Sarah J; Graham, Amanda L

2017-10-01

Cogent arguments have been made against the need for biochemical verification in population-based studies with low-demand characteristics. Despite this fact, studies involving digital interventions (low-demand) are often required in peer review to report biochemically verified abstinence. To address this discrepancy, we examined the feasibility and costs of biochemical verification in a web-based study conducted with a national sample. Participants were 600U.S. adult current smokers who registered on a web-based smoking cessation program and completed surveys at baseline and 3months. Saliva sampling kits were sent to participants who reported 7-day abstinence at 3months, and analyzed for cotinine. The response rate at 3-months was 41.2% (n=247): 93 participants reported 7-day abstinence (38%) and were mailed a saliva kit (71% returned). The discordance rate was 36.4%. Participants with discordant responses were more likely to report 3-month use of nicotine replacement therapy or e-cigarettes than those with concordant responses (79.2% vs. 45.2%, p=0.007). The total cost of saliva sampling was $8280 ($125/sample). Biochemical verification was both time- and cost-intensive, and yielded a relatively small number of samples due to low response rates and use of other nicotine products during the follow-up period. There was a high rate of discordance of self-reported abstinence and saliva testing. Costs for data collection may be prohibitive for studies with large sample sizes or limited budgets. Our findings echo previous statements that biochemical verification is not necessary in population-based studies, and add evidence specific to technology-based studies. Copyright © 2017 Elsevier Ltd. All rights reserved.

Biochemical abnormalities in Pearson syndrome.

PubMed

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.

Physiological, Biochemical, and Molecular Mechanisms of Heat Stress Tolerance in Plants

PubMed Central

Hasanuzzaman, Mirza; Nahar, Kamrun; Alam, Md. Mahabub; Roychowdhury, Rajib; Fujita, Masayuki

2013-01-01

High temperature (HT) stress is a major environmental stress that limits plant growth, metabolism, and productivity worldwide. Plant growth and development involve numerous biochemical reactions that are sensitive to temperature. Plant responses to HT vary with the degree and duration of HT and the plant type. HT is now a major concern for crop production and approaches for sustaining high yields of crop plants under HT stress are important agricultural goals. Plants possess a number of adaptive, avoidance, or acclimation mechanisms to cope with HT situations. In addition, major tolerance mechanisms that employ ion transporters, proteins, osmoprotectants, antioxidants, and other factors involved in signaling cascades and transcriptional control are activated to offset stress-induced biochemical and physiological alterations. Plant survival under HT stress depends on the ability to perceive the HT stimulus, generate and transmit the signal, and initiate appropriate physiological and biochemical changes. HT-induced gene expression and metabolite synthesis also substantially improve tolerance. The physiological and biochemical responses to heat stress are active research areas, and the molecular approaches are being adopted for developing HT tolerance in plants. This article reviews the recent findings on responses, adaptation, and tolerance to HT at the cellular, organellar, and whole plant levels and describes various approaches being taken to enhance thermotolerance in plants. PMID:23644891

In silico characterization of microbial electrosynthesis for metabolic engineering of biochemicals

PubMed Central

2011-01-01

Background A critical concern in metabolic engineering is the need to balance the demand and supply of redox intermediates such as NADH. Bioelectrochemical techniques offer a novel and promising method to alleviate redox imbalances during the synthesis of biochemicals and biofuels. Broadly, these techniques reduce intracellular NAD+ to NADH and therefore manipulate the cell's redox balance. The cellular response to such redox changes and the additional reducing power available to the cell can be harnessed to produce desired metabolites. In the context of microbial fermentation, these bioelectrochemical techniques can be used to improve product yields and/or productivity. Results We have developed a method to characterize the role of bioelectrosynthesis in chemical production using the genome-scale metabolic model of E. coli. The results in this paper elucidate the role of bioelectrosynthesis and its impact on biomass growth, cellular ATP yields and biochemical production. The results also suggest that strain design strategies can change for fermentation processes that employ microbial electrosynthesis and suggest that dynamic operating strategies lead to maximizing productivity. Conclusions The results in this paper provide a systematic understanding of the benefits and limitations of bioelectrochemical techniques for biochemical production and highlight how electrical enhancement can impact cellular metabolism and biochemical production. PMID:21967745

Flow Injection Technique for Biochemical Analysis with Chemiluminescence Detection in Acidic Media

PubMed Central

Chen, Jing; Fang, Yanjun

2007-01-01

A review with 90 references is presented to show the development of acidic chemiluminescence methods for biochemical analysis by use of flow injection technique in the last 10 years. A brief discussion of both the chemiluminescence and flow injection technique is given. The proposed methods for biochemical analysis are described and compared according to the used chemiluminescence system.

Label-free optical resonant sensors for biochemical applications

NASA Astrophysics Data System (ADS)

Ciminelli, Caterina; Campanella, Clarissa Martina; Dell'Olio, Francesco; Campanella, Carlo Edoardo; Armenise, Mario Nicola

2013-03-01

For a number of years, the scientific community has been paying growing attention to the monitoring and enhancement of public health and the quality of life through the detection of all dangerous agents for the human body, including gases, proteins, virus, and bacterial agents. When these agents are detected through label-free biochemical sensors, the molecules are not modified structurally or functionally by adding fluorescent or radioactive dyes. This work focuses on label-free optical ring resonator-based configurations suited for bio-chemical sensing, highlighting their physical aspects and specific applications. Resonant wavelength shift and the modal splitting occurring when the analyte interacts with microresonant structures are the two major physical aspects analyzed in this paper. Competitive optical platforms proposed in the literature are also illustrated together with their properties and performance.

Exploring the remote sensing of foliar biochemical concentrations with AVIRIS data

NASA Technical Reports Server (NTRS)

Smith, Geoffrey M.; Curran, Paul J.

1992-01-01

Airborne Visible/Infrared Imaging Spectrometer (AVIRIS) data shows promise for the estimation of foliar biochemical concentrations at the scale of the canopy. There are, however, several problems associated with the use of AVIRIS data in this way and these are detailed in recent Plant Biochemical Workshop Report. The research reported was concentrated upon three of these problems: field sampling of forest canopies, wet laboratory assay of foliar chemicals, and the visualization of AVIRIS data.

[Biochemical principles of early saturnism recognition].

PubMed

Tsimakuridze, M P; Mansuradze, E A; Zurashvili, D G; Tsimakuridze, M P

2009-03-01

The aim of the work is to determine the major sensitive criteria of biochemical indicators that allow timely discovery of negative influence of lead on organism and assist in early diagnosis of primary stages of saturnism. The workers of Georgian typographies, performing technological processes of letterpress printing were observed. Professional groups having contact with lead aerosols (main group of 66 people) and the workers of the same typography not being in touch with the poison (control group of 24 people) were studied. It was distinguished that, protracted professional contact with lead causes moderate increase of lead, coproporphyrin and DALA in daily urine in most cases; it is more clearly evidenced in the professional groups of lead smelters and lino operators and less clearly among typesetter and printers. Upon the checkup of people, having a direct contact with lead, biochemical analysis of urine should be given a preference, especially the determination of quantitative content of lead and coproporphyrin in urine with the aim of revealing the lead carrier, which is one of the first signals for occupational lookout and medical monitoring of the similar contingent.

Pheochromocytoma-paraganglioma: Biochemical and genetic diagnosis.

PubMed

Cano Megías, Marta; Rodriguez Puyol, Diego; Fernández Rodríguez, Loreto; Sención Martinez, Gloria Lisette; Martínez Miguel, Patricia

Pheochromocytomas and paragangliomas are tumours derived from neural crest cells, which can be diagnosed by biochemical measurement of metanephrine and methoxytyramine. Advances in genetic research have identified many genes involved in the pathogenesis of these tumours, suggesting that up to 35-45% may have an underlying germline mutation. These genes have a singular transcriptional signature and can be grouped into 2 clusters (or groups): cluster 1 (VHL and SHDx), involved in angiogenesis and hypoxia pathways; and cluster 2 (MEN2 and NF1), linked to the kinase signalling pathway. In turn, these genes are associated with a characteristic biochemical phenotype (noradrenergic and adrenergic), and clinical features (location, biological behaviour, age of presentation, etc.) in a large number of cases. Early diagnosis of these tumours, accompanied by a correct genetic diagnosis, should eventually become a priority to enable better treatment, early detection of complications, proper screening of family members and related tumours, as well as an improvement in the overall prognosis of these patients. Copyright © 2016 Sociedad Española de Nefrología. Published by Elsevier España, S.L.U. All rights reserved.

Physiological, biochemical and transcriptional analysis of onion bulbs during storage

PubMed Central

Chope, Gemma A.; Cools, Katherine; Hammond, John P.; Thompson, Andrew J.; Terry, Leon A.

2012-01-01

Background and Aims During the transition from endo-dormancy to eco-dormancy and subsequent growth, the onion bulb undergoes the transition from sink organ to source, to sustain cell division in the meristematic tissue. The mechanisms controlling these processes are not fully understood. Here, a detailed analysis of whole onion bulb physiological, biochemical and transcriptional changes in response to sprouting is reported, enabling a better knowledge of the mechanisms regulating post-harvest onion sprout development. Methods Biochemical and physiological analyses were conducted on different cultivars (‘Wellington’, ‘Sherpa’ and ‘Red Baron’) grown at different sites over 3 years, cured at different temperatures (20, 24 and 28 °C) and stored under different regimes (1, 3, 6 and 6 → 1 °C). In addition, the first onion oligonucleotide microarray was developed to determine differential gene expression in onion during curing and storage, so that transcriptional changes could support biochemical and physiological analyses. Key Results There were greater transcriptional differences between samples at harvest and before sprouting than between the samples taken before and after sprouting, with some significant changes occurring during the relatively short curing period. These changes are likely to represent the transition from endo-dormancy to sprout suppression, and suggest that endo-dormancy is a relatively short period ending just after curing. Principal component analysis of biochemical and physiological data identified the ratio of monosaccharides (fructose and glucose) to disaccharide (sucrose), along with the concentration of zeatin riboside, as important factors in discriminating between sprouting and pre-sprouting bulbs. Conclusions These detailed analyses provide novel insights into key regulatory triggers for sprout dormancy release in onion bulbs and provide the potential for the development of biochemical or transcriptional markers for sprout

Biochemical Applications in the Analytical Chemistry Lab

ERIC Educational Resources Information Center

Strong, Cynthia; Ruttencutter, Jeffrey

2004-01-01

An HPLC and a UV-visible spectrophotometer are identified as instruments that helps to incorporate more biologically-relevant experiments into the course, in order to increase the students understanding of selected biochemistry topics and enhances their ability to apply an analytical approach to biochemical problems. The experiment teaches…

Survey of Biochemical Education in Japanese Universities.

ERIC Educational Resources Information Center

Kagawa, Yasuo

1995-01-01

Reports findings of questionnaires sent to faculty in charge of biochemical education in medical schools and other programs from dentistry to agriculture. Total class hours have declined since 1984. New trends include bioethics and computer-assisted learning. Tables show trends in lecture hours, lecture content, laboratory hours, core subject…

40 CFR 158.2082 - Experimental use permit biochemical pesticides residue data requirements table.

Code of Federal Regulations, 2010 CFR

2010-07-01

... biochemical agents in the biochemical human health assessment data requirements, § 158.2050. 2. The same... human or livestock consumption. 12. Data on fish are required for all pesticides applied directly to... pesticides residue data requirements table. 158.2082 Section 158.2082 Protection of Environment ENVIRONMENTAL...

40 CFR 158.2082 - Experimental use permit biochemical pesticides residue data requirements table.

Code of Federal Regulations, 2011 CFR

2011-07-01

... biochemical agents in the biochemical human health assessment data requirements, § 158.2050. 2. The same... human or livestock consumption. 12. Data on fish are required for all pesticides applied directly to... pesticides residue data requirements table. 158.2082 Section 158.2082 Protection of Environment ENVIRONMENTAL...

40 CFR 158.2082 - Experimental use permit biochemical pesticides residue data requirements table.

Code of Federal Regulations, 2012 CFR

2012-07-01

... biochemical agents in the biochemical human health assessment data requirements, § 158.2050. 2. The same... human or livestock consumption. 12. Data on fish are required for all pesticides applied directly to... pesticides residue data requirements table. 158.2082 Section 158.2082 Protection of Environment ENVIRONMENTAL...

40 CFR 158.2082 - Experimental use permit biochemical pesticides residue data requirements table.

Code of Federal Regulations, 2014 CFR

2014-07-01

... biochemical agents in the biochemical human health assessment data requirements, § 158.2050. 2. The same... human or livestock consumption. 12. Data on fish are required for all pesticides applied directly to... pesticides residue data requirements table. 158.2082 Section 158.2082 Protection of Environment ENVIRONMENTAL...

40 CFR 158.2082 - Experimental use permit biochemical pesticides residue data requirements table.

Code of Federal Regulations, 2013 CFR

2013-07-01

... biochemical agents in the biochemical human health assessment data requirements, § 158.2050. 2. The same... human or livestock consumption. 12. Data on fish are required for all pesticides applied directly to... pesticides residue data requirements table. 158.2082 Section 158.2082 Protection of Environment ENVIRONMENTAL...

Salivary testosterone may not serve as a screening test in the diagnosis of biochemical hyperandrogenism.

PubMed

Ambroziak, Urszula; Kuryłowicz, Alina; Kępczyńska-Nyk, Anna; Kondracka, Agnieszka; Gajda, Sylvia; Sieńko, Damian

2018-06-01

The diagnosis of biochemical hyperandrogenism is still challenging because a set of appropriate, recommended diagnostic tests has not been established. In our study, we aimed to answer the question of whether salivary testosterone is a reliable test to establish the diagnosis of biochemical hyperandrogenism as compared to serum total testosterone (TT) measured either by liquid chromatography-tandem mass spectrometry (LC-MS/MS) or immunoassay and to assess which set of biochemical tests would be the most appropriate for the identification of biochemical hyperandrogenism. A total of 39 women, aged 18-45 years, with clinical or biochemical hyperandrogenism and 41 healthy individuals, aged 19-45 years, were enrolled in the study. Salivary testosterone was measured using the Salimetrics test. Serum TT was measured either using the LC-MS/MS method or immunoassay, and dehydroepiandrosterone sulphate (DHEA-S) and androstenedione were measured using LC-MS/MS. In 15 of 17 (88%) patients with elevated serum TT measured by LC-MS/MS and in 14 of 16 (87%) measured with immunoassay, salivary testosterone showed normal levels. In 11 of 39 women (28%) with normal serum testosterone levels, DHEA-S was elevated. All patients with elevated androstenedione presented with an elevated concentration of either serum testosterone or DHEA-S. Salivary testosterone measurement may lead to the underdiagnosis of biochemical hyperandrogenism. Both serum testosterone and DHEA-S should be measured in the endocrine work-up toward biochemical hyperandrogenism. © 2018 Japan Society of Obstetrics and Gynecology.

Biochemical Disincentives to Fertilizing Cellulosic Ethanol Crops

NASA Astrophysics Data System (ADS)

Gallagher, M. E.; Hockaday, W. C.; Snapp, S.; McSwiney, C.; Baldock, J.

2010-12-01

Corn grain biofuel crops produce the highest yields when the cropping ecosystem is not nitrogen (N)-limited, achieved by application of fertilizer. There are environmental consequences for excessive fertilizer application to crops, including greenhouse gas emissions, hypoxic “dead zones,” and health problems from N runoff into groundwater. The increase in corn acreage in response to demand for alternative fuels (i.e. ethanol) could exacerbate these problems, and divert food supplies to fuel production. A potential substitute for grain ethanol that could reduce some of these impacts is cellulosic ethanol. Cellulosic ethanol feedstocks include grasses (switchgrass), hardwoods, and crop residues (e.g. corn stover, wheat straw). It has been assumed that these feedstocks will require similar N fertilization rates to grain biofuel crops to maximize yields, but carbohydrate yield versus N application has not previously been monitored. We report the biochemical stocks (carbohydrate, protein, and lignin in Mg ha-1) of a corn ecosystem grown under varying N levels. We measured biochemical yield in Mg ha-1 within the grain, leaf and stem, and reproductive parts of corn plants grown at seven N fertilization rates (0-202 kg N ha-1), to evaluate the quantity and quality of these feedstocks across a N fertilization gradient. The N fertilization rate study was performed at the Kellogg Biological Station-Long Term Ecological Research Site (KBS-LTER) in Michigan. Biochemical stocks were measured using 13C nuclear magnetic resonance spectroscopy (NMR), combined with a molecular mixing model (Baldock et al. 2004). Carbohydrate and lignin are the main biochemicals of interest in ethanol production since carbohydrate is the ethanol feedstock, and lignin hinders the carbohydrate to ethanol conversion process. We show that corn residue carbohydrate yields respond only weakly to N fertilization compared to grain. Grain carbohydrate yields plateau in response to fertilization at

[Biochemical changes in apoptosis and methods for their determination (review)].

PubMed

Sedláková, A; Kohút, A; Kalina, I

1999-08-01

Apoptosis or programmed cell death is a physiological process which occurs at different biological states as well as at disease process. Morphologically it is characterized by the chromatine condensation and other changes with preserved integrity of plasmatic membrane. The major and most frequently studied biochemical characteristic of apoptosis is a DNA fragmentation. In our paper attention is directed to the early biochemical changes in cell membranes, i.g., the externalization of phosphatidylserine, hydrolysis of sphingomyeline on the ceramide and activation of phospholipases especially phospholipase A2. In one part we described the changes of cysteine proteases (caspases), which play a key role in the execution of apoptosis. These biochemical changes are associated with ceramide signalization of apoptosis. Briefly are presented also some dates about apoptosis induction with reactive oxygen radicals and the role of the arachidonic acid metabolites in this process. We consider the investigation and determination of these changes as important parameters of apoptosis at some diseases, e.g., cancer or degenerative diseases, and of their treatment.

Design of a biochemical circuit motif for learning linear functions

PubMed Central

Lakin, Matthew R.; Minnich, Amanda; Lane, Terran; Stefanovic, Darko

2014-01-01

Learning and adaptive behaviour are fundamental biological processes. A key goal in the field of bioengineering is to develop biochemical circuit architectures with the ability to adapt to dynamic chemical environments. Here, we present a novel design for a biomolecular circuit capable of supervised learning of linear functions, using a model based on chemical reactions catalysed by DNAzymes. To achieve this, we propose a novel mechanism of maintaining and modifying internal state in biochemical systems, thereby advancing the state of the art in biomolecular circuit architecture. We use simulations to demonstrate that the circuit is capable of learning behaviour and assess its asymptotic learning performance, scalability and robustness to noise. Such circuits show great potential for building autonomous in vivo nanomedical devices. While such a biochemical system can tell us a great deal about the fundamentals of learning in living systems and may have broad applications in biomedicine (e.g. autonomous and adaptive drugs), it also offers some intriguing challenges and surprising behaviours from a machine learning perspective. PMID:25401175

Design of a biochemical circuit motif for learning linear functions.

PubMed

Lakin, Matthew R; Minnich, Amanda; Lane, Terran; Stefanovic, Darko

2014-12-06

Learning and adaptive behaviour are fundamental biological processes. A key goal in the field of bioengineering is to develop biochemical circuit architectures with the ability to adapt to dynamic chemical environments. Here, we present a novel design for a biomolecular circuit capable of supervised learning of linear functions, using a model based on chemical reactions catalysed by DNAzymes. To achieve this, we propose a novel mechanism of maintaining and modifying internal state in biochemical systems, thereby advancing the state of the art in biomolecular circuit architecture. We use simulations to demonstrate that the circuit is capable of learning behaviour and assess its asymptotic learning performance, scalability and robustness to noise. Such circuits show great potential for building autonomous in vivo nanomedical devices. While such a biochemical system can tell us a great deal about the fundamentals of learning in living systems and may have broad applications in biomedicine (e.g. autonomous and adaptive drugs), it also offers some intriguing challenges and surprising behaviours from a machine learning perspective.

MATLAB-Based Teaching Modules in Biochemical Engineering

ERIC Educational Resources Information Center

Lee, Kilho; Comolli, Noelle K.; Kelly, William J.; Huang, Zuyi

2015-01-01

Mathematical models play an important role in biochemical engineering. For example, the models developed in the field of systems biology have been used to identify drug targets to treat pathogens such as Pseudomonas aeruginosa in biofilms. In addition, competitive binding models for chromatography processes have been developed to predict expanded…

An Inductive Logic Programming Approach to Validate Hexose Binding Biochemical Knowledge.

PubMed

Nassif, Houssam; Al-Ali, Hassan; Khuri, Sawsan; Keirouz, Walid; Page, David

2010-01-01

Hexoses are simple sugars that play a key role in many cellular pathways, and in the regulation of development and disease mechanisms. Current protein-sugar computational models are based, at least partially, on prior biochemical findings and knowledge. They incorporate different parts of these findings in predictive black-box models. We investigate the empirical support for biochemical findings by comparing Inductive Logic Programming (ILP) induced rules to actual biochemical results. We mine the Protein Data Bank for a representative data set of hexose binding sites, non-hexose binding sites and surface grooves. We build an ILP model of hexose-binding sites and evaluate our results against several baseline machine learning classifiers. Our method achieves an accuracy similar to that of other black-box classifiers while providing insight into the discriminating process. In addition, it confirms wet-lab findings and reveals a previously unreported Trp-Glu amino acids dependency.

A novel optical fiber biochemical sensor based on long period grating

NASA Astrophysics Data System (ADS)

Mao, Xianhui; Liao, Yanbiao; Zhang, Min; Lai, Shurong; Yin, Haibo

2007-09-01

In this paper, our present work, which aimed at investigating a novel optical fiber biochemical sensor based on long period grating (LPG), is introduced. Biochemical sensor is one of the most attractive fields of sensor research, especially with the development and occurrence of all kinds of novel theory and technology such as LPG. When there is a refraction index periodic perturbation, the guiding mode and cladding mode in LPG couple with each other. This make the LPG is sensitive to the ambient refractive index. This means it can be a novel bio-chemical sensor when it is applied in the fields of biochemistry. After investigating the principle of coupling in LPG, where the formulas of resonance wave length and band width are induced by 3-layer step index model, we developed an optical fiber biochemical sensor. The structure of its probe is designed by coating some function films whose thickness is between several tens and several hundreds nanometers on the cladding of optical fiber. Experiments of monitoring the saline separateness process of Bovine Serum Albumin (BSA) and Mice-Immunoglobulin G (M-IgG) by using the developed LPG sensor have been done. The monitoring indicated that for the BSA, the saline separateness occurs when the saturation is between 50% and 60%, for the M-IgG, the percentage is between 30%-40%. Besides the monitoring, the experiments could also analyze the effects of protein type (different molecule structure), protein consistency and saline saturation to saline separateness. The experimental results show that the optical fiber biochemical sensor based on LPG has many advantages such as simple structure, high sensitivity and miniature. It has a promising future in many research fields and application fields.

Improved prediction of biochemical recurrence after radical prostatectomy by genetic polymorphisms.

PubMed

Morote, Juan; Del Amo, Jokin; Borque, Angel; Ars, Elisabet; Hernández, Carlos; Herranz, Felipe; Arruza, Antonio; Llarena, Roberto; Planas, Jacques; Viso, María J; Palou, Joan; Raventós, Carles X; Tejedor, Diego; Artieda, Marta; Simón, Laureano; Martínez, Antonio; Rioja, Luis A

2010-08-01

Single nucleotide polymorphisms are inherited genetic variations that can predispose or protect individuals against clinical events. We hypothesized that single nucleotide polymorphism profiling may improve the prediction of biochemical recurrence after radical prostatectomy. We performed a retrospective, multi-institutional study of 703 patients treated with radical prostatectomy for clinically localized prostate cancer who had at least 5 years of followup after surgery. All patients were genotyped for 83 prostate cancer related single nucleotide polymorphisms using a low density oligonucleotide microarray. Baseline clinicopathological variables and single nucleotide polymorphisms were analyzed to predict biochemical recurrence within 5 years using stepwise logistic regression. Discrimination was measured by ROC curve AUC, specificity, sensitivity, predictive values, net reclassification improvement and integrated discrimination index. The overall biochemical recurrence rate was 35%. The model with the best fit combined 8 covariates, including the 5 clinicopathological variables prostate specific antigen, Gleason score, pathological stage, lymph node involvement and margin status, and 3 single nucleotide polymorphisms at the KLK2, SULT1A1 and TLR4 genes. Model predictive power was defined by 80% positive predictive value, 74% negative predictive value and an AUC of 0.78. The model based on clinicopathological variables plus single nucleotide polymorphisms showed significant improvement over the model without single nucleotide polymorphisms, as indicated by 23.3% net reclassification improvement (p = 0.003), integrated discrimination index (p <0.001) and likelihood ratio test (p <0.001). Internal validation proved model robustness (bootstrap corrected AUC 0.78, range 0.74 to 0.82). The calibration plot showed close agreement between biochemical recurrence observed and predicted probabilities. Predicting biochemical recurrence after radical prostatectomy based on

Scalable Parameter Estimation for Genome-Scale Biochemical Reaction Networks

PubMed Central

Kaltenbacher, Barbara; Hasenauer, Jan

2017-01-01

Mechanistic mathematical modeling of biochemical reaction networks using ordinary differential equation (ODE) models has improved our understanding of small- and medium-scale biological processes. While the same should in principle hold for large- and genome-scale processes, the computational methods for the analysis of ODE models which describe hundreds or thousands of biochemical species and reactions are missing so far. While individual simulations are feasible, the inference of the model parameters from experimental data is computationally too intensive. In this manuscript, we evaluate adjoint sensitivity analysis for parameter estimation in large scale biochemical reaction networks. We present the approach for time-discrete measurement and compare it to state-of-the-art methods used in systems and computational biology. Our comparison reveals a significantly improved computational efficiency and a superior scalability of adjoint sensitivity analysis. The computational complexity is effectively independent of the number of parameters, enabling the analysis of large- and genome-scale models. Our study of a comprehensive kinetic model of ErbB signaling shows that parameter estimation using adjoint sensitivity analysis requires a fraction of the computation time of established methods. The proposed method will facilitate mechanistic modeling of genome-scale cellular processes, as required in the age of omics. PMID:28114351

Study on color difference estimation method of medicine biochemical analysis

NASA Astrophysics Data System (ADS)

Wang, Chunhong; Zhou, Yue; Zhao, Hongxia; Sun, Jiashi; Zhou, Fengkun

2006-01-01

The biochemical analysis in medicine is an important inspection and diagnosis method in hospital clinic. The biochemical analysis of urine is one important item. The Urine test paper shows corresponding color with different detection project or different illness degree. The color difference between the standard threshold and the test paper color of urine can be used to judge the illness degree, so that further analysis and diagnosis to urine is gotten. The color is a three-dimensional physical variable concerning psychology, while reflectance is one-dimensional variable; therefore, the estimation method of color difference in urine test can have better precision and facility than the conventional test method with one-dimensional reflectance, it can make an accurate diagnose. The digital camera is easy to take an image of urine test paper and is used to carry out the urine biochemical analysis conveniently. On the experiment, the color image of urine test paper is taken by popular color digital camera and saved in the computer which installs a simple color space conversion (RGB -> XYZ -> L *a *b *)and the calculation software. Test sample is graded according to intelligent detection of quantitative color. The images taken every time were saved in computer, and the whole illness process will be monitored. This method can also use in other medicine biochemical analyses that have relation with color. Experiment result shows that this test method is quick and accurate; it can be used in hospital, calibrating organization and family, so its application prospect is extensive.

[Analysis of biochemical markers in serum of guinea pigs after death caused by hypothermia].

PubMed

Li, Shi-ying; Deng, Kai-fei; Shao, Yu; Li, Zheng-dong; Qin, Zhi-qiang; Chen, Yi-jiu; Huang, Ping

2014-08-01

To explore the changes and rules of biochemical markers in serum of guinea pigs after death caused by hypothermia and to provide references for fatal hypothermia diagnosis by serum biochemical markers. Twenty guinea pigs were randomly divided into experimental group and control group. The guinea pigs in the experimental group were kept at -30 °C until death, while the ones in control group were decapitated after same survival intervals at 25 °C. The serum was extracted from the whole blood of right ventricular immediately. Subsequently, a series of serum biochemical markers were analyzed by auto bio-chemical analyzer. The levels of glucose, uric acid, creatinine and urea nitrogen in the experimental group were significantly higher than those in control group, respectively (P<0.05). Compared with the control group, the levels of total protein and albumin were significantly lower in the experimental group (P<0.05). There were no significantly differences of the levels of other markers such as serum enzymes and ions observed between the two groups. There are characteristic changes of some specific serum biochemical markers in fatal hypothermia, which may be potentially useful for auxiliary diagnosis of fatal hypothermia.

Linear analysis near a steady-state of biochemical networks: control analysis, correlation metrics and circuit theory.

PubMed

Heuett, William J; Beard, Daniel A; Qian, Hong

2008-05-15

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. 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 RTBS and STBS 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. 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).

BioCluster: tool for identification and clustering of Enterobacteriaceae based on biochemical data.

PubMed

Abdullah, Ahmed; Sabbir Alam, S M; Sultana, Munawar; Hossain, M Anwar

2015-06-01

Presumptive identification of different Enterobacteriaceae species is routinely achieved based on biochemical properties. Traditional practice includes manual comparison of each biochemical property of the unknown sample with known reference samples and inference of its identity based on the maximum similarity pattern with the known samples. This process is labor-intensive, time-consuming, error-prone, and subjective. Therefore, automation of sorting and similarity in calculation would be advantageous. Here we present a MATLAB-based graphical user interface (GUI) tool named BioCluster. This tool was designed for automated clustering and identification of Enterobacteriaceae based on biochemical test results. In this tool, we used two types of algorithms, i.e., traditional hierarchical clustering (HC) and the Improved Hierarchical Clustering (IHC), a modified algorithm that was developed specifically for the clustering and identification of Enterobacteriaceae species. IHC takes into account the variability in result of 1-47 biochemical tests within this Enterobacteriaceae family. This tool also provides different options to optimize the clustering in a user-friendly way. Using computer-generated synthetic data and some real data, we have demonstrated that BioCluster has high accuracy in clustering and identifying enterobacterial species based on biochemical test data. This tool can be freely downloaded at http://microbialgen.du.ac.bd/biocluster/. Copyright © 2015 The Authors. Production and hosting by Elsevier Ltd.. All rights reserved.

CELLULAR, BIOCHEMICAL, AND MOLECULAR TECHNIQUES IN DEVELOPMENTAL TOXICOLOGY

EPA Science Inventory

Cellular, molecular and biochemical approaches vastly expand the possibilities for revealing the underlying mechanisms of developmental toxicity. The increasing interest in embryonic development as a model system for the study of gene expression has resulted in a cornucopia of i...

Lipemia interferences in routine clinical biochemical tests.

PubMed

Calmarza, Pilar; Cordero, José

2011-01-01

Lipemic specimens are a common and frequent, but yet unresolved problem in clinical chemistry, and may produce significant interferences in the analytical results of different biochemical parameters. The aim of this study was to examine the effect of lipid removal using ultracentrifugation of lipemic samples, on some routine biochemistry parameters. Among all the samples obtained daily in our laboratory, the ones which were visibly muddy were selected and underwent to a process of ultracentrifugation, being determined a variety of biochemical tests before and after ultracentrifugation. A total of 110 samples were studied. We found significant differences in all the parameters studied except for total bilirubin, glucose, gamma-glutamyl transferase (GGT) and aspartate aminotransferase (AST). The greatest differences in the parameters analyzed were found in the concentration of alanine aminotransferase (ALT) (7.36%) and the smallest ones in the concentration of glucose (0.014%). Clinically significant interferences were found for phosphorus, creatinine, total protein and calcium. Lipemia causes clinically significant interferences for phosphorus, creatinine, total protein and calcium measurement and those interferences could be effectively removed by ultracentrifugation.

Adult amphibian epidermal proteins: biochemical characterization and developmental appearance.

PubMed

Reeves, O R

1975-08-01

The keratin-like proteins (KLPs) from the epidermis of adult frogs of the species Xenopus laevis have been isolated and biochemically characterized by means of polyacrylamide gel electrophoresis, amino acid analysis, tryptic peptide mapping, amino-terminal end-group analysis and isoelectric focusing. One particular protein fraction of rather unusual amino acid composition found only in epidermal tissue was isolated in quantity by preparative gel electrophoresis and monospecific antibodies prepared against it. Using this anti-KLP antibody preparation it was possible to show that at least one kine of keratin-like protein characteristic of the adult epidermis first appears within the larval epidermis during metamorphosis. This is the first reported biochemical characterization of a tissue-specific protien from adult amphibian skin.

Exercise-induced biochemical changes and their potential influence on cancer: a scientific review

PubMed Central

Thomas, Robert James; Kenfield, Stacey A; Jimenez, Alfonso

2017-01-01

Aim To review and discuss the available international literature regarding the indirect and direct biochemical mechanisms that occur after exercise, which could positively, or negatively, influence oncogenic pathways. Methods The PubMed, MEDLINE, Embase and Cochrane libraries were searched for papers up to July 2016 addressing biochemical changes after exercise with a particular reference to cancer. The three authors independently assessed their appropriateness for inclusion in this review based on their scientific quality and relevance. Results 168 papers were selected and categorised into indirect and direct biochemical pathways. The indirect effects included changes in vitamin D, weight reduction, sunlight exposure and improved mood. The direct effects included insulin-like growth factor, epigenetic effects on gene expression and DNA repair, vasoactive intestinal peptide, oxidative stress and antioxidant pathways, heat shock proteins, testosterone, irisin, immunity, chronic inflammation and prostaglandins, energy metabolism and insulin resistance. Summary Exercise is one of several lifestyle factors known to lower the risk of developing cancer and is associated with lower relapse rates and better survival. This review highlights the numerous biochemical processes, which explain these potential anticancer benefits. PMID:27993842

Linear analysis near a steady-state of biochemical networks: Control analysis, correlation metrics and circuit theory

PubMed Central

Heuett, William J; Beard, Daniel A; Qian, Hong

2008-01-01

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 RTBS and STBS 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). PMID:18482450

The biochemical properties of antibodies and their fragments

USDA-ARS?s Scientific Manuscript database

Immunoglobulins (Ig) or antibodies are a powerful molecular recognition tools that can be used to identify minute quantities of a given target analyte. Their antigen binding properties define both the sensitivity and selectivity of an immunoassay. Understanding the biochemical properties of this c...

Structure and biochemical characterization of proliferating cellular nuclear antigen from a parasitic protozoon

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cardona-Felix, Cesar S.; Lara-Gonzalez, Samuel; Brieba, Luis G.

2012-02-08

Proliferating cellular nuclear antigen (PCNA) is a toroidal-shaped protein that is involved in cell-cycle control, DNA replication and DNA repair. Parasitic protozoa are early-diverged eukaryotes that are responsible for neglected diseases. In this work, a PCNA from a parasitic protozoon was identified, cloned and biochemically characterized and its crystal structure was determined. Structural and biochemical studies demonstrate that PCNA from Entamoeba histolytica assembles as a homotrimer that is able to interact with and stimulate the activity of a PCNA-interacting peptide-motif protein from E. histolytica, EhDNAligI. The data indicate a conservation of the biochemical mechanisms of PCNA-mediated interactions between metazoa, yeastmore » and parasitic protozoa.« less

Hierarchical thinking in network biology: the unbiased modularization of biochemical networks.

PubMed

Papin, Jason A; Reed, Jennifer L; Palsson, Bernhard O

2004-12-01

As reconstructed biochemical reaction networks continue to grow in size and scope, there is a growing need to describe the functional modules within them. Such modules facilitate the study of biological processes by deconstructing complex biological networks into conceptually simple entities. The definition of network modules is often based on intuitive reasoning. As an alternative, methods are being developed for defining biochemical network modules in an unbiased fashion. These unbiased network modules are mathematically derived from the structure of the whole network under consideration.

Chapter A7. Section 7.0. Five-Day Biochemical Oxygen Demand

USGS Publications Warehouse

Delzer, Gregory C.; McKenzie, Stuart W.

1999-01-01

The presence of a sufficient concentration of dissolved oxygen is critical to maintaining the aquatic life and aesthetic quality of streams and lakes. Determinng how organic matter affects the concentration of dissolved oxygen (DO) in a stream or lake is integral to water-quality management. The decay of organic matter in water is measured as biochemical or chemical oxygen demand. This report describes the field protocols used by U.S. Geological Survey (USGS) personnel to determine the five-day test for biochemical oxygen demand.

Biochemical markers in the assessment of bone disease

NASA Technical Reports Server (NTRS)

Bikle, D. D.

1997-01-01

As the mean age of our population increases, increasing attention has been paid to the diseases associated with aging, including diseases of the skeleton such as osteoporosis. Effective means of treating and possibly preventing such skeletal disorders are emerging, making their early recognition an important goal for the primary care physician. Although bone density measurements and skeletal imaging studies remain of primary diagnostic importance in this regard, a large number of assays for biochemical markers of bone formation and resorption are being developed that promise to complement the densitometry measurements and imaging studies, providing an assessment of the rates of bone turnover and an earlier evaluation of the effects of therapy. In this review, emphasizing the recent literature, the major biochemical markers currently in use or under active investigation are described, and their application in a number of diseases of the skeleton including osteoporosis is evaluated.

A stopped-flow calorimeter for biochemical applications.

PubMed Central

Howarth, J V; Millar, N C; Gutfreund, H

1987-01-01

A rapid-response stopped-flow calorimeter for small samples of reagents is described. The construction, performance characteristics and operational limitations are described, along with an example of its ability to resolve the kinetics of an enzyme-catalysed hydrolysis. It is thought likely that the method would find useful application in a variety of chemical and biochemical investigations. PMID:3435478

MARCKS promotes invasion and is associated with biochemical recurrence in prostate cancer

PubMed Central

Dorris, Emma; O'Neill, Amanda; Hanrahan, Karen; Treacy, Ann; Watson, R. William

2017-01-01

Background Overtreatment of low-grade prostate cancer is a recognised problem for clinicians and patients. However, under-treatment runs the risk of missing the opportunity for cure in those who could benefit. Identification of new biomarkers of disease progression, including metastases, is required to better stratify and appropriately treat these patients. The ability to predict if prostate cancer will recur is an important clinical question that would impact treatment options for patients. Studies in other cancers have associated MARCKS with metastasis. Methods Tissue microarrays of local prostatectomy samples from a cohort of biochemical recurrent and non-biochemical recurrent tumours were assayed for MARCKS protein expression. Prostate cancer cell lines were transfected with siRNA targeting MARCKS or a control and functional endpoints of migration, invasion, proliferation, viability and apoptosis were measured. Actin was visualised by fluorescent microscopy and evidence of a cadherin switch and activation of the AKT pathway were assayed. Results MARCKS was upregulated in biochemical recurrent patients compared to non-biochemical recurrent. Knockdown of MARCKS reduced migration and invasion of prostate cancer cells, reduced MMP9 mRNA expression, as well as decreasing cell spreading and increased cell:cell adhesion in prostate cancer cell colonies. Knockdown of MARCKS had no effect on proliferation, viability or apoptosis of the prostate cancer cells. Conclusions In conclusion, MARCKS promotes migration and invasion and is associated with biochemical recurrence in localised prostate cancer tumours. The mechanisms by which this occurs have yet to be fully elucidated but lack of a cadherin switch indicates it is not via epithelial-to-mesenchymal transition. Actin rearrangement indicates that MARCKS promotes invasion through regulating the architecture of the cell. PMID:29069765

A Novel MiRNA-Based Predictive Model for Biochemical Failure Following Post-Prostatectomy Salvage Radiation Therapy

PubMed Central

Stegmaier, Petra; Drendel, Vanessa; Mo, Xiaokui; Ling, Stella; Fabian, Denise; Manring, Isabel; Jilg, Cordula A.; Schultze-Seemann, Wolfgang; McNulty, Maureen; Zynger, Debra L.; Martin, Douglas; White, Julia; Werner, Martin; Grosu, Anca L.; Chakravarti, Arnab

2015-01-01

Purpose To develop a microRNA (miRNA)-based predictive model for prostate cancer patients of 1) time to biochemical recurrence after radical prostatectomy and 2) biochemical recurrence after salvage radiation therapy following documented biochemical disease progression post-radical prostatectomy. Methods Forty three patients who had undergone salvage radiation therapy following biochemical failure after radical prostatectomy with greater than 4 years of follow-up data were identified. Formalin-fixed, paraffin-embedded tissue blocks were collected for all patients and total RNA was isolated from 1mm cores enriched for tumor (>70%). Eight hundred miRNAs were analyzed simultaneously using the nCounter human miRNA v2 assay (NanoString Technologies; Seattle, WA). Univariate and multivariate Cox proportion hazards regression models as well as receiver operating characteristics were used to identify statistically significant miRNAs that were predictive of biochemical recurrence. Results Eighty eight miRNAs were identified to be significantly (p<0.05) associated with biochemical failure post-prostatectomy by multivariate analysis and clustered into two groups that correlated with early (≤ 36 months) versus late recurrence (>36 months). Nine miRNAs were identified to be significantly (p<0.05) associated by multivariate analysis with biochemical failure after salvage radiation therapy. A new predictive model for biochemical recurrence after salvage radiation therapy was developed; this model consisted of miR-4516 and miR-601 together with, Gleason score, and lymph node status. The area under the ROC curve (AUC) was improved to 0.83 compared to that of 0.66 for Gleason score and lymph node status alone. Conclusion miRNA signatures can distinguish patients who fail soon after radical prostatectomy versus late failures, giving insight into which patients may need adjuvant therapy. Notably, two novel miRNAs (miR-4516 and miR-601) were identified that significantly improve

Recursively constructing analytic expressions for equilibrium distributions of stochastic biochemical reaction networks

PubMed Central

Baetica, Ania-Ariadna; Singhal, Vipul; Murray, Richard M.

2017-01-01

Noise is often indispensable to key cellular activities, such as gene expression, necessitating the use of stochastic models to capture its dynamics. The chemical master equation (CME) is a commonly used stochastic model of Kolmogorov forward equations that describe how the probability distribution of a chemically reacting system varies with time. Finding analytic solutions to the CME can have benefits, such as expediting simulations of multiscale biochemical reaction networks and aiding the design of distributional responses. However, analytic solutions are rarely known. A recent method of computing analytic stationary solutions relies on gluing simple state spaces together recursively at one or two states. We explore the capabilities of this method and introduce algorithms to derive analytic stationary solutions to the CME. We first formally characterize state spaces that can be constructed by performing single-state gluing of paths, cycles or both sequentially. We then study stochastic biochemical reaction networks that consist of reversible, elementary reactions with two-dimensional state spaces. We also discuss extending the method to infinite state spaces and designing the stationary behaviour of stochastic biochemical reaction networks. Finally, we illustrate the aforementioned ideas using examples that include two interconnected transcriptional components and biochemical reactions with two-dimensional state spaces. PMID:28566513

Recursively constructing analytic expressions for equilibrium distributions of stochastic biochemical reaction networks.

PubMed

Meng, X Flora; Baetica, Ania-Ariadna; Singhal, Vipul; Murray, Richard M

2017-05-01

Noise is often indispensable to key cellular activities, such as gene expression, necessitating the use of stochastic models to capture its dynamics. The chemical master equation (CME) is a commonly used stochastic model of Kolmogorov forward equations that describe how the probability distribution of a chemically reacting system varies with time. Finding analytic solutions to the CME can have benefits, such as expediting simulations of multiscale biochemical reaction networks and aiding the design of distributional responses. However, analytic solutions are rarely known. A recent method of computing analytic stationary solutions relies on gluing simple state spaces together recursively at one or two states. We explore the capabilities of this method and introduce algorithms to derive analytic stationary solutions to the CME. We first formally characterize state spaces that can be constructed by performing single-state gluing of paths, cycles or both sequentially. We then study stochastic biochemical reaction networks that consist of reversible, elementary reactions with two-dimensional state spaces. We also discuss extending the method to infinite state spaces and designing the stationary behaviour of stochastic biochemical reaction networks. Finally, we illustrate the aforementioned ideas using examples that include two interconnected transcriptional components and biochemical reactions with two-dimensional state spaces. © 2017 The Author(s).

Connecting Biochemical Photosynthesis Models with Crop Models to Support Crop Improvement

PubMed Central

Wu, Alex; Song, Youhong; van Oosterom, Erik J.; Hammer, Graeme L.

2016-01-01

The next advance in field crop productivity will likely need to come from improving crop use efficiency of resources (e.g., light, water, and nitrogen), aspects of which are closely linked with overall crop photosynthetic efficiency. Progress in genetic manipulation of photosynthesis is confounded by uncertainties of consequences at crop level because of difficulties connecting across scales. Crop growth and development simulation models that integrate across biological levels of organization and use a gene-to-phenotype modeling approach may present a way forward. There has been a long history of development of crop models capable of simulating dynamics of crop physiological attributes. Many crop models incorporate canopy photosynthesis (source) as a key driver for crop growth, while others derive crop growth from the balance between source- and sink-limitations. Modeling leaf photosynthesis has progressed from empirical modeling via light response curves to a more mechanistic basis, having clearer links to the underlying biochemical processes of photosynthesis. Cross-scale modeling that connects models at the biochemical and crop levels and utilizes developments in upscaling leaf-level models to canopy models has the potential to bridge the gap between photosynthetic manipulation at the biochemical level and its consequences on crop productivity. Here we review approaches to this emerging cross-scale modeling framework and reinforce the need for connections across levels of modeling. Further, we propose strategies for connecting biochemical models of photosynthesis into the cross-scale modeling framework to support crop improvement through photosynthetic manipulation. PMID:27790232

Diagnostic value of the biochemical tests in patients with purulent pericarditis.

PubMed

Ekim, Meral; Ekim, Hasan

2014-07-01

Purulent pericarditis is a collection of purulent effusion in the pericardial space. It has become a rare entity with the increased availability and use of antibiotics. In contrast to pleural empyema, there are few data regarding the biochemical parameters of purulent pericardial effusion to aid diagnosis. Therefore, in this study, we have evaluated the diagnostic utility of biochemical tests in patients with purulent pericarditis. Between September 2004 and September 2012, we treated fifteen children with purulent pericarditis and tamponade. There were 8 boys and 7 girls, ranging in age from 8 months to 14 years, with a mean age of 5.3 ± 3.2 years. Echocardiographic diagnosis of cardiac tamponade was made in all patients. All patients underwent immediate surgical drainage due to cardiac tamponade. The diagnosis of purulent pericarditis was supported by biochemical tests. Anterior mini-thoracotomy or subxiphoid approach was performed for surgical drainage. The most common clinical findings were tamponade, hepatomegaly, tachycardia, fever refractory antibiotic therapy, dyspnea, tachypnea, cough, and increased jugular venous pressure. Central venous pressure decreased and arterial tension increased immediately after the evacuation of purulent effusion during operation in all patients. The pericardial effusion had high lactic dehydrogenase, and low glucose concentration, confirming purulent pericarditis. Also, pH (mean± SD) was 7.01 ± 0.06. The culture of pericardial effusions and blood samples were negative. Biochemical tests are useful guideline when assessing the pericardial effusions. However, these tests should be interpreted with the clinical and operative findings.

Connecting Biochemical Photosynthesis Models with Crop Models to Support Crop Improvement.

PubMed

Wu, Alex; Song, Youhong; van Oosterom, Erik J; Hammer, Graeme L

2016-01-01

The next advance in field crop productivity will likely need to come from improving crop use efficiency of resources (e.g., light, water, and nitrogen), aspects of which are closely linked with overall crop photosynthetic efficiency. Progress in genetic manipulation of photosynthesis is confounded by uncertainties of consequences at crop level because of difficulties connecting across scales. Crop growth and development simulation models that integrate across biological levels of organization and use a gene-to-phenotype modeling approach may present a way forward. There has been a long history of development of crop models capable of simulating dynamics of crop physiological attributes. Many crop models incorporate canopy photosynthesis (source) as a key driver for crop growth, while others derive crop growth from the balance between source- and sink-limitations. Modeling leaf photosynthesis has progressed from empirical modeling via light response curves to a more mechanistic basis, having clearer links to the underlying biochemical processes of photosynthesis. Cross-scale modeling that connects models at the biochemical and crop levels and utilizes developments in upscaling leaf-level models to canopy models has the potential to bridge the gap between photosynthetic manipulation at the biochemical level and its consequences on crop productivity. Here we review approaches to this emerging cross-scale modeling framework and reinforce the need for connections across levels of modeling. Further, we propose strategies for connecting biochemical models of photosynthesis into the cross-scale modeling framework to support crop improvement through photosynthetic manipulation.

Surrogate biochemical markers: precise measurement for strategic drug and biologics development.

PubMed

Lee, J W; Hulse, J D; Colburn, W A

1995-05-01

More efficient drug and biologics development is necessary for future success of pharmaceutical and biotechnology companies. One way to achieve this objective is to use rationally selected surrogate markers to improve the early decision-making process. Using typical clinical chemistry methods to measure biochemical markers may not ensure adequate precision and reproducibility. In contrast, using analytical methods that meet good laboratory practices along with rational selection and validation of biochemical markers can give those who use them a competitive advantage over those who do not by providing meaningful data for earlier decision making.

Temperature-programmed natural convection for micromixing and biochemical reaction in a single microfluidic chamber.

PubMed

Kim, Sung-Jin; Wang, Fang; Burns, Mark A; Kurabayashi, Katsuo

2009-06-01

Micromixing is a crucial step for biochemical reactions in microfluidic networks. A critical challenge is that the system containing micromixers needs numerous pumps, chambers, and channels not only for the micromixing but also for the biochemical reactions and detections. Thus, a simple and compatible design of the micromixer element for the system is essential. Here, we propose a simple, yet effective, scheme that enables micromixing and a biochemical reaction in a single microfluidic chamber without using any pumps. We accomplish this process by using natural convection in conjunction with alternating heating of two heaters for efficient micromixing, and by regulating capillarity for sample transport. As a model application, we demonstrate micromixing and subsequent polymerase chain reaction (PCR) for an influenza viral DNA fragment. This process is achieved in a platform of a microfluidic cartridge and a microfabricated heating-instrument with a fast thermal response. Our results will significantly simplify micromixing and a subsequent biochemical reaction that involves reagent heating in microfluidic networks.

A Geometric Method for Model Reduction of Biochemical Networks with Polynomial Rate Functions.

PubMed

Samal, Satya Swarup; Grigoriev, Dima; Fröhlich, Holger; Weber, Andreas; Radulescu, Ovidiu

2015-12-01

Model reduction of biochemical networks relies on the knowledge of slow and fast variables. We provide a geometric method, based on the Newton polytope, to identify slow variables of a biochemical network with polynomial rate functions. The gist of the method is the notion of tropical equilibration that provides approximate descriptions of slow invariant manifolds. Compared to extant numerical algorithms such as the intrinsic low-dimensional manifold method, our approach is symbolic and utilizes orders of magnitude instead of precise values of the model parameters. Application of this method to a large collection of biochemical network models supports the idea that the number of dynamical variables in minimal models of cell physiology can be small, in spite of the large number of molecular regulatory actors.

Biochemical diagnosis of phaeochromocytoma: two instructive case reports.

PubMed Central

Stewart, M F; Reed, P; Weinkove, C; Moriarty, K J; Ralston, A J

1993-01-01

The biochemical features of two patients with phaeochromocytomas illustrate the inadvisability of depending on a single group of analytes for the diagnosis. The first case presented as a surgical emergency with retroperitoneal haemorrhage. Biochemical diagnosis was difficult since total 24 hour urinary free catecholamine excretion was within normal limits in two out of three samples, and only marginally raised in the third with an atypical preponderance of adrenaline. Plasma catecholamine concentrations were also normal. But urinary excretion of the catecholamine metabolites, metadrenaline and 4-hydroxy-3-methoxy mandelic acid (HMMA), was consistently raised. In contrast, the second patient presenting with headache and labile hypertension showed normal metabolite excretion in the face of grossly increased free noradrenaline excretion and raised plasma noradrenaline concentrations. It is therefore recommend that, as well as urinary free catecholamines, one group of their main metabolites, the 3-methoxy amines (normetadrenaline and metadrenaline) or HMMA, should routinely be measured whenever a phaeochromocytoma is suspected. PMID:8463426

A Biochemical Oscillator Using Excitatory Molecules for Nanonetworks.

PubMed

Shitiri, Ethungshan; Cho, Ho-Shin

2016-10-01

For nanonetworks to be able to achieve large-scale functionality, such as to respond collectively to a trigger, synchrony between nanomachines is essential. However, to facilitate synchronization, some sort of physical clocking mechanism is required, such as the oscillators driven by auto-inhibitory molecules or by auto-inducing molecules. In this study, taking inspiration from the widely studied biological oscillatory phenomena called Calcium (Ca 2+ ) oscillations, we undertake a different approach to design an oscillator. Our model employs three different types of excitatory molecules that work in tandem to generate oscillatory phenomenon in the concentration levels of the molecule of interest. The main objective of the study is to model a high frequency biochemical oscillator, along with the investigations to identify and determine the parameters that affect the period of the oscillations. The investigations entail and highlight the design of the reserve unit, a reservoir of the molecule of interest, as a key factor in realizing a high frequency stable biochemical oscillator.

Biochemically Silent Abdominal Paragangliomas in Patients with Mutations in the Succinate Dehydrogenase Subunit B Gene

PubMed Central

Timmers, Henri J. L. M.; Pacak, Karel; Huynh, Thanh T.; Abu-Asab, Mones; Tsokos, Maria; Merino, Maria J.; Baysal, Bora E.; Adams, Karen T.; Eisenhofer, Graeme

2008-01-01

Context: Patients with adrenal and extra-adrenal abdominal paraganglioma (PGL) almost invariably have increased plasma and urine concentrations of metanephrines, the O-methylated metabolites of catecholamines. We report four cases of biochemically silent abdominal PGL, in which metanephrines were normal despite extensive disease. Objective: Our objective was to identify the mechanism underlying the lack of catecholamine hypersecretion and metabolism to metanephrines in biochemically silent PGL. Design: This is a descriptive study. Setting: The study was performed at a referral center. Patients: One index case and three additional patients with large abdominal PGL and metastases but with the lack of evidence of catecholamine production, six patients with metastatic catecholamine-producing PGL and a mutation of the succinate dehydrogenase subunit B (SDHB) gene, and 136 random patients with catecholamine-producing PGL were included in the study. Main Outcome Measures: Plasma, urine, and tumor tissue concentrations of catecholamines and metabolites were calculated with electron microscopy and tyrosine hydroxylase immunohistochemistry. Results: All four patients with biochemically silent PGL had an underlying SDHB mutation. In the index case, the tumor tissue concentration of catecholamines (1.8 nmol/g) was less than 0.01% that of the median (20,410 nmol/g) for the 136 patients with catecholamine-producing tumors. Electron microscopy showed the presence of normal secretory granules in all four biochemically silent PGLs. Tyrosine hydroxylase immunoreactivity was negligible in the four biochemically silent PGLs but abundant in catecholamine-producing PGLs. Conclusions: Patients with SDHB mutations may present with biochemically silent abdominal PGLs due to defective catecholamine synthesis resulting from the absence of tyrosine hydroxylase. Screening for tumors in patients with SDHB mutations should not be limited to biochemical tests of catecholamine excess. PMID

Influence of Biochemical Features of Burkholderia pseudomallei Strains on Identification Reliability by Vitek 2 System.

PubMed

Zakharova, Irina B; Lopasteyskaya, Yana A; Toporkov, Andrey V; Viktorov, Dmitry V

2018-01-01

Burkholderia pseudomallei is a Gram-negative saprophytic soil bacterium that causes melioidosis, a potentially fatal disease endemic in wet tropical areas. The currently available biochemical identification systems can misidentify some strains of B. pseudomallei . The aim of the present study was to identify the biochemical features of B. pseudomallei , which can affect its correct identification by Vitek 2 system. The biochemical patterns of 40 B. pseudomallei strains were obtained using Vitek 2 GN cards. The average contribution of biochemical tests in overall dissimilarities between correctly and incorrectly identified strains was assessed using nonmetric multidimensional scaling. It was found ( R statistic of 0.836, P = 0.001) that a combination of negative N-acetyl galactosaminidase, β-N-acetyl glucosaminidase, phosphatase, and positive D-cellobiase (dCEL), tyrosine arylamidase (TyrA), and L-proline arylamidase (ProA) tests leads to low discrimination of B. pseudomallei , whereas a set of positive dCEL and negative N-acetyl galactosaminidase, TyrA, and ProA determines the wrong identification of B. pseudomallei as Burkholderia cepacia complex. The further expansion of the Vitek 2 identification keys is needed for correct identification of atypical or regionally distributed biochemical profiles of B. pseudomallei .

Biochemical and cytochemical evaluation of heterozygote individuals with glucose-6-phosphate dehydrogenase deficiency.

PubMed

Gurbuz, Nilgun; Aksu, Tevfik Aslan; Van Noorden, Cornelis J F

2005-01-01

The aim of this study was to diagnose heterozygous glucose-6-phosphate dehydrogenase (G6PD) deficient females by an inexpensive cytochemical G6PD staining method that is easy to perform, allowing diagnosis of G6PD deficiency without cumbersome genetic analysis. Three subject groups were included in the study. The first group consisted of 15 hemizygous deficient males. The second and the third group were composed of 15 heterozygous deficient females and 15 healthy individuals, respectively. Biochemical determination and cytochemical staining of G6PD activity were performed in samples of all subjects. Results obtained with the cytochemical staining method correlated significantly with the biochemical data (p < 0.001), but a only 51-68% of the erythrocytes were stained positively in females with normal biochemical G6PD activity despite their having a G6PD-deficient child. This observation clearly indicates that these individuals are heterozygously deficient. These findings show that the cytochemical staining method to detect G6PD activity in erythrocytes is reliable, sensitive and specific and is superior to the biochemical method. Therefore, this method can be used routinely to detect heterozygous G6PD deficiency.

A biochemical method for assessing the neurotoxic effects of misonidazole in the rat.

PubMed Central

Rose, G. P.; Dewar, A. J.; Stratford, I. J.

1980-01-01

A proven biochemical method for assessing chemically induced neurotoxicity has been applied to the study of the toxic effects of misonidazole (MISO) in the rat. This involves the fluorimetric measurement of beta-glucuronidase and beta-galactosidase activities in homogenates of rat nervous tissue. The tissues analysed were sciatic/posterior tibial nerve (SPTN) cut into 4 sections, trigeminal ganglia and cerebellum. MISO administered i.p. to Wistar rats in doses greater than 300 mg/kg/day for 7 consecutive days produced maximal increases in both beta-glucuronidase and beta-galactosidase activities in th SPTN at 4 weeks (140-180% of control values). The highest increases were associated with the most distal secretion of the nerve. Significant enzyme-activity changes were also found in the trigeminal ganglia and cerebellum of MISO-dosed rats. The greatest activity occurred 4-5 weeks after dosing, and was dose-related. It is concluded that, in the rat, MISO can produce biochemical changes consistent with a dying-back peripheral neuropathy, and biochemical changes suggestive of cerebellar damage. This biochemical approach would appear to offer a convenient quantitative method for the detection of neurotoxic effects of other potential radio-sensitizing drugs. PMID:7459223

A comparative biochemical profile of some cyprinids fish in Dukan Lake, Kurdistan-Iraq

NASA Astrophysics Data System (ADS)

Azeez, Darya Mohammed; Mohammed, Sarbaz Ibrahim

2017-09-01

The present study was carried out to demonstrate the baseline values for some serum biochemical parameters for 64 adult freshwater fish including seven species belong to family Cyprinidae, have been collected in Dukan Lake, Kurdistan region-Iraq. Fishes were weighed, measured, and collect blood for blood chemistry. Serum biochemical analyses were determined using (Cobas C 311) full automatic chemical analyzer. The result of comparative study of serum biochemical parameters of all Cyprinidae species showed that serum glucose was (459.10±106.99 mg/dl) and direct bilirubin was (0.056±0.021mg/dl) in Barbus grypus, serum total protein (3.511± 0.0484gm/dl) and HDL (133.11±0.4231mg/dl) in Cyprinus carpio, serum cholesterol (338.33±43.923 mg/dl) and LDL (86.11±11.871mg/dl) in Carassius carassius, serum triglyceride (420.0±28.8mg/dl) and ALK (113.93±20.65U/L) in Chondrostoma regium, serum AST and serum ALT in Capoeta trutta, were significantly higher when compared to other species. In a conclusion there is variation in biochemical values among species of same family.

Biochemical and Biophysical Cues in Matrix Design for Chronic and Diabetic Wound Treatment

PubMed Central

Xiao, Yun; Ahadian, Samad

2017-01-01

Progress in biomaterial science and engineering and increasing knowledge in cell biology have enabled us to develop functional biomaterials providing appropriate biochemical and biophysical cues for tissue regeneration applications. Tissue regeneration is particularly important to treat chronic wounds of people with diabetes. Understanding and controlling the cellular microenvironment of the wound tissue are important to improve the wound healing process. In this study, we review different biochemical (e.g., growth factors, peptides, DNA, and RNA) and biophysical (e.g., topographical guidance, pressure, electrical stimulation, and pulsed electromagnetic field) cues providing a functional and instructive acellular matrix to heal diabetic chronic wounds. The biochemical and biophysical signals generally regulate cell–matrix interactions and cell behavior and function inducing the tissue regeneration for chronic wounds. Some technologies and devices have already been developed and used in the clinic employing biochemical and biophysical cues for wound healing applications. These technologies can be integrated with smart biomaterials to deliver therapeutic agents to the wound tissue in a precise and controllable manner. This review provides useful guidance in understanding molecular mechanisms and signals in the healing of diabetic chronic wounds and in designing instructive biomaterials to treat them. PMID:27405960

Scientific issues and potential remote-sensing requirements for plant biochemical content

NASA Technical Reports Server (NTRS)

Peterson, David L.; Hubbard, G. S.

1992-01-01

Application of developments in imaging spectrometry to the study of terrestrial ecosystems, which began in 1983, demonstrate the potential to estimate lignin and nitrogen concentrations of plant canopies by remote-sensing techniques. Estimation of these parameters from the first principles of radiative transfer and the interactions of light with plant materials is not presently possible, principally because of lack of knowledge about internal leaf scattering and specific absorption involving biochemical compounds. From the perspective of remote-sensing instrumentation, sensors are needed to support derivative imaging spectroscopy. Biochemical absorption features tend to occur in functional groupings throughout the 1100- to 2500-nm region. Derivative spectroscopy improves the information associated with the weaker, narrower absorption features of biochemical absorption that are superimposed on the strong absolute variations due to foliar biomass, pigments, and leaf water content of plant canopies. Preliminary sensor specifications call for 8-nm bandwidths at 2-nm centers in four spectral regions (about 400 bands total) and a signal-to-noise performance of at least 1000:1 for 20 percent albedo targets in the 2000-nm region.

ESTIMATING GASEOUS EXCHANGES BETWEEN THE ATMOSPHERE AND PLANTS USING A COUPLED BIOCHEMICAL DRY DEPOSITION MODEL

EPA Science Inventory

To study gaseous exchanges between the soil, biosphere and atmosphere, a biochemical model was coupled with the latest version of Meyers Multi-Layer Deposition Model. The biochemical model describes photosynthesis and respiration and their coupling with stomatal resistance for...

Complexity reduction of biochemical rate expressions.

PubMed

Schmidt, Henning; Madsen, Mads F; Danø, Sune; Cedersund, Gunnar

2008-03-15

The current trend in dynamical modelling of biochemical systems is to construct more and more mechanistically detailed and thus complex models. The complexity is reflected in the number of dynamic state variables and parameters, as well as in the complexity of the kinetic rate expressions. However, a greater level of complexity, or level of detail, does not necessarily imply better models, or a better understanding of the underlying processes. Data often does not contain enough information to discriminate between different model hypotheses, and such overparameterization makes it hard to establish the validity of the various parts of the model. Consequently, there is an increasing demand for model reduction methods. We present a new reduction method that reduces complex rational rate expressions, such as those often used to describe enzymatic reactions. The method is a novel term-based identifiability analysis, which is easy to use and allows for user-specified reductions of individual rate expressions in complete models. The method is one of the first methods to meet the classical engineering objective of improved parameter identifiability without losing the systems biology demand of preserved biochemical interpretation. The method has been implemented in the Systems Biology Toolbox 2 for MATLAB, which is freely available from http://www.sbtoolbox2.org. The Supplementary Material contains scripts that show how to use it by applying the method to the example models, discussed in this article.

Mathematical treatment of isotopologue and isotopomer speciation and fractionation in biochemical kinetics

DOE Office of Scientific and Technical Information (OSTI.GOV)

Maggi, F.M.; Riley, W.J.

2009-11-01

We present a mathematical treatment of the kinetic equations that describe isotopologue and isotopomer speciation and fractionation during enzyme-catalyzed biochemical reactions. These equations, presented here with the name GEBIK (general equations for biochemical isotope kinetics) and GEBIF (general equations for biochemical isotope fractionation), take into account microbial biomass and enzyme dynamics, reaction stoichiometry, isotope substitution number, and isotope location within each isotopologue and isotopomer. In addition to solving the complete GEBIK and GEBIF, we also present and discuss two approximations to the full solutions under the assumption of biomass-free and enzyme steady-state, and under the quasi-steady-state assumption as applied tomore » the complexation rate. The complete and approximate approaches are applied to observations of biological denitrification in soils. Our analysis highlights that the full GEBIK and GEBIF provide a more accurate description of concentrations and isotopic compositions of substrates and products throughout the reaction than do the approximate forms. We demonstrate that the isotopic effects of a biochemical reaction depend, in the most general case, on substrate and complex concentrations and, therefore, the fractionation factor is a function of time. We also demonstrate that inverse isotopic effects can occur for values of the fractionation factor smaller than 1, and that reactions that do not discriminate isotopes do not necessarily imply a fractionation factor equal to 1.« less

BioNetCAD: design, simulation and experimental validation of synthetic biochemical networks

PubMed Central

Rialle, Stéphanie; Felicori, Liza; Dias-Lopes, Camila; Pérès, Sabine; El Atia, Sanaâ; Thierry, Alain R.; Amar, Patrick; Molina, Franck

2010-01-01

Motivation: Synthetic biology studies how to design and construct biological systems with functions that do not exist in nature. Biochemical networks, although easier to control, have been used less frequently than genetic networks as a base to build a synthetic system. To date, no clear engineering principles exist to design such cell-free biochemical networks. Results: We describe a methodology for the construction of synthetic biochemical networks based on three main steps: design, simulation and experimental validation. We developed BioNetCAD to help users to go through these steps. BioNetCAD allows designing abstract networks that can be implemented thanks to CompuBioTicDB, a database of parts for synthetic biology. BioNetCAD enables also simulations with the HSim software and the classical Ordinary Differential Equations (ODE). We demonstrate with a case study that BioNetCAD can rationalize and reduce further experimental validation during the construction of a biochemical network. Availability and implementation: BioNetCAD is freely available at http://www.sysdiag.cnrs.fr/BioNetCAD. It is implemented in Java and supported on MS Windows. CompuBioTicDB is freely accessible at http://compubiotic.sysdiag.cnrs.fr/ Contact: stephanie.rialle@sysdiag.cnrs.fr; franck.molina@sysdiag.cnrs.fr Supplementary information: Supplementary data are available at Bioinformatics online. PMID:20628073

Biochemical and Structural Characterisation of DNA Ligases from Bacteria and Archaea.

PubMed

Pergolizzi, Giulia; Wagner, Gerd K; Bowater, Richard Peter

2016-08-31

DNA ligases are enzymes that seal breaks in the backbones of DNA, leading to them being essential for the survival of all organisms. DNA ligases have been studied from many different types of cells and organisms and shown to have diverse sizes and sequences, with well conserved specific sequences that are required for enzymatic activity. A significant number of DNA ligases have been isolated or prepared in recombinant forms and, here, we review their biochemical and structural characterisation. All DNA ligases contain an essential lysine that transfers an adenylate group from a co-factor to the 5'-phosphate of the DNA end that will ultimately be joined to the 3'-hydroxyl of the neighbouring DNA strand. The essential DNA ligases in bacteria use nicotinamide adenine dinucleotide ( β -NAD + ) as their co-factor whereas those that are essential in other cells use adenosine-5'-triphosphate (ATP) as their co-factor. This observation suggests that the essential bacterial enzyme could be targeted by novel antibiotics and the complex molecular structure of β -NAD + affords multiple opportunities for chemical modification. Several recent studies have synthesised novel derivatives and their biological activity against a range of DNA ligases has been evaluated as inhibitors for drug discovery and/or non-natural substrates for biochemical applications. Here, we review the recent advances that herald new opportunities to alter the biochemical activities of these important enzymes. The recent development of modified derivatives of nucleotides highlights that the continued combination of structural, biochemical and biophysical techniques will be useful in targeting these essential cellular enzymes. ©2016 The Author(s).

Diagnostic value of the biochemical tests in patients with purulent pericarditis

PubMed Central

Ekim, Meral; Ekim, Hasan

2014-01-01

Objectives: Purulent pericarditis is a collection of purulent effusion in the pericardial space. It has become a rare entity with the increased availability and use of antibiotics. In contrast to pleural empyema, there are few data regarding the biochemical parameters of purulent pericardial effusion to aid diagnosis. Therefore, in this study, we have evaluated the diagnostic utility of biochemical tests in patients with purulent pericarditis. Methods: Between September 2004 and September 2012, we treated fifteen children with purulent pericarditis and tamponade. There were 8 boys and 7 girls, ranging in age from 8 months to 14 years, with a mean age of 5.3 ± 3.2 years. Echocardiographic diagnosis of cardiac tamponade was made in all patients. All patients underwent immediate surgical drainage due to cardiac tamponade. The diagnosis of purulent pericarditis was supported by biochemical tests. Anterior mini-thoracotomy or subxiphoid approach was performed for surgical drainage. Results: The most common clinical findings were tamponade, hepatomegaly, tachycardia, fever refractory antibiotic therapy, dyspnea, tachypnea, cough, and increased jugular venous pressure. Central venous pressure decreased and arterial tension increased immediately after the evacuation of purulent effusion during operation in all patients. The pericardial effusion had high lactic dehydrogenase, and low glucose concentration, confirming purulent pericarditis. Also, pH (mean± SD) was 7.01 ± 0.06. The culture of pericardial effusions and blood samples were negative. Conclusion: Biochemical tests are useful guideline when assessing the pericardial effusions. However, these tests should be interpreted with the clinical and operative findings. PMID:25097529

Scaling up semi-arid grassland biochemical content from the leaf to the canopy level: challenges and opportunities.

PubMed

He, Yuhong; Mui, Amy

2010-01-01

Remote sensing imagery is being used intensively to estimate the biochemical content of vegetation (e.g., chlorophyll, nitrogen, and lignin) at the leaf level. As a result of our need for vegetation biochemical information and our increasing ability to obtain canopy spectral data, a few techniques have been explored to scale leaf-level biochemical content to the canopy level for forests and crops. However, due to the contribution of non-green materials (i.e., standing dead litter, rock, and bare soil) from canopy spectra in semi-arid grasslands, it is difficult to obtain information about grassland biochemical content from remote sensing data at the canopy level. This paper summarizes available methods used to scale biochemical information from the leaf level to the canopy level and groups these methods into three categories: direct extrapolation, canopy-integrated approach, and inversion of physical models. As for semi-arid heterogeneous grasslands, we conclude that all methods are useful, but none are ideal. It is recommended that future research should explore a systematic upscaling framework which combines spatial pattern analysis, canopy-integrated approach, and modeling methods to retrieve vegetation biochemical content at the canopy level.

Biochemical identification of the mallard, Anas platyrhynchos, and black duck, A. rubripes

USGS Publications Warehouse

Morgan, R.P.; Noe, L.A.; Henny, C.J.

1976-01-01

1. Eleven tissue systems from mallards and black ducks were examined for soluble proteins, lactate dehydrogenases and non-specific esterases through discontinuous polyacrylamide techniques.2. Biochemical relationships between the black duck and mallard are extremely similar.3. Hemoglobins and lactate dehydrogenase appear to be common in electrophoretic mobility between the two species.4. Approximately 89% of the soluble proteins and 58% of the non-specific esterases are common among the two species, indicating both biochemical similarity at the genus level and species-specificity.

[Some biochemical parameters in the placenta in discoordinated and powerless labors].

PubMed

Sitnikova, O G; Peretiatko, L P; Sharygin, S A; Kuz'menko, G N; Popova, I G

2009-11-01

A number of biochemical parameters (total nitrites and nitrates (NO(x)), cyclic guanosine monophosphate (cGMP), nitrotyrosine, medium-weight molecules (MCM) in the placenta were determined in women with gestosis during discoordinated and powerless labor. Thirty placentas (10 placentas from parturients after discoordinated labor, 10 from those after powerless labor, 10 placentas as a control group) were examined. Changes in the parameters under study were found to result in the development of nitroxide and oxidant stresses and endotoxicosis. The biochemical parameters should be considered as placental criteria for the differential diagnosis of labor anomalies in gestosis, such as powerless and discoordinated labors.

Biochemical functionalization of peptide nanotubes with phage displayed peptides

NASA Astrophysics Data System (ADS)

Swaminathan, Swathi; Cui, Yue

2016-09-01

The development of a general approach for the biochemical functionalization of peptide nanotubes (PNTs) could open up existing opportunities in both fundamental studies as well as a variety of applications. PNTs are spontaneously assembled organic nanostructures made from peptides. Phage display has emerged as a powerful approach for identifying selective peptide binding motifs. Here, we demonstrate for the first time the biochemical functionalization of PNTs via peptides identified from a phage display peptide library. The phage-displayed peptides are shown to recognize PNTs. These advances further allow for the development of bifunctional peptides for the capture of bacteria and the self-assembly of silver particles onto PNTs. We anticipate that these results could provide significant opportunities for using PNTs in both fundamental studies and practical applications, including sensors and biosensors nanoelectronics, energy storage devices, drug delivery, and tissue engineering.

Sonographic assessment of petroleum-induced hepatotoxicity in Nigerians: does biochemical assessment underestimate liver damage?

PubMed

Anakwue, Angel-Mary; Anakwue, Raphael; Okeji, Mark; Idigo, Felicitas; Agwu, Kenneth; Nwogu, Uloma

2017-03-01

Exposure to petroleum products has been shown to have significant adverse effects on the liver which can manifest either as morphological or physiological changes. The aim of the study was to assess the effects of chronic exposure to some petroleum products on the liver of exposed workers using sonography and to determine whether biochemical assessments underestimated hepatotoxicity. Abdominal ultrasound was performed on 415 exposed workers in order to evaluate liver echogenicity and size. Also, biochemical assessment of the liver was done to evaluate its function. Statistically significant increase in the liver parenchymal echogenicity and the liver size was seen in the exposed workers compared with control (p ≤ 0.05). These increased as the exposure duration increased. It was also noted that out of 16.87% (N=70) exposed workers with abnormal liver echopattern, only 2.65% (N=11) had alanine aminotransferase above the reference range. The study revealed evidence of ultrasound detectable hepatotoxicity among the exposed subjects. Sonography appeared to detect petroleum products-induced hepatic toxicity more than biochemical assays suggesting that biochemical assessment may have underestimated toxicity.

Linking stable isotopes and biochemical responses in Balanus glandula under sewage influence.

PubMed

Laitano, M V; Díaz-Jaramillo, M; Rodriguez, Y E; Ducós, E; Panarello, H O; Fernández-Gimenez, A V

2018-02-01

In the present study, we analyzed the influence of untreated sewage exposure on carbon (δ13C) and nitrogen (δ15N) isotopic composition and several biochemical responses in the barnacle Balanus glandula. The main objective was to evaluate whether changes in stable isotopes signature do reflect biochemical sub-lethal effects in a sewage influence gradient. Stable isotopes analysis showed differences in isotope signatures between close sewage influence and distant sites, being δ13C signatures stronger than that of δ15N. Regarding biochemical effects, although organisms close to the effluent would be clearly exposed to contaminants (increased GST activity) the oxidative stress would not be too evident (peroxidases and ACAP not affected). The most affected physiological aspect was the digestive one, reflected in increased alkaline proteases and lipases activities. A clear relation between δ15N and GST activity was found, showing to δ15N as an indicator of potential exposure to chemical contaminants.

Non-equilibrium phase transition in mesoscopic biochemical systems: from stochastic to nonlinear dynamics and beyond

PubMed Central

Ge, Hao; Qian, Hong

2011-01-01

A theory for an non-equilibrium phase transition in a driven biochemical network is presented. The theory is based on the chemical master equation (CME) formulation of mesoscopic biochemical reactions and the mathematical method of large deviations. The large deviations theory provides an analytical tool connecting the macroscopic multi-stability of an open chemical system with the multi-scale dynamics of its mesoscopic counterpart. It shows a corresponding non-equilibrium phase transition among multiple stochastic attractors. As an example, in the canonical phosphorylation–dephosphorylation system with feedback that exhibits bistability, we show that the non-equilibrium steady-state (NESS) phase transition has all the characteristics of classic equilibrium phase transition: Maxwell construction, a discontinuous first-derivative of the ‘free energy function’, Lee–Yang's zero for a generating function and a critical point that matches the cusp in nonlinear bifurcation theory. To the biochemical system, the mathematical analysis suggests three distinct timescales and needed levels of description. They are (i) molecular signalling, (ii) biochemical network nonlinear dynamics, and (iii) cellular evolution. For finite mesoscopic systems such as a cell, motions associated with (i) and (iii) are stochastic while that with (ii) is deterministic. Both (ii) and (iii) are emergent properties of a dynamic biochemical network. PMID:20466813

Influence of Biochemical Features of Burkholderia pseudomallei Strains on Identification Reliability by Vitek 2 System

PubMed Central

Zakharova, Irina B; Lopasteyskaya, Yana A; Toporkov, Andrey V; Viktorov, Dmitry V

2018-01-01

Background: Burkholderia pseudomallei is a Gram-negative saprophytic soil bacterium that causes melioidosis, a potentially fatal disease endemic in wet tropical areas. The currently available biochemical identification systems can misidentify some strains of B. pseudomallei. The aim of the present study was to identify the biochemical features of B. pseudomallei, which can affect its correct identification by Vitek 2 system. Materials and Methods: The biochemical patterns of 40 B. pseudomallei strains were obtained using Vitek 2 GN cards. The average contribution of biochemical tests in overall dissimilarities between correctly and incorrectly identified strains was assessed using nonmetric multidimensional scaling. Results: It was found (R statistic of 0.836, P = 0.001) that a combination of negative N-acetyl galactosaminidase, β-N-acetyl glucosaminidase, phosphatase, and positive D-cellobiase (dCEL), tyrosine arylamidase (TyrA), and L-proline arylamidase (ProA) tests leads to low discrimination of B. pseudomallei, whereas a set of positive dCEL and negative N-acetyl galactosaminidase, TyrA, and ProA determines the wrong identification of B. pseudomallei as Burkholderia cepacia complex. Conclusion: The further expansion of the Vitek 2 identification keys is needed for correct identification of atypical or regionally distributed biochemical profiles of B. pseudomallei. PMID:29563716

A MULTILAYER BIOCHEMICAL DRY DEPOSITION MODEL 1. MODEL FORMULATION

EPA Science Inventory

A multilayer biochemical dry deposition model has been developed based on the NOAA Multilayer Model (MLM) to study gaseous exchanges between the soil, plants, and the atmosphere. Most of the parameterizations and submodels have been updated or replaced. The numerical integration ...

The dynamics of blood biochemical parameters in cosmonauts during long-term space flights

NASA Astrophysics Data System (ADS)

Markin, Andrei; Strogonova, Lubov; Balashov, Oleg; Polyakov, Valery; Tigner, Timoty

Most of the previously obtained data on cosmonauts' metabolic state concerned certain stages of the postflight period. In this connection, all conclusions, as to metabolism peculiarities during the space flight, were to a large extent probabilistic. The purpose of this work was study of metabolism characteristics in cosmonauts directly during long-term space flights. In the capillary blood samples taken from a finger, by "Reflotron IV" biochemical analyzer, "Boehringer Mannheim" GmbH, Germany, adapted to weightlessness environments, the activity of GOT, GPT, CK, gamma-GT, total and pancreatic amylase, as well as concentration of hemoglobin, glucose, total bilirubin, uric acid, urea, creatinine, total, HDL- and LDL cholesterol, triglycerides had been determined. HDL/LDL-cholesterol ratio also was computed. The crewmembers of 6 main missions to the "Mir" orbital station, a total of 17 cosmonauts, were examined. Biochemical tests were carryed out 30-60 days before lounch, and in the flights different stages between the 25-th and the 423-rd days of flights. In cosmonauts during space flight had been found tendency to increase, in compare with basal level, GOT, GPT, total amylase activity, glucose and total cholesterol concentration, and tendency to decrease of CK activity, hemoglobin, HDL-cholesterol concentration, and HDL/LDL — cholesterol ratio. Some definite trends in variations of other determined biochemical parameters had not been found. The same trends of mentioned biochemical parameters alterations observed in majority of tested cosmonauts, allows to suppose existence of connection between noted metabolic alterations with influence of space flight conditions upon cosmonaut's body. Variations of other studied blood biochemical parameters depends on, probably, pure individual causes.

Influence of low-frequency vibration on changes of biochemical parameters of living rats

NASA Astrophysics Data System (ADS)

Kasprzak, Cezary; Damijan, Zbigniew; Panuszka, Ryszard

2004-05-01

The aim of the research was to investigate how some selected biochemical parameters of living rats depend on exposure of low-frequency vibrations. Experiments were run on 30 Wistar rats randomly segregated into three groups: (I) 20 days old (before puberty), (II) 70th day after; (III) control group. The exposure was repeated seven times, for 3 h, at the same time of day. Vibrations applied during the first tests of the experiment had acceleration 1.22 m/s2 and frequency 20 Hz. At the 135th day the rats' bones were a subject of morphometric/biochemical examination. The results of biochemical tests proved decrease in LDL and HDL cholesterol levels for exposed rats as well as the Ca contents in blood plasma. There was evident increasing of Ca in blood plasma in exposed rats for frequency of exposition.

Microbial diversity and dynamics during the production of May bryndza cheese.

PubMed

Pangallo, Domenico; Saková, Nikoleta; Koreňová, Janka; Puškárová, Andrea; Kraková, Lucia; Valík, Lubomír; Kuchta, Tomáš

2014-01-17

paradoxus. The diversity of yeasts and fungi encompassed Alternaria alternata, "Ascomycete sp.", Aspergillus fumigatus, Beauveria brongniartii, Candida xylopsoci, C. inconspicua, Cladosporium cladosporioides, Debaromyces hansenii, Fomes fomentarius, Galactomyces candidus, Gymnoascus reesii, Chaetomium globosum, Kluyveromyces marxianus, Metarhizium anisopliae, Penicillium aurantiogriseum, P. camemberti, P. freii, P. polonicum, P. viridicatum, Pichia kudriavzevii, Sordaria alcina, Trichosporon lactis and Yarrowia lipolytica. © 2013.

Protease and lipase activities of fungal and bacterial strains derived from an artisanal raw ewe's milk cheese.

PubMed

Ozturkoglu-Budak, Sebnem; Wiebenga, Ad; Bron, Peter A; de Vries, Ronald P

2016-11-21

We previously identified the microbiota present during cheese ripening and observed high protease and lipase activity in Divle Cave cheese. To determine the contribution of individual isolates to enzyme activities, we investigated a range of species representing this microbiota for their proteolytic and lipolytic ability. In total, 17 fungal, 5 yeast and 18 bacterial strains, previously isolated from Divle Cave cheese, were assessed. Qualitative protease and lipase activities were performed on skim-milk agar and spirit-blue lipase agar, respectively, and resulted in a selection of strains for quantitative assays. For the quantitative assays, the strains were grown on minimal medium containing irradiated Divle Cave cheese, obtained from the first day of ripening. Out of 16 selected filamentous fungi, Penicillium brevicompactum, Penicillium cavernicola and Penicillium olsonii showed the highest protease activity, while Mucor racemosus was the best lipase producer. Yarrowia lipolytica was the best performing yeast with respect to protease and lipase activity. From the 18 bacterial strains, 14 and 11 strains, respectively showed protease and lipase activity in agar plates. Micrococcus luteus, Bacillus stratosphericus, Brevibacterium antiquum, Psychrobacter glacincola and Pseudomonas proteolytica displayed the highest protease and lipase activity. The proteases of yeast and filamentous fungi were identified as mainly aspartic protease by specific inhibition with Pepstatin A, whereas inhibition by PMSF (phenylmethylsulfonyl fluoride) indicated that most bacterial enzymes belong to serine type protease. Our results demonstrate that aspartic proteases, which usually have high milk clotting activity, are predominantly derived from fungal strains, and therefore fungal enzymes appear to be more suitable for use in the cheese industry. Microbial enzymes studied in this research might be alternatives for rennin (chymosin) from animal source because of their low cost and stable

Purification and characterization of an alkaline protease from the marine yeast Aureobasidium pullulans for bioactive peptide production from different sources.

PubMed

Ma, Chunling; Ni, Xiumei; Chi, Zhenming; Ma, Liyan; Gao, Lingmei

2007-01-01

The extracellular alkaline protease in the supernatant of cell culture of the marine yeast Aureobasidium pullulans 10 was purified to homogeneity with a 2.1-fold increase in specific protease activity as compared to that in the supernatant by ammonium sulfate fractionation, gel filtration chromatography (Sephadex G-75), and anion-exchange chromatography (DEAE Sepharose Fast Flow). According to the sodium dodecyl sulfate-polyacrylamide gel electrophoresis data, the molecular mass of the purified enzyme was estimated to be 32.0 kDa. The optimal pH and temperature of the purified enzyme were 9.0 and 45 degrees C, respectively. The enzyme was activated by Cu(2+) (at a concentration of 1.0 mM) and Mn(2+) and inhibited by Hg(2+), Fe(2+), Fe(3+), Zn(2+), and Co(2+). The enzyme was strongly inhibited by phenylmethylsulfonyl fluoride, but weakly inhibited by EDTA, 1-10-phenanthroline, and iodoacetic acid. The K(m) and V(max) values of the purified enzyme for casein were 0.25 mg/ml and 0.0286 micromol/min/mg of protein, respectively. After digestion of shrimp protein, spirulina (Arthospira platensis) protein, proteins of marine yeast strains N3C (Yarrowia lipolytica) and YA03a (Hanseniaspora uvarum), milk protein, and casein with the purified alkaline protease, angiotensin I converting enzyme (ACE) inhibitory activities of the resulting peptides reached 85.3%, 12.1%, 29.8%, 22.8%, 14.1%, and 15.5%, respectively, while the antioxidant activities of these were 52.1%. 54.6%, 25.1%, 35%, 12.5%, and 24.2%, respectively, indicating that ACE inhibitory activity of the resulting peptides from the shrimp protein and antioxidant activity of those produced from the spirulina protein were the highest, respectively. These results suggest that the bioactive peptides produced by digestion of the shrimp protein with the purified alkaline protease have potential applications in the food and pharmaceutical industries.

Chronic histiocytic intervillositis - Clinical, biochemical and radiological findings: An observational study.

PubMed

Koby, Lawrence; Keating, Sarah; Malinowski, Ann Kinga; D'Souza, Rohan

2018-04-01

Chronic histiocytic intervillositis (CHI) of the placenta although rare, has a high recurrence rate, is associated with serious adverse pregnancy outcomes and has no available treatment. This study aims to determine clinical, biochemical and radiological factors associated with CHI, to guide management of subsequent pregnancies. This retrospective observational study included consecutive cases with a histopathologic diagnosis of CHI after 18 weeks of gestation, between 2001 and 2014, and no controls. Clinical (maternal, fetal and delivery outcomes), biochemical (first- and second-trimester biomarkers for fetal aneuploidy and serum alkaline phosphatase) and radiological (second- and third-trimester fetal, placental and Doppler ultrasound) factors associated with a histopathological diagnosis of CHI were identified and results presented as percentages. Outcomes of subsequent pregnancies were described. Of 231 identified cases of 'intervillositis', 33 were confirmed to have CHI, of which only 4/33 (12.1%) had prior uncomplicated term deliveries. During pregnancy, 10/18 (55.5%) had abnormal first-trimester screening, 4/16 (25%) had abnormal second-trimester screening, 6/19 (31.6%) had at least one elevated alkaline phosphatase level, and 15/20 (75%) had at least one abnormal feature on mid-trimester placental ultrasound. In subsequent pregnancies that were closely followed with a combination of biochemical and radiologic tests, there were no cases of fetal loss, and lower incidence of fetal growth restriction and preterm birth. No clinical, biochemical or radiological finding is consistently associated with CHI and adverse outcomes thereof. Whether the incorporation of these tests in individualized care-plans could improve outcomes in subsequent pregnancies needs to be studied further. Copyright © 2018 Elsevier Ltd. All rights reserved.

A scalable moment-closure approximation for large-scale biochemical reaction networks

PubMed Central

Kazeroonian, Atefeh; Theis, Fabian J.; Hasenauer, Jan

2017-01-01

Abstract Motivation: Stochastic molecular processes are a leading cause of cell-to-cell variability. Their dynamics are often described by continuous-time discrete-state Markov chains and simulated using stochastic simulation algorithms. As these stochastic simulations are computationally demanding, ordinary differential equation models for the dynamics of the statistical moments have been developed. The number of state variables of these approximating models, however, grows at least quadratically with the number of biochemical species. This limits their application to small- and medium-sized processes. Results: In this article, we present a scalable moment-closure approximation (sMA) for the simulation of statistical moments of large-scale stochastic processes. The sMA exploits the structure of the biochemical reaction network to reduce the covariance matrix. We prove that sMA yields approximating models whose number of state variables depends predominantly on local properties, i.e. the average node degree of the reaction network, instead of the overall network size. The resulting complexity reduction is assessed by studying a range of medium- and large-scale biochemical reaction networks. To evaluate the approximation accuracy and the improvement in computational efficiency, we study models for JAK2/STAT5 signalling and NFκB signalling. Our method is applicable to generic biochemical reaction networks and we provide an implementation, including an SBML interface, which renders the sMA easily accessible. Availability and implementation: The sMA is implemented in the open-source MATLAB toolbox CERENA and is available from https://github.com/CERENADevelopers/CERENA. Contact: jan.hasenauer@helmholtz-muenchen.de or atefeh.kazeroonian@tum.de Supplementary information: Supplementary data are available at Bioinformatics online. PMID:28881983

Early hypofractionated salvage radiotherapy for postprostatectomy biochemical recurrence.

PubMed

Kruser, Tim J; Jarrard, David F; Graf, Andrew K; Hedican, Sean P; Paolone, David R; Wegenke, John D; Liu, Glenn; Geye, Heather M; Ritter, Mark A

2011-06-15

Postprostatectomy adjuvant or salvage radiotherapy, when using standard fractionation, requires 6.5 to 8 weeks of treatment. The authors report on the safety and efficacy of an expedited radiotherapy course for salvage prostate radiotherapy. A total of 108 consecutive patients were treated with salvage radiation therapy to 65 grays (Gy) in 26 fractions of 2.5 Gy. Median follow-up was 32.4 months. Median presalvage prostate-specific antigen (PSA) was 0.44 (range, 0.05-9.50). Eighteen (17%) patients received androgen deprivation after surgery or concurrently with radiation. The actuarial freedom from biochemical failure for the entire group at 4 years was 67% ± 5.3%. An identical 67% control rate was seen at 5 years for the first 50 enrolled patients, whose median follow-up was longer at 43 months. One acute grade 3 genitourinary toxicity occurred, with no acute grade 3 gastrointestinal and no late grade 3 toxicities observed. On univariate analysis, higher Gleason score (P = .006), PSA doubling time ≤12 months (P = .03), perineural invasion (P = .06), and negative margins (P = .06) showed association with unsuccessful salvage. On multivariate analysis, higher Gleason score (P = .057) and negative margins (P = .088) retained an association with biochemical failure. Hypofractionated radiotherapy (65 Gy in 2.5 Gy fractions in about 5 weeks) reduces the length of treatment by from 1-½ to 3 weeks relative to other treatment schedules commonly used, produces low rates of toxicity, and demonstrates encouraging efficacy at 4 to 5 years. Hypofractionation may provide a convenient, resource-efficient, and well-tolerated salvage approach for the estimated 20,000 to 35,000 US men per year experiencing biochemical recurrence after prostatectomy. Copyright © 2010 American Cancer Society.

Recent Advances on the Use of Biochemical Extracts as Filaricidal Agents

PubMed Central

Al-Abd, Nazeh M.; Nor, Zurainee Mohamed; Al-Adhroey, Abdulelah H.; Suhaimi, Anwar; Sivanandam, S.

2013-01-01

Lymphatic filariasis is a parasitic infection that causes a devastating public health and socioeconomic burden with an estimated infection of over 120 million individuals worldwide. The infection is caused by three closely related nematode parasites, namely, Wuchereria bancrofti, Brugia malayi, and B. timori, which are transmitted to human through mosquitoes of Anopheles, Culex, and Aedes genera. The species have many ecological variants and are diversified in terms of their genetic fingerprint. The rapid spread of the disease and the genetic diversification cause the lymphatic filarial parasites to respond differently to diagnostic and therapeutic interventions. This in turn prompts the current challenge encountered in its management. Furthermore, most of the chemical medications used are characterized by adverse side effects. These complications urgently warrant intense prospecting on bio-chemicals that have potent efficacy against either the filarial worms or thier vector. In lieu of this, we presented a review on recent literature that reported the efficacy of filaricidal biochemicals and those employed as vector control agents. In addition, methods used for biochemical extraction, screening procedures, and structure of the bioactive compounds were also presented. PMID:24298292

Is Preoperative Biochemical Testing for Pheochromocytoma Necessary for All Adrenal Incidentalomas?

PubMed Central

Jun, Joo Hyun; Ahn, Hyun Joo; Lee, Sangmin M.; Kim, Jie Ae; Park, Byung Kwan; Kim, Jee Soo; Kim, Jung Han

2015-01-01

Abstract This study examined whether imaging phenotypes obtained from computed tomography (CT) can replace biochemical tests to exclude pheochromocytoma among adrenal incidentalomas (AIs) in the preoperative setting. We retrospectively reviewed the medical records of all patients (n = 251) who were admitted for operations and underwent adrenal-protocol CT for an incidentally discovered adrenal mass from January 2011 to December 2012. Various imaging phenotypes were assessed for their screening power for pheochromocytoma. Final diagnosis was confirmed by biopsy, biochemical tests, and follow-up CT. Pheochromocytomas showed similar imaging phenotypes as malignancies, but were significantly different from adenomas. Unenhanced attenuation values ≤10 Hounsfield units (HU) showed the highest specificity (97%) for excluding pheochromocytoma as a single phenotype. A combination of size ≤3 cm, unenhanced attenuation values ≤ 10 HU, and absence of suspicious morphology showed 100% specificity for excluding pheochromocytoma. Routine noncontrast CT can be used as a screening tool for pheochromocytoma by combining 3 imaging phenotypes: size ≤3 cm, unenhanced attenuation values ≤10 HU, and absence of suspicious morphology, and may substitute for biochemical testing in the preoperative setting. PMID:26559265

Adiabatic coarse-graining and simulations of stochastic biochemical networks

PubMed Central

Sinitsyn, N. A.; Hengartner, Nicolas; Nemenman, Ilya

2009-01-01

We propose a universal approach for analysis and fast simulations of stiff stochastic biochemical networks, which rests on elimination of fast chemical species without a loss of information about mesoscopic, non-Poissonian fluctuations of the slow ones. Our approach is similar to the Born–Oppenheimer approximation in quantum mechanics and follows from the stochastic path integral representation of the cumulant generating function of reaction events. In applications with a small number of chemical reactions, it produces analytical expressions for cumulants of chemical fluxes between the slow variables. This allows for a low-dimensional, interpretable representation and can be used for high-accuracy, low-complexity coarse-grained numerical simulations. As an example, we derive the coarse-grained description for a chain of biochemical reactions and show that the coarse-grained and the microscopic simulations agree, but the former is 3 orders of magnitude faster. PMID:19525397

Prostate cancer: role of pretreatment multiparametric 3-T MRI in predicting biochemical recurrence after radical prostatectomy.

PubMed

Park, Jung Jae; Kim, Chan Kyo; Park, Sung Yoon; Park, Byung Kwan; Lee, Hyun Moo; Cho, Seong Whi

2014-05-01

The purpose of this study is to retrospectively investigate whether pretreatment multiparametric MRI findings can predict biochemical recurrence in patients who underwent radical prostatectomy (RP) for localized prostate cancer. In this study, 282 patients with biopsy-proven prostate cancer who received RP underwent pretreatment MRI using a phased-array coil at 3 T, including T2-weighted imaging (T2WI), diffusion-weighted imaging (DWI), and dynamic contrast-enhanced MRI (DCE-MRI). MRI variables included apparent tumor presence on combined imaging sequences, extracapsular extension, and tumor size on DWI or DCE-MRI. Clinical variables included baseline prostate-specific antigen (PSA) level, clinical stage, and Gleason score at biopsy. The relationship between clinical and imaging variables and biochemical recurrence was evaluated using Cox regression analysis. After a median follow-up of 26 months, biochemical recurrence developed in 61 patients (22%). Univariate analysis revealed that all the imaging and clinical variables were significantly associated with biochemical recurrence (p < 0.01). On multivariate analysis, however, baseline PSA level (p = 0.002), Gleason score at biopsy (p = 0.024), and apparent tumor presence on combined T2WI, DWI, and DCE-MRI (p = 0.047) were the only significant independent predictors of biochemical recurrence. Of the independent predictors, apparent tumor presence on combined T2WI, DWI, and DCE-MRI showed the highest hazard ratio (2.38) compared with baseline PSA level (hazard ratio, 1.05) and Gleason score at biopsy (hazard ratio, 1.34). The apparent tumor presence on combined T2WI, DWI, and DCE-MRI of pretreatment MRI is an independent predictor of biochemical recurrence after RP. This finding may be used to construct a predictive model for biochemical recurrence after surgery.

Modelling biochemical reaction systems by stochastic differential equations with reflection.

PubMed

Niu, Yuanling; Burrage, Kevin; Chen, Luonan

2016-05-07

In this paper, we gave a new framework for modelling and simulating biochemical reaction systems by stochastic differential equations with reflection not in a heuristic way but in a mathematical way. The model is computationally efficient compared with the discrete-state Markov chain approach, and it ensures that both analytic and numerical solutions remain in a biologically plausible region. Specifically, our model mathematically ensures that species numbers lie in the domain D, which is a physical constraint for biochemical reactions, in contrast to the previous models. The domain D is actually obtained according to the structure of the corresponding chemical Langevin equations, i.e., the boundary is inherent in the biochemical reaction system. A variant of projection method was employed to solve the reflected stochastic differential equation model, and it includes three simple steps, i.e., Euler-Maruyama method was applied to the equations first, and then check whether or not the point lies within the domain D, and if not perform an orthogonal projection. It is found that the projection onto the closure D¯ is the solution to a convex quadratic programming problem. Thus, existing methods for the convex quadratic programming problem can be employed for the orthogonal projection map. Numerical tests on several important problems in biological systems confirmed the efficiency and accuracy of this approach. Copyright © 2016 Elsevier Ltd. All rights reserved.

A Diagrammatic Language for Biochemical Networks

NASA Astrophysics Data System (ADS)

Maimon, Ron

2002-03-01

I present a diagrammatic language for representing the structure of biochemical networks. The language is designed to represent modular structure in a computational fasion, with composition of reactions replacing functional composition. This notation is used to represent arbitrarily large networks efficiently. The notation finds its most natural use in representing biological interaction networks, but it is a general computing language appropriate to any naturally occuring computation. Unlike lambda-calculus, or text-derived languages, it does not impose a tree-structure on the diagrams, and so is more effective at representing biological fucntion than competing notations.

Biochemical and biomechanical characterisation of equine cervical facet joint cartilage.

PubMed

O'Leary, S A; White, J L; Hu, J C; Athanasiou, K A

2018-04-15

The equine cervical facet joint is a site of significant pathology. Located bilaterally on the dorsal spine, these diarthrodial joints work in conjunction with the intervertebral disc to facilitate appropriate spinal motion. Despite the high prevalence of pathology in this joint, the facet joint is understudied and thus lacking in viable treatment options. The goal of this study was to characterise equine facet joint cartilage and provide a comprehensive database describing the morphological, histological, biochemical and biomechanical properties of this tissue. Descriptive cadaver studies. A total of 132 facet joint surfaces were harvested from the cervical spines of six skeletally mature horses (11 surfaces per animal) for compiling biomechanical and biochemical properties of hyaline cartilage of the equine cervical facet joints. Gross morphometric measurements and histological staining were performed on facet joint cartilage. Creep indentation and uniaxial strain-to-failure testing were used to determine the biomechanical compressive and tensile properties. Biochemical assays included quantification of total collagen, sulfated glycosaminoglycan and DNA content. The facet joint surfaces were ovoid in shape with a flat articular surface. Histological analyses highlighted structures akin to articular cartilage of other synovial joints. In general, biomechanical and biochemical properties did not differ significantly between the inferior and superior joint surfaces as well as among spinal levels. Interestingly, compressive and tensile properties of cervical facet articular cartilage were lower than those of articular cartilage from other previously characterised equine joints. Removal of the superficial zone reduced the tissue's tensile strength, suggesting that this zone is important for the tensile integrity of the tissue. Facet surfaces were sampled at a single, central location and do not capture the potential topographic variation in cartilage properties. This