Cell wall glycoproteins at interaction sites between parasitic giant dodder (Cuscuta reflexa) and its host Pelargonium zonale.

PubMed

Striberny, Bernd; Krause, Kirsten

2015-01-01

The process of host plant penetration by parasitic dodder (genus Cuscuta) is accompanied by molecular and structural changes at the host/parasite interface. Recently, changes in pectin methyl esterification levels in the host cell walls abutting parasitic cells in established infection sites were reported. In addition to that, we show here that the composition of cell wall glycoproteins in Cuscuta-infected Pelargonium zonale undergoes substantial changes. While several arabinogalactan protein epitopes exhibit decreased abundances in the vicinity of the Cuscuta reflexa haustorium, extensins tend to increase in the infected areas.

The N-Linked Outer Chain Mannans and the Dfg5p and Dcw1p Endo-α-1,6-Mannanases Are Needed for Incorporation of Candida albicans Glycoproteins into the Cell Wall

PubMed Central

Ao, Jie; Chinnici, Jennifer L.; Maddi, Abhiram

2015-01-01

A biochemical pathway for the incorporation of cell wall protein into the cell wall of Neurospora crassa was recently proposed. In this pathway, the DFG-5 and DCW-1 endo-α-1,6-mannanases function to covalently cross-link cell wall protein-associated N-linked galactomannans, which are structurally related to the yeast outer chain mannans, into the cell wall glucan-chitin matrix. In this report, we demonstrate that the mannosyltransferase enzyme Och1p, which is needed for the synthesis of the N-linked outer chain mannan, is essential for the incorporation of cell wall glycoproteins into the Candida albicans cell wall. Using endoglycosidases, we show that C. albicans cell wall proteins are cross-linked into the cell wall via their N-linked outer chain mannans. We further demonstrate that the Dfg5p and Dcw1p α-1,6-mannanases are needed for the incorporation of cell wall glycoproteins into the C. albicans cell wall. Our results support the hypothesis that the Dfg5p and Dcw1p α-1,6-mannanases incorporate cell wall glycoproteins into the C. albicans cell wall by cross-linking outer chain mannans into the cell wall glucan-chitin matrix. PMID:26048011

Interdisciplinary cantilever physics: Elasticity of carrot, celery, and plasticware

NASA Astrophysics Data System (ADS)

Pestka, Kenneth A.

2014-05-01

This article presents several simple cantilever-based experiments using common household items (celery, carrot, and a plastic spoon) that are appropriate for introductory undergraduate laboratories or independent student projects. By applying Hooke's law and Euler beam theory, students are able to determine Young's modulus, fracture stress, yield stress, strain energy, and sound speed of these apparently disparate materials. In addition, a cellular foam elastic model is introduced—applicable to biologic materials as well as an essential component in the development of advanced engineering composites—that provides a mechanism to determine Young's modulus of the cell wall material found in celery and carrot. These experiments are designed to promote exploration of the similarities and differences between common inorganic and organic materials, fill a void in the typical undergraduate curriculum, and provide a foundation for more advanced material science pursuits within biology, botany, and food science as well as physics and engineering.

Process quality planning of quality function deployment for carrot syrup

NASA Astrophysics Data System (ADS)

Ekawati, Yurida; Noya, Sunday; Widjaja, Filemon

2017-06-01

Carrot products are rarely available in the market. Based on previous research that had been done using QFD to generate product design of carrots products, the research to produce the process quality planning had been carried out. The carrot product studied was carrot syrup. The research resulted in a process planning matrix for carrot syrup. The matrix gives information about critical process plan and the priority of the critical process plan. The critical process plan on the production process of carrot syrup consists of carrots sorting, carrots peeling, carrots washing, blanching process, carrots cutting, the making of pureed carrots, filtering carrot juice, the addition of sugar in carrot juice, the addition of food additives in carrot juice, syrup boiling, syrup filtering, syrup filling into the bottle, the bottle closure and cooling. The information will help the design of the production process of carrot syrup.

Electrohydrodynamic drying of carrot slices.

PubMed

Ding, Changjiang; Lu, Jun; Song, Zhiqing

2015-01-01

Carrots have one of the highest levels of carotene, and they are rich in vitamins, fiber and minerals. However, since fresh carrots wilt rapidly after harvest under inappropriate storage conditions, drying has been used to improve their shelf life and retain nutritional quality. Therefore, to further investigate the potential of this method, carrot slices were dried in an EHD system in order to study the effect of different voltages on drying rate. As measures of quality, carotene content and rehydration ratio were, respectively, compared against the conventional oven drying regime. Carotene, the main component of the dried carrot, and rehydration characteristics of the dried product can both indicate quality by physical and chemical changes during the drying process. Mathematical modeling and simulation of drying curves were also performed, using root mean square error, reduced mean square of the deviation and modeling efficiency as the primary criteria to select the equation that best accounts for the variation in the drying curves of the dried samples. Theoretically, the Page model was best suited for describing the drying rate curve of carrot slices at 10kV to 30kV. Experimentally, the drying rate of carrots was notably greater in the EHD system when compared to control, and quality, as determined by carotene content and rehydration ratio, was also improved when compared to oven drying. Therefore, this work presents a facile and effective strategy for experimentally and theoretically determining the drying properties of carrots, and, as a result, it provides deeper insight into the industrial potential of the EHD drying technique.

(Hydroxyproline-rich glycoprotein of the plant cell wall): Report on work from June 1987 to June 1988

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

Not Available

1988-01-01

In soybean seed costs the accumulation of the hydroxproline-rich glycoprotein extensin is regulated in a developmental and tissue-specific manner. The time course of appearance of extensin during seed development was studied by Western blot analysis and by immunogold-silver localization. Using these techniques extensin was first detected at 16 to 18 d after anthesis, increasing during development to high levels at 24 d after anthesis. Immunogold-silver localization of extensin in the seed coat showed marked depostion of the glycoprotein in the walls of palisade epidermal cells and hourglass cells. The immunolocalization of extensin in developing soybean seeds was also made bymore » a new technique - tissue printing on nitrocellulose paper. This technique shows that extensin is primarily localized in the seed coal, hilum, and vascular elements of the seed.« less

Carbohydrate-dependent binding of langerin to SodC, a cell wall glycoprotein of Mycobacterium leprae.

PubMed

Kim, Hee Jin; Brennan, Patrick J; Heaslip, Darragh; Udey, Mark C; Modlin, Robert L; Belisle, John T

2015-02-01

Langerhans cells participate in the immune response in leprosy by their ability to activate T cells that recognize the pathogen, Mycobacterium leprae, in a langerin-dependent manner. We hypothesized that langerin, the distinguishing C-type lectin of Langerhans cells, would recognize the highly mannosylated structures in pathogenic Mycobacterium spp. The coding region for the extracellular and neck domain of human langerin was cloned and expressed to produce a recombinant active trimeric form of human langerin (r-langerin). Binding assays performed in microtiter plates, by two-dimensional (2D) Western blotting, and by surface plasmon resonance demonstrated that r-langerin possessed carbohydrate-dependent affinity to glycoproteins in the cell wall of M. leprae. This lectin, however, yielded less binding to mannose-capped lipoarabinomannan (ManLAM) and even lower levels of binding to phosphatidylinositol mannosides. However, the superoxide dismutase C (SodC) protein of the M. leprae cell wall was identified as a langerin-reactive ligand. Tandem mass spectrometry verified the glycosylation of a recombinant form of M. leprae SodC (rSodC) produced in Mycobacterium smegmatis. Analysis of r-langerin affinity by surface plasmon resonance revealed a carbohydrate-dependent affinity of rSodC (equilibrium dissociation constant [KD] = 0.862 μM) that was 20-fold greater than for M. leprae ManLAM (KD = 18.69 μM). These data strongly suggest that a subset of the presumptively mannosylated M. leprae glycoproteins act as ligands for langerin and may facilitate the interaction of M. leprae with Langerhans cells. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Carbohydrate-Dependent Binding of Langerin to SodC, a Cell Wall Glycoprotein of Mycobacterium leprae

PubMed Central

Kim, Hee Jin; Brennan, Patrick J.; Heaslip, Darragh; Udey, Mark C.; Modlin, Robert L.

2014-01-01

Langerhans cells participate in the immune response in leprosy by their ability to activate T cells that recognize the pathogen, Mycobacterium leprae, in a langerin-dependent manner. We hypothesized that langerin, the distinguishing C-type lectin of Langerhans cells, would recognize the highly mannosylated structures in pathogenic Mycobacterium spp. The coding region for the extracellular and neck domain of human langerin was cloned and expressed to produce a recombinant active trimeric form of human langerin (r-langerin). Binding assays performed in microtiter plates, by two-dimensional (2D) Western blotting, and by surface plasmon resonance demonstrated that r-langerin possessed carbohydrate-dependent affinity to glycoproteins in the cell wall of M. leprae. This lectin, however, yielded less binding to mannose-capped lipoarabinomannan (ManLAM) and even lower levels of binding to phosphatidylinositol mannosides. However, the superoxide dismutase C (SodC) protein of the M. leprae cell wall was identified as a langerin-reactive ligand. Tandem mass spectrometry verified the glycosylation of a recombinant form of M. leprae SodC (rSodC) produced in Mycobacterium smegmatis. Analysis of r-langerin affinity by surface plasmon resonance revealed a carbohydrate-dependent affinity of rSodC (equilibrium dissociation constant [KD] = 0.862 μM) that was 20-fold greater than for M. leprae ManLAM (KD = 18.69 μM). These data strongly suggest that a subset of the presumptively mannosylated M. leprae glycoproteins act as ligands for langerin and may facilitate the interaction of M. leprae with Langerhans cells. PMID:25422308

Recent advance in carrot genomics

USDA-ARS?s Scientific Manuscript database

In recent years there has been an effort towards the development of genomic resources in carrot. The number of available sequences for carrot in public databases has increased recently. This has allowed the design of SSRs markers, COS markers and a high-throughput SNP assay for genotyping. Additiona...

First Report of 'Candidatus Liberibacter solanacearum' in Carrots in Europe

USDA-ARS?s Scientific Manuscript database

Carrot (Daucus carota) plants exhibiting symptoms that resembled those of carrot psyllid (Trioza apicalis) damage were observed in commercial fields in southern Finland in August 2008. Carrot psyllid is a serious pest of carrots in northern and central Europe, where it can cause up to 100% yield los...

Roles of Tyrosine-Rich Precursor Glycoproteins and Dityrosine- and 3,4-Dihydroxyphenylalanine-Mediated Protein Cross-Linking in Development of the Oocyst Wall in the Coccidian Parasite Eimeria maxima

PubMed Central

Belli, Sabina I.; Wallach, Michael G.; Luxford, Catherine; Davies, Michael J.; Smith, Nicholas C.

2003-01-01

The oocyst wall of apicomplexan parasites protects them from the harsh external environment, preserving their survival prior to transmission to the next host. If oocyst wall formation could be disrupted, then logically, the cycle of disease transmission could be stopped, and strategies to control infection by several organisms of medical and veterinary importance such as Eimeria, Plasmodium, Toxoplasma, Cyclospora, and Neospora could be developed. Here, we show that two tyrosine-rich precursor glycoproteins, gam56 and gam82, found in specialized organelles (wall-forming bodies) in the sexual stage (macrogamete) of Eimeria maxima are proteolytically processed into smaller glycoproteins, which are then incorporated into the developing oocyst wall. The identification of high concentrations of dityrosine and 3,4-dihydroxyphenylalanine (DOPA) in oocyst extracts by high-pressure liquid chromatography, together with the detection of a UV autofluorescence in intact oocysts, implicates dityrosine- and possibly DOPA-protein cross-links in oocyst wall hardening. In addition, the identification of peroxidase activity in the wall-forming bodies of macrogametes supports the hypothesis that dityrosine- and DOPA-mediated cross-linking might be an enzyme-catalyzed event. As such, the mechanism of oocyst wall formation in Eimeria, is analogous to the underlying mechanisms involved in the stabilization of extracellular matrices in a number of organisms, widely distributed in nature, including insect resilin, nematode cuticles, yeast cell walls, mussel byssal threads, and sea urchin fertilization membranes. PMID:12796290

21 CFR 73.300 - Carrot oil.

Code of Federal Regulations, 2011 CFR

2011-04-01

... in color additive mixtures for coloring foods. (b) Specifications. Carrot oil shall contain no more... coloring foods generally, in amounts consistent with good manufacturing practice, except that it may not be... 21 Food and Drugs 1 2011-04-01 2011-04-01 false Carrot oil. 73.300 Section 73.300 Food and Drugs...

Solid Loss of Carrots During Simulated Gastric Digestion.

PubMed

Kong, Fanbin; Singh, R Paul

2011-03-01

The knowledge of solid loss kinetics of foods during digestion is crucial for understanding the factors that constrain the release of nutrients from the food matrix and their fate of digestion. The objective of this study was to investigate the solid loss of carrots during simulated gastric digestion as affected by pH, temperature, viscosity of gastric fluids, mechanical force present in stomach, and cooking. Cylindrical carrot samples were tested by static soaking method and using a model stomach system. The weight retention, moisture, and loss of dry mass were determined. The results indicated that acid hydrolysis is critical for an efficient mass transfer and carrot digestion. Internal resistance rather than external resistance is dominant in the transfer of soluble solids from carrot to gastric fluid. Increase in viscosity of gastric fluid by adding 0.5% gum (w/w) significantly increased the external resistance and decreased mass transfer rate of carrots in static soaking. When mechanical force was not present, 61% of the solids in the raw carrot samples were released into gastric fluid after 4 h of static soaking in simulated gastric juice. Mechanical force significantly increased solid loss by causing surface erosion. Boiling increased the disintegration of carrot during digestion that may favor the loss of solids meanwhile reducing the amount of solids available for loss in gastric juice. Weibull function was successfully used to describe the solid loss of carrot during simulated digestion. The effective diffusion coefficients of solids were calculated using the Fick's second law of diffusion for an infinite cylinder, which are between 0.75 × 10(-11) and 8.72 × 10(-11) m(2)/s, depending on the pH of the gastric fluid.

cDNA cloning of carrot extracellular beta-fructosidase and its expression in response to wounding and bacterial infection.

PubMed

Sturm, A; Chrispeels, M J

1990-11-01

We isolated a full-length cDNA for apoplastic (extracellular or cell wall-bound) beta-fructosidase (invertase), determined its nucleotide sequence, and used it as a probe to measure changes in mRNA as a result of wounding of carrot storage roots and infection of carrot plants with the bacterial pathogen Erwinia carotovora. The derived amino acid sequence of extracellular beta-fructosidase shows that it is a basic protein (pl 9.9) with a signal sequence for entry into the endoplasmic reticulum and a propeptide at the N terminus that is not present in the mature protein. Amino acid sequence comparison with yeast and bacterial invertases shows that the overall homology is only about 28%, but that there are short conserved motifs, one of which is at the active site. Maturing carrot storage roots contain barely detectable levels of mRNA for extracellular beta-fructosidase and these levels rise slowly but dramatically after wounding with maximal expression after 12 hours. Infection of roots and leaves of carrot plants with E. carotovora results in a very fast increase in the mRNA levels with maximal expression after 1 hour. These results indicate that apoplastic beta-fructosidase is probably a new and hitherto unrecognized pathogenesis-related protein [Van Loon, L.C. (1985). Plant Mol. Biol. 4, 111-116]. Suspension-cultured carrot cells contain high levels of mRNA for extracellular beta-fructosidase and these levels remain the same whether the cells are grown on sucrose, glucose, or fructose.

The Most Abundant Glycoprotein of Amebic Cyst Walls (Jacob) Is a Lectin with Five Cys-Rich, Chitin-Binding Domains

PubMed Central

Frisardi, Marta; Ghosh, Sudip K.; Field, Jessica; Van Dellen, Katrina; Rogers, Rick; Robbins, Phillips; Samuelson, John

2000-01-01

The infectious stage of amebae is the chitin-walled cyst, which is resistant to stomach acids. In this study an extraordinarily abundant, encystation-specific glycoprotein (Jacob) was identified on two-dimensional protein gels of cyst walls purified from Entamoeba invadens. Jacob, which was acidic and had an apparent molecular mass of ∼100 kDa, contained sugars that bound to concanavalin A and ricin. The jacob gene encoded a 45-kDa protein with a ladder-like series of five Cys-rich domains. These Cys-rich domains were reminiscent of but not homologous to the Cys-rich chitin-binding domains of insect chitinases and peritrophic matrix proteins that surround the food bolus in the insect gut. Jacob bound purified chitin and chitin remaining in sodium dodecyl sulfate-treated cyst walls. Conversely, the E. histolytica plasma membrane Gal/GalNAc lectin bound sugars of intact cyst walls and purified Jacob. In the presence of galactose, E. invadens formed wall-less cysts, which were quadranucleate and contained Jacob and chitinase (another encystation-specific protein) in secretory vesicles. A galactose lectin was found to be present on the surface of wall-less cysts, which phagocytosed bacteria and mucin-coated beads. These results suggest that the E. invadens cyst wall forms when the plasma membrane galactose lectin binds sugars on Jacob, which in turn binds chitin via its five chitin-binding domains. PMID:10858239

Influence of technical processing units on chemical composition and antimicrobial activity of carrot (Daucus carrot L.) juice essential oil.

PubMed

Ma, Tingting; Luo, Jiyang; Tian, Chengrui; Sun, Xiangyu; Quan, Meiping; Zheng, Cuiping; Kang, Lina; Zhan, Jicheng

2015-03-01

The effect of three processing units (blanching, enzyme liquefaction, pasteurisation) on chemical composition and antimicrobial activity of carrot juice essential oil was investigated in this paper. A total of 36 compounds were identified by GC-MS from fresh carrot juice essential oil. The main constituents were carotol (20.20%), sabinene (12.80%), β-caryophyllene (8.04%) and α-pinene (6.05%). Compared with the oil of fresh juice, blanching and pasteurisation could significantly decrease the components of the juice essential oil, whereas enzyme liquefaction had no considerable effect on the composition of juice essential oil. With regard to the antimicrobial activity, carrot juice essential oil could cause physical damage and morphological alteration on microorganisms, while the three different processing units showed noticeable differences on the species of microorganisms, the minimum inhibitory concentration and minimum bactericidal concentration. Results revealed that the carrot juice essential oil has great potential for application as a natural antimicrobial applied in pharmaceutical and food industries. Copyright © 2014 Elsevier Ltd. All rights reserved.

cDNA cloning of carrot extracellular beta-fructosidase and its expression in response to wounding and bacterial infection.

PubMed Central

Sturm, A; Chrispeels, M J

1990-01-01

We isolated a full-length cDNA for apoplastic (extracellular or cell wall-bound) beta-fructosidase (invertase), determined its nucleotide sequence, and used it as a probe to measure changes in mRNA as a result of wounding of carrot storage roots and infection of carrot plants with the bacterial pathogen Erwinia carotovora. The derived amino acid sequence of extracellular beta-fructosidase shows that it is a basic protein (pl 9.9) with a signal sequence for entry into the endoplasmic reticulum and a propeptide at the N terminus that is not present in the mature protein. Amino acid sequence comparison with yeast and bacterial invertases shows that the overall homology is only about 28%, but that there are short conserved motifs, one of which is at the active site. Maturing carrot storage roots contain barely detectable levels of mRNA for extracellular beta-fructosidase and these levels rise slowly but dramatically after wounding with maximal expression after 12 hours. Infection of roots and leaves of carrot plants with E. carotovora results in a very fast increase in the mRNA levels with maximal expression after 1 hour. These results indicate that apoplastic beta-fructosidase is probably a new and hitherto unrecognized pathogenesis-related protein [Van Loon, L.C. (1985). Plant Mol. Biol. 4, 111-116]. Suspension-cultured carrot cells contain high levels of mRNA for extracellular beta-fructosidase and these levels remain the same whether the cells are grown on sucrose, glucose, or fructose. PMID:2152110

Radicinols and radicinin phytotoxins produced by Alternaria radicina on carrots.

PubMed

Solfrizzo, Michele; Vitti, Carolina; De Girolamo, Annalisa; Visconti, Angelo; Logrieco, Antonio; Fanizzi, Francesco P

2004-06-02

The phytotoxin epi-radicinol, a diastereomer of radicinol, was isolated from large cultures of Alternaria radicina grown on carrot slices and identified by GC-MS, LC-MS, (1)H NMR, and (13)C NMR. Four strains of A. radicina isolated from rotted carrot produced epi-radicinol as the major metabolite (up to 39414 microg/g) together with radicinol (up to 2423 microg/g), and, to a lesser extent, radicinin when cultured on carrot slices, whereas on rice they mainly produced radicinin (2486-53800 microg/g). Radicinin and epi-radicinol reduced root elongation of germinating carrot seeds at concentrations of 10-20 microg/mL. Carrot samples naturally infected by A. radicina contained detectable quantities of epi-radicinol also in combination with lower levels of radicinin or radicinol. Accumulation of radicinols and radicinin in stored carrots, either naturally contaminated or artificially inoculated with A. radicina, was stimulated by successive temperature rises from 1 to 10 degrees C and from 10 to 20 degrees C, reaching maximum levels of 60 microg/g epi-radicinol and 26 microg/g radicinin. This is the first report on the production of radicinols by A. radicina and its natural occurrence in carrots in association with radicinin.

Vegetables of temperate climates: Carrot, parsnip and beetroot

USDA-ARS?s Scientific Manuscript database

Carrots, parsnips and beets are horticultural root crops eaten around the world. These root vegetables have a long shelf-life when kept in cold storage. Among them are sources of vitamin A, vitamin C, fiber, and other colorful pigments that provide antioxidant activity. Carrots come in many color...

Polynuclear aromatic hydrocarbon uptake by carrots grown in sludge-amended soil

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

Wild, S.R.; Jones, K.C.

The uptake of polynuclear aromatic hydrocarbons (PAHs) from sewage sludge-amended soils by carrots (Daucus carota) was investigated. Carrots were grown in control soils and soils amended with three sludge application rates, 15, 55, and 180 t/ha. Applied sludge contained 17.2 mg [summation]PAH/kg, a concentration typical for a sludge derived from a rural area. Carrot foliage, root peels and root cores were analyzed for 15 PAH compounds. Carrots foliage PAH concentrations were unaffected by sludge applications (PAH loadings), but root peel PAH concentrations increased to a plateau concentration with increasing soil PAH levels. Low molecular weight PAH compounds dominate dindividual componentsmore » of the [summation]PAH load in the root tissues. The PAH concentrations detected in the root peels were all significantly lower than in the foliage, which receives PAH inputs from the atmosphere. Carrot core [summation]PAH concentrations were unaffected by sludge application, implying little or no transfer of PAHs from the peels to the core. About 70% of the PAH burden found in carrots was associated with the peels. Fresh weight carrot core concentrations were all <4.2 [mu]g/kg. Overall, this investigation suggests that the risks posed to human health by PAHs applied in sewage sludge to arable soils are minimal.« less

Detection and prediction of post harvest carrot diseases

USDA-ARS?s Scientific Manuscript database

Carrot is the most important outdoor vegetable crop in Norway with a market value of NOK 675 million in 2010. Because demand for carrots is year-round but the growing season is short, the crop is typically stored after harvest for as long as 6 months. Diseases that develop during low temperature sto...

Titanium dioxide/UV photocatalytic disinfection in fresh carrots.

PubMed

Cho, Mihee; Choi, Yoonjung; Park, Hyojin; Kim, Kwansik; Woo, Gun-Jo; Park, Jiyong

2007-01-01

Increased occurrences of fresh produce-related outbreaks of foodborne illness have focused attention on effective washing processes for fruits and vegetables. A titanium dioxide (TiO2) photocatalytic reaction under UV radiation provides a high rate of disinfection. The photo-killing effects of TiO2 on bacteria in liquid cultures under experimental conditions have been widely studied. However, the disinfection effects of the TiO2 photocatalytic reaction on fresh vegetables during a washing process have not been evaluated. Our objectives were to design a pilot-scale TiO2/UV photocatalytic reactor for fresh carrots and to compare the bactericidal effects of the TiO2/UV reaction against bacteria in liquid media and on carrots. TiO2/UV photocatalytic reactions for 40, 60, and 30 s were required for the complete killing of Escherichia coli, Salmonella Typhimurium, and Bacillus cereus (initial counts of approximately 6.7 log CFU/ml), respectively. The counts of total aerobic bacteria in fresh carrots and foodborne pathogenic bacteria in inoculated carrots were also measured. Counts of total aerobic bacteria were reduced by 1.8 log CFU/g after TiO2/UV photocatalytic disinfection for 20 min compared with a 1.1-log CFU/g reduction by UV alone. E. coli, Salmonella Typhimurium, and B. cereus (8 log CFU/ml) were inoculated onto carrots, and the number of surviving bacteria in carrots was determined after treatment. The TiO2/UV treatment exhibited 2.1-, 2.3-, and 1.8-log CFU/g reductions in the counts of E. coli, Salmonella Typhimurium, and B. cereus, respectively, compared with 1.3-, 1.2-, and 1.2-log CFU/g reductions by UV alone. The TiO2/UV photocatalyst reaction showed significant bactericidal effects, indicating that this process is applicable to nonthermal disinfection of fresh vegetables.

Efficient CRISPR/Cas9-based genome editing in carrot cells.

PubMed

Klimek-Chodacka, Magdalena; Oleszkiewicz, Tomasz; Lowder, Levi G; Qi, Yiping; Baranski, Rafal

2018-04-01

The first report presenting successful and efficient carrot genome editing using CRISPR/Cas9 system. Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/CRISPR-associated (Cas9) is a powerful genome editing tool that has been widely adopted in model organisms recently, but has not been used in carrot-a model species for in vitro culture studies and an important health-promoting crop grown worldwide. In this study, for the first time, we report application of the CRISPR/Cas9 system for efficient targeted mutagenesis of the carrot genome. Multiplexing CRISPR/Cas9 vectors expressing two single-guide RNA (gRNAs) targeting the carrot flavanone-3-hydroxylase (F3H) gene were tested for blockage of the anthocyanin biosynthesis in a model purple-colored callus using Agrobacterium-mediated genetic transformation. This approach allowed fast and visual comparison of three codon-optimized Cas9 genes and revealed that the most efficient one in generating F3H mutants was the Arabidopsis codon-optimized AteCas9 gene with up to 90% efficiency. Knockout of F3H gene resulted in the discoloration of calli, validating the functional role of this gene in the anthocyanin biosynthesis in carrot as well as providing a visual marker for screening successfully edited events. Most resulting mutations were small Indels, but long chromosome fragment deletions of 116-119 nt were also generated with simultaneous cleavage mediated by two gRNAs. The results demonstrate successful site-directed mutagenesis in carrot with CRISPR/Cas9 and the usefulness of a model callus culture to validate genome editing systems. Given that the carrot genome has been sequenced recently, our timely study sheds light on the promising application of genome editing tools for boosting basic and translational research in this important vegetable crop.

A carrot leucine-rich-repeat protein that inhibits ice recrystallization.

PubMed

Worrall, D; Elias, L; Ashford, D; Smallwood, M; Sidebottom, C; Lillford, P; Telford, J; Holt, C; Bowles, D

1998-10-02

Many organisms adapted to live at subzero temperatures express antifreeze proteins that improve their tolerance to freezing. Although structurally diverse, all antifreeze proteins interact with ice surfaces, depress the freezing temperature of aqueous solutions, and inhibit ice crystal growth. A protein purified from carrot shares these functional features with antifreeze proteins of fish. Expression of the carrot complementary DNA in tobacco resulted in the accumulation of antifreeze activity in the apoplast of plants grown at greenhouse temperatures. The sequence of carrot antifreeze protein is similar to that of polygalacturonase inhibitor proteins and contains leucine-rich repeats.

Trace Determination of Linear Alkylbenzene Sulfonates: Application in Artificially Polluted Soil—Carrots System

PubMed Central

Sablayrolles, Caroline; Montréjaud-Vignoles, Mireille; Silvestre, Jérôme; Treilhou, Michel

2009-01-01

Surfactants are widely used in household and industrial products. The risk of incorporation of linear alkylbenzene sulfonates (LAS) from biosolids, wastewater, and fertilizers land application to the food chain is being assessed at present by the European Union. In the present work, a complete analytical method for LAS trace determination has been developed and successfully applied to LAS (C10–C13) uptake in carrot plants used as model. These carrots were grown in soil with the trace organics compounds added directly into the plant containers in pure substances form. LAS trace determination (μg kg−1 dry matter) in carrots samples was achieved by Soxtec apparatus and high-performance liquid chromatography-fluorescence detection. The methodology developed provides LAS determination at low detection limits (5 μg kg−1 dry matter) for carrot sample (2 g dry matter) with good recoveries rate (>90%). Transfer of LAS has been followed into the various parts of the carrot plant. LAS are generally found in the carrot leaves and percentage transfer remains very low (0.02%). PMID:20107562

First Report of ‘Candidatus Liberibacter solanacearum’ Associated with Psyllid-Infested Carrots in Germany

USDA-ARS?s Scientific Manuscript database

Carrot plants with symptoms resembling those associated with the bacterium “Candidatus Liberibacter solanacearum” were observed in commercial carrot fields in Lower Saxony, Germany in September 2014. The fields were infested with the carrot psyllid Trioza apicalis and the infection rate was about 50...

Effect of UV-B light and different cutting styles on antioxidant enhancement of commercial fresh-cut carrot products

USDA-ARS?s Scientific Manuscript database

The results of the present study show that exposing five commercial fresh-cut carrot products (baby carrots, carrot sticks, shredded carrots, crinkle cut coins, and oblong chips) to UV-B radiation can significantly increase the antioxidant capacity of carrots. Optimization of the nutritional benefic...

Partial Resistance of Carrot to Alternaria dauci Correlates with In Vitro Cultured Carrot Cell Resistance to Fungal Exudates

PubMed Central

Voisine, Linda; Gatto, Julia; Hélesbeux, Jean-Jacques; Séraphin, Denis; Peña-Rodriguez, Luis M.; Richomme, Pascal; Boedo, Cora; Yovanopoulos, Claire; Gyomlai, Melvina; Briard, Mathilde; Simoneau, Philippe; Poupard, Pascal; Berruyer, Romain

2014-01-01

Although different mechanisms have been proposed in the recent years, plant pathogen partial resistance is still poorly understood. Components of the chemical warfare, including the production of plant defense compounds and plant resistance to pathogen-produced toxins, are likely to play a role. Toxins are indeed recognized as important determinants of pathogenicity in necrotrophic fungi. Partial resistance based on quantitative resistance loci and linked to a pathogen-produced toxin has never been fully described. We tested this hypothesis using the Alternaria dauci – carrot pathosystem. Alternaria dauci, causing carrot leaf blight, is a necrotrophic fungus known to produce zinniol, a compound described as a non-host selective toxin. Embryogenic cellular cultures from carrot genotypes varying in resistance against A. dauci were confronted with zinniol at different concentrations or to fungal exudates (raw, organic or aqueous extracts). The plant response was analyzed through the measurement of cytoplasmic esterase activity, as a marker of cell viability, and the differentiation of somatic embryos in cellular cultures. A differential response to toxicity was demonstrated between susceptible and partially resistant genotypes, with a good correlation noted between the resistance to the fungus at the whole plant level and resistance at the cellular level to fungal exudates from raw and organic extracts. No toxic reaction of embryogenic cultures was observed after treatment with the aqueous extract or zinniol used at physiological concentration. Moreover, we did not detect zinniol in toxic fungal extracts by UHPLC analysis. These results suggest that strong phytotoxic compounds are present in the organic extract and remain to be characterized. Our results clearly show that carrot tolerance to A. dauci toxins is one component of its partial resistance. PMID:24983469

NM-Scale Anatomy of an Entire Stardust Carrot Track

NASA Technical Reports Server (NTRS)

Nakamura-Messenger, K.; Keller, L. P.; Clemett, S. J.; Messenger, S.

2009-01-01

Comet Wild-2 samples collected by NASA s Stardust mission are extremely complex, heterogeneous, and have experienced wide ranges of alteration during the capture process. There are two major types of track morphologies: "carrot" and "bulbous," that reflect different structural/compositional properties of the impactors. Carrot type tracks are typically produced by compact or single mineral grains which survive essentially intact as a single large terminal particle. Bulbous tracks are likely produced by fine-grained or organic-rich impactors [1]. Owing to their challenging nature and especially high value of Stardust samples, we have invested considerable effort in developing both sample preparation and analytical techniques tailored for Stardust sample analyses. Our report focuses on our systematic disassembly and coordinated analysis of Stardust carrot track #112 from the mm to nm-scale.

The Role of Pectin in Pb Binding by Carrot Peel Biosorbents: Isoterm Adsorption Study

NASA Astrophysics Data System (ADS)

Hastuti, B.; Totiana, F.; Winiasih, R.

2018-04-01

Cheaply and abundantly biosorption available materials such as carrot peels can be a cost-efficient method for removing heavy metals from wastewater. To investigate the role pectin plays in metal binding by carrot peels, commerce pectin was compared. FTIR spectra confirmed the presence of carboxyl and hydroxyl groups in commerce pectin and carrot pectin. Isoterm experiments showed that all materials could remove Pb (II) ion. All of materials binding Pb (II) follow Freundlich models adsorption. The commerce pectin bindsPb (II) by involving energy 16.6 KJ/mole whereas pectin from carrot peel involves energy 21.09 KJ/mole. It indicates that commerce pectin binds the Pb (II) by physics adsorption whereas pectin from carrot peel by physics and chemical adsorption.

Frequent Insect Visitors Are Not Always Pollen Carriers in Hybrid Carrot Pollination.

PubMed

Gaffney, Ann; Bohman, Björn; Quarrell, Stephen R; Brown, Philip H; Allen, Geoff R

2018-06-07

Insect crop visitations do not necessarily translate to carriage or transfer of pollen. To evaluate the potential of the various insects visiting hybrid carrot flowers to facilitate pollen transfer, this study examines insect visitation rates to hybrid carrot seed crops in relation to weather, time of day and season, pollen carrying capacity, inter-row movement, and visitation frequency to male-fertile and male-sterile umbels. The highest pollen loads were carried by nectar scarabs, honey bees, and the hover fly Eristalis tenax (Linnaeus). Honey bees and muscoid flies were observed to forage mostly within the male fertile carrot row while nectar scarabs and E. tenax foraged across rows, carrying equal pollen loads regardless of their distance from the pollen source. All observed insect taxa were more frequently seen visiting male-fertile than male-sterile umbels. In contrast to other visiting insects, honey bees were abundant and frequent visitors and were observed carrying high pollen loads. Consequently, we suggest both optimizing honey bee management and improving the attraction of carrot lines to honey bees to improve pollination rates for hybrid carrot seed crops.

Plant defense genes are regulated by ethylene

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

Ecker, J.R.; Davis, R.W.

One of the earliest detectable events during plant-pathogen interaction is a rapid increase in ethylene biosynthesis. This gaseous plant stress hormone may be a signal for plants to activate defense mechanisms against invading pathogens such as bacteria, fungi, and viruses. The effect of ethylene on four plant genes involved in three separate plant defense response pathways was examined; these included (i and ii) genes that encode L-phenylalanine ammonia-lyase (EC 4.3.1.5) and 4-coumarate:CoA ligase (4-coumarate:CoA ligase (AMP-forming), EC 6.2.1.12), enzymes of the phenylpropanoid pathway, (iii) the gene encoding chalcone synthase, an enzyme of the flavonoid glycoside pathway, and (iv) the genesmore » encoding hydroxyproline-rich glycoprotein, a major protein component(s) of plant cell walls. Blot hybridization analysis of mRNA from ethylene-treated carrot roots reveals marked increases in the levels of phenylalanine ammonia-lyase mRNA, 4-coumarate CoA ligase mRNA, chalcone synthase mRNA, and certain hydroxyproline-rich glycoprotein transcripts. The effect of ethylene on hydroxyproline-rich glycoprotein mRNA accumulation was different from that of wounding. Ethylene induces two hydroxyproline-rich glycoprotein mRNAs (1.8 and 4.0 kilobases), whereas wounding of carrot root leads to accumulation of an additional hydroxyproline-rich mRNA (1.5 kilobases). These results indicate that at least two distinct signals, ethylene and a wound signal, can affect the expression of plant defense-response genes.« less

A novel dehydration technique for carrot slices implementing ultrasound and vacuum drying methods.

PubMed

Chen, Zhi-Gang; Guo, Xiao-Yu; Wu, Tao

2016-05-01

A novel drying technique using a combination of ultrasound and vacuum dehydration was developed to shorten the drying time and improve the quality of carrot slices. Carrot slices were dried with ultrasonic vacuum (USV) drying and vacuum drying at 65 °C and 75 °C. The drying rate was significantly influenced by the drying techniques and temperatures. Compared with vacuum drying, USV drying resulted in a 41-53% decrease in the drying time. The drying time for the USV and vacuum drying techniques at 75 °C was determined to be 140 and 340 min for carrot slices, respectively. The rehydration potential, nutritional value (retention of β-carotene and ascorbic acid), color, and textural properties of USV-dried carrot slices are predominately better compared to vacuum-dried carrot slices. Moreover, lower energy consumption was used in the USV technique. The drying data (time versus moisture ratio) were successfully fitted to Wang and Singh model. Copyright © 2015. Published by Elsevier B.V.

The next generation of carotenoid studies in carrot (Daucus carota L.)

USDA-ARS?s Scientific Manuscript database

Orange carrot (Daucus carota L.) is one of the richest sources of naturally occurring ß-carotene while red and yellow carrot varieties contain large quantities of lycopene and lutein. The human body utilizes carotenoids, particularly ß-carotene (provitamin A) as a precursor for the production of ret...

Chemical and sensory quality of processed carrot puree as influenced by stress-induced phenolic compounds.

PubMed

Talcott, S T; Howard, L R

1999-04-01

Physicochemical analysis of processed strained product was performed on 10 carrot genotypes grown in Texas (TX) and Georgia (GA). Carrots from GA experienced hail damage during growth, resulting in damage to their tops. Measurements included pH, moisture, soluble phenolics, total carotenoids, sugars, organic acids, and isocoumarin (6-MM). Sensory analysis was conducted using a trained panel to evaluate relationships between chemical and sensory attributes of the genotypes and in carrots spiked with increasing levels of 6-MM. Preharvest stress conditions in GA carrots seemed to elicit a phytoalexic response, producing compounds that impacted the perception of bitter and sour flavors. Spiking 6-MM into strained carrots demonstrated the role bitter compounds have in lowering sweetness scores while increasing the perception of sour flavor. Screening fresh carrots for the phytoalexin 6-MM has the potential to significantly improve the sensory quality of processed products.

Incentivizing wellness in the workplace: sticks (not carrots) send stigmatizing signals.

PubMed

Tannenbaum, David; Valasek, Chad J; Knowles, Eric D; Ditto, Peter H

2013-08-01

Companies often provide incentives for employees to maintain healthy lifestyles. These incentives can take the form of either discounted premiums for healthy-weight employees ("carrot" policies) or increased premiums for overweight employees ("stick" policies). In the three studies reported here, we demonstrated that even when stick and carrot policies are formally equivalent, they do not necessarily convey the same information to employees. Stick but not carrot policies were viewed as reflecting negative company attitudes toward overweight employees (Study 1a) and were evaluated especially negatively by overweight participants (Study 1b). This was true even when overweight employees paid less money under the stick than under the carrot policy. When acting as policymakers (Study 2), participants with high levels of implicit overweight bias were especially likely to choose stick policies-often on the grounds that such policies were cost-effective-even when doing so was more costly to the company. Policymakers should realize that the framing of incentive programs can convey tacit, and sometimes stigmatizing, messages.

Assessing the Likelihood of Transmission of Candidatus Liberibacter solanacearum to Carrot by Potato Psyllid, Bactericera cockerelli (Hemiptera: Triozidae)

PubMed Central

Munyaneza, Joseph E.; Mustafa, Tariq; Fisher, Tonja W.; Sengoda, Venkatesan G.; Horton, David R.

2016-01-01

‘Candidatus Liberibacter solanacearum’ (Lso) is a phloem-limited bacterium that severely affects important Solanaceae and Apiaceae crops, including potato, tomato, pepper, tobacco, carrot and celery. This bacterium is transmitted to solanaceous species by potato psyllid, Bactericera cockerelli, and to Apiaceae by carrot psyllids, including Trioza apicalis and Bactericera trigonica. Five haplotypes of Lso have so far been described, two are associated with solanaceous species and potato psyllids, whereas the other three are associated with carrot and celery crops and carrot psyllids. Little is known about cross-transmission of Lso to carrot by potato psyllids or to potato by carrot psyllids. Thus, the present study assessed whether potato psyllid can transmit Lso to carrot and whether Lso haplotypes infecting solanaceous species can also infect carrot and lead to disease symptom development. In addition, the stylet probing behavior of potato psyllid on carrot was assessed using electropenetrography (EPG) technology to further elucidate potential Lso transmission to Apiaceae by this potato insect pest. Results showed that, while potato psyllids survived on carrot for several weeks when confined on the plants under controlled laboratory and field conditions, the insects generally failed to infect carrot plants with Lso. Only three of the 200 carrot plants assayed became infected with Lso and developed characteristic disease symptoms. Lso infection in the symptomatic carrot plants was confirmed by polymerase chain reaction assay and Lso in the carrots was determined to be of the haplotype B, which is associated with solanaceous species. EPG results further revealed that potato psyllids readily feed on carrot xylem but rarely probe into the phloem tissue, explaining why little to no Lso infection occurred during the controlled laboratory and field cage transmission trials. Results of our laboratory and field transmission studies, combined with our EPG results

7 CFR 319.56-43 - Baby corn and baby carrots from Zambia.

Code of Federal Regulations, 2010 CFR

2010-01-01

... § 319.56-43 Baby corn and baby carrots from Zambia. (a) Immature, dehusked “baby” sweet corn (Zea mays L... consignments only. (b) Immature “baby” carrots (Daucus carota L. ssp. sativus) for consumption measuring 10 to...

Formulation and evaluation of carrot seed oil-based cosmetic emulsions.

PubMed

Singh, Shalini; Lohani, Alka; Mishra, Arun Kumar; Verma, Anurag

2018-05-08

The present study deals with the evaluation of antiaging potential of carrot seed oil-based cosmetic emulsions. Briefly, cosmetic emulsions composed of carrot seed oil in varying proportions (2, 4, and 6% w/v) were prepared using the hydrophile-lipophile balance (HLB) technique. Coconut oil, nonionic surfactants (Tween 80 and Span 80), and xanthan gum were used as the oil phase, emulgent, and emulsion stabilizer, respectively. The formed emulsions were evaluated for various physical, chemical, and biochemical parameters such as the zeta potential, globule size measurement, antioxidant activity, sun protection factor (SPF), skin irritation, and biochemical studies. The zeta potential values ranged from -43.2 to -48.3, indicating good stability. The polydispersity index (PDI) of various emulsion formulations ranged from 0.353 to 0.816. 1,1-Diphenyl-2-picrylhydrazyl- (DPPH) and nitric oxide-free radical scavenging activity showed the antioxidant potential of the prepared carrot seed oil emulsions. The highest SPF value (6.92) was shown by F3 having 6%w/v carrot seed oil. Histopathological data and biochemical analysis (ascorbic acid (ASC) and total protein content) suggest that these cosmetic emulsions have sufficient potential to be used as potential skin rejuvenating preparations.

Color changes kinetics during deep fat frying of carrot slice

NASA Astrophysics Data System (ADS)

Salehi, Fakhreddin

2018-05-01

Heat and mass transfer phenomena take place during frying cause physicochemical changes, which affect the colour and surface of the fried products. The effect of frying temperature on the colour changes and heat transfer during deep fat frying of carrot has been investigated. The colour scale parameters redness (a*), yellowness (b*) and lightness (L*), and color change intensity (ΔE) were used to estimate colour changes during frying as a function of oil temperature. L* value of fried carrot decreased during frying. The redness of fried carrot decreased during the early stages of frying, while it increased afterwards (become more red). A first-order kinetic equation was used for each one of the three colour parameters, in which the rate constant is a function of oil temperatures. The results showed that oil temperature has a significant effect on the colour parameters. Different kinetic models were used to fit the experimental data and the results revealed that the quadratic model was the most suitable to describe the color change intensity (ΔE) (R > 0.96). Center temperature of carrot slice increased with increase in oil temperature and time during frying.

Incidence and severity of cavity spot of carrot as affected by pigmentation, temperature, and rainfall

USDA-ARS?s Scientific Manuscript database

Field trials to determine the effect of carrot pigmentation and weather parameters on cavity spot of carrot (CS) were conducted in the Holland/Bradford Marsh region of Ontario between 2002 and 2009. Twenty three colored carrots from the USDA-ARS breeding program at the University of Wisconsin (5) an...

Optimization of physicochemical and textural properties of pizza cheese fortified with soybean oil and carrot extract.

PubMed

Motevalizadeh, Ehsan; Mortazavi, Seyed Ali; Milani, Elnaz; Hooshmand-Dalir, Moosa Al-Reza

2018-03-01

Response surface methodology (RSM) was used to optimize pizza cheese containing carrot extract. The effects of two important independent variables including soybean oil (5%-20%) and carrot extract (5%-20%) were studied on physicochemical and textural properties of pizza cheese containing carrot extract. According to the results, RSM was successfully used for optimizing formulation of pizza cheese containing carrot juice. Results of this study revealed that oil (A), carrot (B), AB, square term of carrot (B 2 ), B, AB, square term of oil (A 2 ), B 2 , AB, AB, A 2 B, A 2 , A 2 , A, A 2 , A 2 , AB, and AB 2 had the most effect on moisture, acidity, stretch, L*, a*, b*, hardness, meltability, springiness, peroxide value (PV), cohesiveness, chewiness, gumminess, fracture force, adhesiveness force, stiffness, flavor, and overall acceptability, respectively. A formulation upon 20% oil and 10.88% carrot extract was found as the optimal formulation for pizza cheese containing carrot extract. At the optimal formulation, PV, L*, a*, b*, meltability, stretch, cohesiveness, springiness, gumminess, chewiness, adhesive force, flavor, texture, and overall acceptability at the optimum formulation were measured 2.23, 82.51, -3.69, 18.05, 17.86, 85.61, 0.41, 7.874, 23.7, 0.27, 0.61, 3.50, 3.95, and 3.65, respectively.

Crotonic acid as a bioactive factor in carrot seeds (Daucus carota L.).

PubMed

Jasicka-Misiak, Izabela; Wieczorek, Piotr P; Kafarski, Paweł

2005-06-01

Water extracts from the carrot seed (Daucus carota L.) var. Perfekcja exhibit plant growth inhibitory properties against cress, cucumber, onion and carrot in a dose-dependant manner. This property results from the action of low-and high-molecular components of the extract. The low-molecular component was identified as crotonic acid ((E)-2-butenoic acid). Its presence was also confirmed in other late varieties of carrot. The determined strong herbicidal properties of crotonic acid and its availability after release to soil combined with its high level in seeds suggest that it might be considered as an allelopathic and autotoxic factor in the seeds.

Descriptive analysis and early-stage consumer acceptance of yogurts fermented with carrot juice.

PubMed

Cliff, M A; Fan, L; Sanford, K; Stanich, K; Doucette, C; Raymond, N

2013-07-01

This research explored the sensory characteristics and consumer acceptance of novel probiotic unsweetened yogurts. Yogurts were made with 4 carrot juice levels (8, 16, 24, and 32%), 2 firmness levels (regular, 45g/L milk solids; firm, 90g/L milk solids), and 2 starter cultures (C1, C2). The sensory profile characterized the color intensity (before and after stirring), carrot flavor, sourness, and 7 texture/mouth-feel attributes (astringency, chalkiness, mouth-coating, thickness, smoothness, creaminess, and graininess). The influence of carrot juice level and firmness level were evaluated using ANOVA, polynomial contrasts, and principal component analysis. Mean scores and standard errors were calculated. Consumer acceptance panels in Wolfville, Nova Scotia (n=56), and in Vancouver, British Columbia (Asian n=72, non-Asian n=72), evaluated the hedonic responses to the C1 and C2 formulations, respectively. We observed increases in color intensity, carrot flavor, creaminess, mouth-coating, and chalkiness with increasing carrot juice levels, as well as increases in color intensity, carrot flavor, creaminess, mouth-coating, thickness, and astringency with increasing milk solids concentrations of the C1 and C2 yogurts. Mean hedonic scores for color, appearance, and texture/mouth-feel were greater than hedonic scores for aroma, flavor/taste, and overall liking. This research identified the sensory qualities that need further development and demonstrated the importance of early-stage consumer acceptance research for directing new product development. Copyright © 2013 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Drinking carrot juice increases total antioxidant status and decreases lipid peroxidation in adults.

PubMed

Potter, Andrew S; Foroudi, Shahrzad; Stamatikos, Alexis; Patil, Bhimanagouda S; Deyhim, Farzad

2011-09-24

High prevalence of obesity and cardiovascular disease is attributable to sedentary lifestyle and eating diets high in fat and refined carbohydrate while eating diets low in fruit and vegetables. Epidemiological studies have confirmed a strong association between eating diets rich in fruits and vegetables and cardiovascular health. The aim of this pilot study was to determine whether drinking fresh carrot juice influences antioxidant status and cardiovascular risk markers in subjects not modifying their eating habits. An experiment was conducted to evaluate the effects of consuming 16 fl oz of daily freshly squeezed carrot juice for three months on cardiovascular risk markers, C-reactive protein, insulin, leptin, interleukin-1α, body fat percentage, body mass index (BMI), blood pressure, antioxidant status, and malondialdehyde production. Fasting blood samples were collected pre-test and 90 days afterward to conclude the study. Drinking carrot juice did not affect (P > 0.1) the plasma cholesterol, triglycerides, Apo A, Apo B, LDL, HDL, body fat percentage, insulin, leptin, interleukin-1α, or C-reactive protein. Drinking carrot juice decreased (P = 0.06) systolic pressure, but did not influence diastolic pressure. Drinking carrot juice significantly (P < 0.05) increased the plasma total antioxidant capacity and decreased (P < 0.05) the plasma malondialdehyde production. Drinking carrot juice may protect the cardiovascular system by increasing total antioxidant status and by decreasing lipid peroxidation independent of any of the cardiovascular risk markers measured in the study.

Elusive carrot: tax incentives for R and D

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

Brown, K.M.

The 1981 Economic Recovery Tax Act created a tax credit for firms that increased their research and development (R and D) spending, but this provision has had some unintended consequences because it provided a carrot that was no longer needed and which could actually speed the decline of R and D efforts. The credit is having little impact on long-term research planning because it expires at the end of 1985. When Congress decides whether to extend, modify, or drop the credit, it would do better to replace the carrot and stick approach with a stable, predictable policy environment that willmore » benefit industrial innovation.« less

Assessing the impact of geogenic chromium uptake by carrots (Daucus carota) grown in Asopos river basin.

PubMed

Lilli, Maria A; Syranidou, Evdokia; Palliou, Andriana; Nikolaidis, Nikolaos P; Karatzas, George; Kalogerakis, Nicolas

2017-01-01

A methodology was developed to assess the impact of geogenic origin hexavalent chromium uptake by carrots, and the risk of human consumption of carrots grown in Asopos River basin in Greece. A field scale experiment was conducted with carrots cultivated in treatment plots, with and without compost amendment, in order to assess the impact of carbon in the mobility and uptake of chromium by plants. The results suggested that there is a trend for chromium mobilization and uptake in the surface and the leaves of the carrots cultivated in the treatment plot with the higher carbon addition, but not in the core of the carrots. Limited mobility of hexavalent chromium in the soil-plant-water system is presented due to the affinity of chromium to be retained in the solid phase and be uptaken by plants. Hexavalent chromium tolerant bacterial strains were isolated from the carrots. These endophytic bacteria, present in all parts of the plant, were able to reduce hexavalent chromium to trivalent form to levels below the detection limit. Finally, a site-specific risk assessment analysis suggested no adverse effects to human health due to the consumption of carrots. These findings are of particular importance since they confirm that carrots grown in soils with geogenic origin chromium does not pose any adverse risk for human consumption, but could also have the beneficial effect of the micronutrient trivalent chromium. Copyright © 2016 Elsevier Inc. All rights reserved.

New carrot and garlic germplasm to advance breeding and understand crop origins

USDA-ARS?s Scientific Manuscript database

The genetic variation provided by diverse plant germplasm is the basic building material used for crop improvement that shapes the crops we grow today. Wild carrot from the U.S. provided the cytoplasm used to develop a reliable system to produce hybrid carrots that account for most of the commercial...

Molecular characterization and pathogenicity of a carrot (Daucus carota) infecting begomovirus and associated betasatellite from India.

PubMed

Kumar, Jitendra; Gunapati, Samatha; Singh, Sudhir P; Gadre, Rekha; Sharma, Naresh C; Tuli, Rakesh

2013-10-24

The yellow mosaic pattern and shortening of leaf petiole are common disease symptoms associated with begomovirus infection in carrot. DNA from field infected carrot leaves was analyzed by rolling circle amplification and sequencing. The results established the presence of ageratum enation virus (AEV), which is referred to here as ageratum enation virus-carrot (AEV-Car). Symptomatic ageratum (Ageratum conyzoides) plants, growing adjacent to the carrot fields, also showed the presence of AEV (AEV-Age). Ageratum yellow leaf curl betasatellite (AYLCB) was also detected in the AEV infected carrot and ageratum samples. AEV-Car and AEV-Age are 95-97% identical in their DNA sequences, represents groups of isolates from the respective plant hosts (carrot and ageratum). Agroinoculation using infectious clones of AEV-Car plus AYLCB or AEV-Age plus AYLCB in carrot, ageratum, tobacco (Nicotiana tabacum) and tomato (Solanum lycopersicum) produced yellow mosaic and curling symptoms in leaves of inoculated plants. Agroinoculation of the two isolates together, along with the betasatellite (AEV-Car plus AEV-Age plus AYLCB) resulted in the enhancement of symptoms in comparison to the plants inoculated with single isolate. Plants with more severe symptoms showed a higher level of viral DNA accumulation, suggesting synergistic interactions between the two isolates of AEV. Copyright © 2013. Published by Elsevier B.V.

Effects of Cover Crops on Pratylenchus penetrans and the Nematode Community in Carrot Production

PubMed Central

Grabau, Zane J.; Zar Maung, Zin Thu; Noyes, D. Corey; Baas, Dean G.; Werling, Benjamin P.; Brainard, Daniel C.; Melakeberhan, Haddish

2017-01-01

Cover cropping is a common practice in U.S. Midwest carrot production for soil conservation, and may affect soil ecology and plant-parasitic nematodes—to which carrots are very susceptible. This study assessed the impact of cover crops—oats (Avena sativa), radish (Raphanus sativus) cv. Defender, rape (Brassica napus) cv. Dwarf Essex, and a mixture of oats and radish—on plant-parasitic nematodes and soil ecology based on the nematode community in Michigan carrot production systems. Research was conducted at two field sites where cover crops were grown in Fall 2014 preceding Summer 2015 carrot production. At Site 1, root-lesion (Pratylenchus penetrans) and stunt (Tylenchorhynchus sp.) nematodes were present at low population densities (less than 25 nematodes/100 cm3 soil), but were not significantly affected (P > 0.05) by cover crops. At Site 2, P. penetrans population densities were increased (P ≤ 0.05) by ‘Defender’ radish compared to other cover crops or fallow control during cover crop growth and midseason carrot production. At both sites, there were few short-term impacts of cover cropping on soil ecology based on the nematode community. At Site 1, only at carrot harvest, radish-oats mixture and ‘Dwarf Essex’ rape alone enriched the soil food web based on the enrichment index (P ≤ 0.05) while rape and radish increased structure index values. At Site 2, bacterivore abundance was increased by oats or radish cover crops compared to control, but only during carrot production. In general, cover crops did not affect the nematode community until nearly a year after cover crop growth suggesting that changes in the soil community following cover cropping may be gradual. PMID:28512383

Unique N-Glycan Moieties of the 66-kDa Cell Wall Glycoprotein from the Red Microalga Porphyridium sp.

PubMed Central

Levy-Ontman, Oshrat; Arad, Shoshana (Malis); Harvey, David J.; Parsons, Thomas B.; Fairbanks, Antony; Tekoah, Yoram

2011-01-01

We report here the structural determination of the N-linked glycans in the 66-kDa glycoprotein, part of the unique sulfated complex cell wall polysaccharide of the red microalga Porphyridium sp. Structures were elucidated by a combination of normal phase/reverse phase HPLC, positive ion MALDI-TOF MS, negative ion electrospray ionization, and MS/MS. The sugar moieties of the glycoprotein consisted of at least four fractions of N-linked glycans, each composed of the same four monosaccharides, GlcNAc, Man, 6-O-MeMan, and Xyl, with compositions Man8–9Xyl1–2Me3GlcNAc2. The present study is the first report of N-glycans with the terminal Xyl attached to the 6-mannose branch of the 6-antenna and to the 3-oxygen of the penultimate (core) GlcNAc. Another novel finding was that all four glycans contain three O-methylmannose residues in positions that have never been reported before. Although it is known that some lower organisms are able to methylate terminal monosaccharides in glycans, the present study on Porphyridium sp. is the first describing an organism that is able to methylate non-terminal mannose residues. This study will thus contribute to understanding of N-glycosylation in algae and might shed light on the evolutionary development from prokaryotes to multicellular organisms. It also may contribute to our understanding of the red algae polysaccharide formation. The additional importance of this research lies in its potential for biotechnological applications, especially in evaluating the use of microalgae as cell factories for the production of therapeutic proteins. PMID:21515680

First report of 'Candidatus Liberibacter solanacearum' associated with psyllid-affected carrots in Norway

USDA-ARS?s Scientific Manuscript database

Carrot (Daucus carota) plants with symptoms resembling those associated with the carrot psyllid Trioza apicalis and the bacterium “Candidatus Liberibacter solanacearum” were observed in 70-80% of commercial fields and experimental plots in southeastern Norway from late July to mid-September 2011; al...

First report of 'Candidatus Liberibacter solanacearum' associated with psyllid-affected carrots in Sweden

USDA-ARS?s Scientific Manuscript database

Carrot (Daucus carota) plants with symptoms resembling those of the carrot psyllid Trioza apicalis and “Candidatus Liberibacter solanacearum” damage were observed in 70% of commercial fields in southern Sweden in August 2011; all cultivars grown were affected, at about 1 to 45% symptomatic plants pe...

Relationship between carrot weevil infestation and parsley yield.

PubMed

Torres, Angel N; Hoy, Casey W

2005-08-01

The relationship between numbers of carrot weevil, Listronotus oregonensis (LeConte), oviposition scars and parsley fresh weight and plant mortality was measured in research plots during 1999 and 2000. Fresh weight was measured in one to two cuttings of parsley planted on two planting dates. The average weight declined with increasing numbers of oviposition scars in the later planting in 1999. Compensatory growth in surviving plants may reduce this effect. Plant mortality increased as number of oviposition scars per plant increased in the second planting in both years and in the first cutting of the first planting in 2000. One oviposition scar per plant is sufficient to result in significant reduction in fresh weight per plant. In commercial parsley fields, the relationship between fresh weight of parsley per 30-cm row section of parsley was best described as a linear function of the proportion of plants with root feeding. Economic damage to parsley that is equivalent to the cost of controlling carrot weevil was estimated to result from approximately 1% of plants with root damage. Based upon this estimated economic injury level, we suggest an action threshold of 1% of plants containing carrot weevil oviposition scars earlier in the growing season when controls could be applied to prevent the damage.

Beyond Carrots and Sticks: Toward a Transformative Model of Division I Athletics

ERIC Educational Resources Information Center

Clubb, Sandy Hatfield

2012-01-01

The old expression about the carrot and the stick, which refers to the application of reward and punishment to induce action, dates back to the days when pack mules were used for transportation. The mules would move toward carrots that dangled just ahead of them--and move all the faster because they feared drivers with sticks behind them. In 2012,…

7 CFR 319.56-43 - Baby corn and baby carrots from Zambia.

Code of Federal Regulations, 2011 CFR

2011-01-01

... 7 Agriculture 5 2011-01-01 2011-01-01 false Baby corn and baby carrots from Zambia. 319.56-43... § 319.56-43 Baby corn and baby carrots from Zambia. (a) Immature, dehusked “baby” sweet corn (Zea mays L..., which is a field, where the corn has been grown must have been inspected at least once during the...

Quality of fresh and stored carrots depending on iodine and nitrogen fertilization.

PubMed

Smoleń, Sylwester; Sady, Włodzimierz; Ledwożyw-Smoleń, Iwona; Strzetelski, Piotr; Liszka-Skoczylas, Marta; Rożek, Stanisław

2014-09-15

Iodine is an important mineral nutrient essential for a proper functioning of human and animal organism. Despite current programmes of iodine prophylaxis (mainly based on salt iodization) approximately 30-38% of human population has insufficient iodine intake. Crop plants can become an efficient vector of this element in the food chain. Iodine is not a nutrient for plants. For that reason, in addition to determining the possibility of increasing iodine content in crop plant it is necessary to describe its impact on yield quality. The aim of the study was to analyze the influence of soil fertilization with iodine and nitrogen on the quality of carrot roots and its storage ability. In 2008-2010 the field study with carrot cv. 'Kazan F1' was conducted. A differential soil fertilization with iodine (in the form of I(-) or IO3(-)) and nitrogen (as NO3(-) or NH4(+)) was applied in the experiment: (1) control without N and I, (2) KI application without N, (3) KIO3 application without N, (4) KI+Ca(NO3)2, (5) KIO3+Ca(NO3)2, (6) KI+(NH4)2SO4 and (7) KIO3+(NH4)2SO4. The experiment was arranged in a split-plot design. Iodine (in both forms) was applied pre-sowing in a dose of 2 kg I ha(-1). Nitrogen in the form of Ca(NO3)2 and (NH4)2SO4 was introduced pre-sowing and as a top dressing, each dose of 100 kg N ha(-1). A diverse, statistically significant influence of tested factors on the activity of free radical-scavenging (DPPH) and the content of: dry matter, glucose, fructose, sucrose, total soluble sugars, soluble solids - Brix %, phenolic compounds, phenylpropanoids, flavonols, anthocyanins and carotenoids was noted in carrot roots directly after the harvest as well as at the end of four-month storage. Iodine applied with relatively high doses of nitrogen decreased the quality of fresh carrot. After storage, opposite relations were noted for tested combinations (with I and N application) with respect to carrot quality when compared to results obtained after the harvest. The

Production and characterization of curcumin microcrystals and evaluation of the antimicrobial and sensory aspects in minimally processed carrots.

PubMed

Silva, Anderson Clayton da; Santos, Priscila Dayane de Freitas; Palazzi, Nicole Campezato; Leimann, Fernanda Vitória; Fuchs, Renata Hernandez Barros; Bracht, Lívia; Gonçalves, Odinei Hess

2017-05-24

Nontoxic conserving agents are in demand by the food industry due to consumers concern about synthetic conservatives, especially in minimally processed food. The antimicrobial activity of curcumin, a natural phenolic compound, has been extensively investigated but hydrophobicity is an issue when applying curcumin to foodstuff. The objective of this work was to evaluate curcumin microcrystals as an antimicrobial agent in minimally processed carrots. The antimicrobial activity of curcumin microcrystals was evaluated in vitro against Gram-positive (Bacillus cereus and Staphylococcus aureus) and Gram-negative (Escherichia coli and Pseudomonas aeruginosa) microorganisms, showing a statistically significant (p < 0.05) decrease in the minimum inhibitory concentration compared to in natura, pristine curcumin. Curcumin microcrystals were effective in inhibiting psychrotrophic and mesophile microorganisms in minimally processed carrots. Sensory analyses were carried out showing no significant difference (p < 0.05) between curcumin microcrystal-treated carrots and non-treated carrots in triangular and tetrahedral discriminative tests. Sensory tests also showed that curcumin microcrystals could be added as a natural preservative in minimally processed carrots without causing noticeable differences that could be detected by the consumer. One may conclude that the analyses of the minimally processed carrots demonstrated that curcumin microcrystals are a suitable natural compound to inhibit the natural microbiota of carrots from a statistical point of view.

Survey of 'Candidatus Liberibacter solanacearum' in carrot crops affected by the psyllid Trioza apicalis (Hemiptera: Triozidae) in Norway

USDA-ARS?s Scientific Manuscript database

The carrot psyllid Trioza apicalis Förster (Hemiptera: Triozidae) is a serious insect pest of carrot (Daucus carota L.) in northern Europe, where it can cause up to 100% crop loss. Although it was long believed that T. apicalis causes damage to carrot by injection of toxins into the plant, it was re...

Effectiveness of some substances in the control of carrot and parsley roots against fungal diseases.

PubMed

Nawrocki, Jacek

2007-01-01

Field experiments were carried out in the years 2005 and 2006 on carrot cv. 'Koral' and 'Perfekcja', and parsley cv. 'Berlinska' and 'Cukrowa'. Effectiveness of substances: Biochikol 020 PC (biologically active substances BAS--chitosan 20 g/dm3), Bioczos BR (extract of garlic 10 g/1 brick) and Biosept 33 SL (extract of grapefruit 33%) on seedling roots of carrot and parsley was studied. As the standard fungicide Zaprawa Funaben T (carbendazim 20% + tiuram 45%) was used. Roots of carrot and parsley were treated one of tested substances spring immediately before planting seedling roots. During vegetation period the growth of seedling shoots and setting of seeds, and their infestation by fungal and bacterial pathogens was noticed. Among substances used for spring dressing of carrot and also parsley seedling roots, the best efficacy exhibited Zaprawa Funaben T in both years of observation. The highest yield of carrot seeds had combination roots cv. 'Koral' and parsley seeds roots cvs 'Berlińska' and 'Cukrowa' dressed Zaprawa Funaben T. Effectiveness of biopreparates Biochikol and Biosept was lower in comparison with the standard fungicide, but their protective effect was significantly higher than in control. Bioczos had the lowest control efficacy.

Morphological characteristics, anatomical structure, and gene expression: novel insights into gibberellin biosynthesis and perception during carrot growth and development.

PubMed

Wang, Guang-Long; Xiong, Fei; Que, Feng; Xu, Zhi-Sheng; Wang, Feng; Xiong, Ai-Sheng

2015-01-01

Gibberellins (GAs) are considered potentially important regulators of cell elongation and expansion in plants. Carrot undergoes significant alteration in organ size during its growth and development. However, the molecular mechanisms underlying gibberellin accumulation and perception during carrot growth and development remain unclear. In this study, five stages of carrot growth and development were investigated using morphological and anatomical structural techniques. Gibberellin levels in leaf, petiole, and taproot tissues were also investigated for all five stages. Gibberellin levels in the roots initially increased and then decreased, but these levels were lower than those in the petioles and leaves. Genes involved in gibberellin biosynthesis and signaling were identified from the carrotDB, and their expression was analyzed. All of the genes were evidently responsive to carrot growth and development, and some of them showed tissue-specific expression. The results suggested that gibberellin level may play a vital role in carrot elongation and expansion. The relative transcription levels of gibberellin pathway-related genes may be the main cause of the different bioactive GAs levels, thus exerting influences on gibberellin perception and signals. Carrot growth and development may be regulated by modification of the genes involved in gibberellin biosynthesis, catabolism, and perception.

Soluble glycoprotein VI dimer inhibits platelet adhesion and aggregation to the injured vessel wall in vivo.

PubMed

Massberg, Steffen; Konrad, Ildiko; Bültmann, Andreas; Schulz, Christian; Münch, Götz; Peluso, Mario; Lorenz, Michael; Schneider, Simon; Besta, Felicitas; Müller, Iris; Hu, Bin; Langer, Harald; Kremmer, Elisabeth; Rudelius, Martina; Heinzmann, Ulrich; Ungerer, Martin; Gawaz, Meinrad

2004-02-01

Platelet-collagen interactions play a fundamental role in the process of arterial thrombosis. The major platelet collagen receptor is the glycoprotein VI (GPVI). Here, we determined the effects of a soluble dimeric form of GPVI on platelet adhesion in vitro and in vivo. We fused the extracellular domain of GPVI with the human immunoglobulin Fc domain. The soluble dimeric form of GPVI (GPVI-Fc) specifically bound to immobilized collagen. Binding of GPVI-Fc to collagen was inhibited competitively by soluble GPVI-Fc, but not control Fc lacking the external GPVI domain. GPVI-Fc inhibited the adhesion of CHO cells that stably express human GPVI and of platelets on collagen and attenuated thrombus formation under shear conditions in vitro. To test the effects of GPVI-Fc in vivo, arterial thrombosis was induced in the mouse carotid artery, and platelet-vessel wall interactions were visualized by intravital fluorescence microscopy. Infusion of GPVI-Fc but not of control Fc virtually abolished stable arrest and aggregation of platelets following vascular injury. Importantly, GPVI-Fc but not control Fc, was detected at areas of vascular injury. These findings further substantiate the critical role of the collagen receptor GPVI in the initiation of thrombus formation at sites of vascular injury and identify soluble GPVI as a promising antithrombotic strategy.

Effect of blanching in water and sugar solutions on texture and microstructure of sliced carrots.

PubMed

Neri, Lilia; Hernando, Isabel Hernando; Pérez-Munuera, Isabel; Sacchetti, Giampiero; Pittia, Paola

2011-01-01

Thermal processing of vegetables has pronounced effects on the cell structure, often negatively affecting the final textural properties of the product. In order to study the effect of thermal processing and the protective effect of sugars on the tissue, sliced carrots were subjected to blanching treatments under different time and temperature combinations both in water and in 4% sugar solutions made of trehalose or maltose. The influence of these process conditions on mass transfer, texture, and microstructure (Cryo-scanning electron microscopy) was thus investigated. The total mass loss of all the samples blanched in water was associated to their cook value (C(100)(18)) except for the overprocessed one (90 °C, 10 min) that showed a total mass change significantly lower due to water uptake. The use of trehalose and maltose in the blanching solution reduced the solute loss while increasing the water loss. Microstructural analysis of the differently blanched carrots showed detachments between adjacent cell walls as well as plasmolysis phenomena as the time and temperature of the thermal treatment were increased. A protective effect of both sugars on cell structures was observed mostly in the sample treated at 90 °C. At macroscopic level, textural changes upon blanching were observed by a penetration test. As blanching time was increased, samples processed at 75 °C showed a hardness increase, while those processed at 90 °C showed a hardness decrease. However, both trehalose and maltose did not exert significant effects on the textural properties of blanched carrots when compared with those blanched in water. Practical Application: The results of this study could offer interesting perspectives in the optimization of the heat treatments in order to preserve the quality of semi-finished processed vegetables. Furthermore, the microstructural analysis is nowadays an important investigation tool that could contribute to a deeper understanding of both the effects of

Time-temperature dependent variations in beta-carotene contents in carrot using different spectrophotometric techniques

NASA Astrophysics Data System (ADS)

Ullah, Rahat; Khan, Saranjam; Shah, Attaullah; Ali, Hina; Bilal, Muhammad

2018-05-01

The current study presents time dependent variations in the concentration of beta-carotene in carrot under different storage-temperature conditions using UV–VIS and Raman spectrophotometric techniques. The UV–VIS absorption spectra of beta-carotene extracted from carrot shows three distinct absorption peaks at 442, 467, and 500 nm with maximum absorption at 467 nm. These absorption peaks are very much reproducible and are assigned to β-carotene. Similarly, Raman spectra of carrot samples also confirmed the three main Raman peaks of beta-carotene at shift positions 1003, 1150, and 1515 cm‑1. An overall decrease in beta-carotene content has been observed for time-temperature conditions. These results depict a decrease of about 40% in the content of beta-carotene when carrot samples were stored in a refrigerator (4 °C) for the first 20 d, whereas a decrease of about 25% was observed when carrot samples were stored in a freezer (‑16 °C) for the same period. The objective of this study is to investigate the possible use of Raman spectroscopy and UV–VIS spectroscopy for quick and detailed analysis of changes (degradation) in beta-carotene content associated with time and temperature in storage (frozen foods) in order to promote quality foods for consumers. Future study with a greater focus on the concentration/content of beta-carotene in other fruits/vegetables is also desirable.

Shelf-life extension of minimally processed carrots by gaseous chlorine dioxide.

PubMed

Gómez-López, V M; Devlieghere, F; Ragaert, P; Debevere, J

2007-05-10

Chlorine dioxide (ClO(2)) gas is a strong oxidizing and sanitizing agent that has a broad and high biocidal effectiveness and big penetration ability; its efficacy to prolong the shelf-life of a minimally processed (MP) vegetable, grated carrots (Daucus carota L.), was tested in this study. Carrots were sorted, their ends removed, hand peeled, cut, washed, spin dried and separated in 2 portions, one to be treated with ClO(2) gas and the other to remain untreated for comparisons. MP carrots were decontaminated in a cabinet at 91% relative humidity and 28 degrees C for up to 6 min, including 30 s of ClO(2) injection to the cabinet, then stored under equilibrium modified atmosphere (4.5% O(2), 8.9% CO(2), 86.6% N(2)) at 7 degrees C for shelf-life studies. ClO(2) concentration in the cabinet rose to 1.33 mg/l after 30 s of treatment, and then fell to nil before 6 min. The shelf-life study included: O(2) and CO(2) headspace concentrations, microbiological quality (mesophilic aerobic bacteria, psychrotrophs, lactic acid bacteria, and yeasts), sensory quality (odour, flavour, texture, overall visual quality, and white blushing), and pH. ClO(2) did not affect respiration rate of MP carrots significantly (alpha< or =0.05), and lowered the pH significantly (alpha< or =0.05). The applied packaging configuration kept O(2) headspace concentrations in treated samples in equilibrium and prevented CO(2) accumulation. After ClO(2) treatment, the decontamination levels (log CFU/g) achieved were 1.88, 1.71, 2.60, and 0.66 for mesophilic aerobic bacteria, psychrotrophs, and yeasts respectively. The initial sensory quality of MP carrots was not impaired significantly (alpha< or =0.05). A lag phase of at least 2 days was observed for mesophilic aerobic bacteria, psychrotrophs, and lactic acid bacteria in treated samples, while mesophilic aerobic bacteria and psychrotrophs increased parallelly. Odour was the only important attribute in sensory deterioration, but it reached an

Extracts from black carrot tissue culture as potent anticancer agents.

PubMed

Sevimli-Gur, Canan; Cetin, Burcu; Akay, Seref; Gulce-Iz, Sultan; Yesil-Celiktas, Ozlem

2013-09-01

Black carrots contain anthocyanins possessing enhanced physiological activities. Explants of young black carrot shoots were cultured in Murashige and Skoog (MS) medium for callus initiation and were transferred to new MS medium supplemented with four different combinations of 2,4-dichlorophenoxyacetic acid and kinetin. Subsequently, the lyophilized calli and black carrot harvested from fields were subjected to ultrasound extraction with ethanol at a ratio of 1:15 (w:v). Obtained extracts were applied to various human cancer cell lines including MCF-7 SK-BR-3 and MDA-MB-231 (human breast adenocarcinomas), HT-29 (human colon adenocarcinoma), PC-3 (human prostate adenocarcinoma), Neuro 2A (Musmusculus neuroblastoma) cancer cell lines and VERO (African green monkey kidney) normal cell line by MTT assay. The highest cytotoxic activity was achieved against Neuro-2A cell lines exhibiting viability of 38-46% at 6.25 μg/ml concentration for all calli and natural extracts. However, a significantly high IC50 value of 170.13 μg/ml was attained in normal cell line VERO indicating that its natural counterpart is an ideal candidate for treatment of brain cancer without causing negative effects to normal healthy cells.

Expression profiles of genes involved in jasmonic acid biosynthesis and signaling during growth and development of carrot.

PubMed

Wang, Guanglong; Huang, Wei; Li, Mengyao; Xu, Zhisheng; Wang, Feng; Xiong, Aisheng

2016-09-01

Jasmonates (JAs) are recognized as essential regulators in response to environmental stimuli and plant development. Carrot is an Apiaceae vegetable with great value and undergoes significant size changes over the course of plant growth. However, JA accumulation and its potential roles in carrot growth remain unclear. Here, methyl JA (MeJA) levels and expression profiles of JA-related genes were analyzed in carrot roots and leaves at five developmental stages. MeJA levels in the roots and leaves were the highest at the first stage and decreased as carrot growth proceeded. Transcript levels of several JA-related genes (Dc13-LOX1, Dc13-LOX2, DcAOS, DcAOC, DcOPR2, DcOPR3, DcOPCL1, DcJAR1, DcJMT, DcCOI1, DcJAZ1, DcJAZ2, DcMYC2, DcCHIB/PR3, DcLEC, and DcVSP2) were not well correlated with MeJA accumulation during carrot root and leaf development. In addition, some JA-related genes (DcJAR1, DcJMT, DcCOI1, DcMYC2, and DcVSP2) showed differential expression between roots and leaves. These results suggest that JAs may regulate carrot plant growth in stage-dependent and organ-specific manners. Our work provides novel insights into JA accumulation and its potential roles during carrot growth and development. © The Author 2016. Published by Oxford University Press on behalf of the Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Morphological characteristics, anatomical structure, and gene expression: novel insights into gibberellin biosynthesis and perception during carrot growth and development

PubMed Central

Wang, Guang-Long; Xiong, Fei; Que, Feng; Xu, Zhi-Sheng; Wang, Feng; Xiong, Ai-Sheng

2015-01-01

Gibberellins (GAs) are considered potentially important regulators of cell elongation and expansion in plants. Carrot undergoes significant alteration in organ size during its growth and development. However, the molecular mechanisms underlying gibberellin accumulation and perception during carrot growth and development remain unclear. In this study, five stages of carrot growth and development were investigated using morphological and anatomical structural techniques. Gibberellin levels in leaf, petiole, and taproot tissues were also investigated for all five stages. Gibberellin levels in the roots initially increased and then decreased, but these levels were lower than those in the petioles and leaves. Genes involved in gibberellin biosynthesis and signaling were identified from the carrotDB, and their expression was analyzed. All of the genes were evidently responsive to carrot growth and development, and some of them showed tissue-specific expression. The results suggested that gibberellin level may play a vital role in carrot elongation and expansion. The relative transcription levels of gibberellin pathway-related genes may be the main cause of the different bioactive GAs levels, thus exerting influences on gibberellin perception and signals. Carrot growth and development may be regulated by modification of the genes involved in gibberellin biosynthesis, catabolism, and perception. PMID:26504574

Insights into the Evolution of Hydroxyproline-Rich Glycoproteins from 1000 Plant Transcriptomes.

PubMed

Johnson, Kim L; Cassin, Andrew M; Lonsdale, Andrew; Wong, Gane Ka-Shu; Soltis, Douglas E; Miles, Nicholas W; Melkonian, Michael; Melkonian, Barbara; Deyholos, Michael K; Leebens-Mack, James; Rothfels, Carl J; Stevenson, Dennis W; Graham, Sean W; Wang, Xumin; Wu, Shuangxiu; Pires, J Chris; Edger, Patrick P; Carpenter, Eric J; Bacic, Antony; Doblin, Monika S; Schultz, Carolyn J

2017-06-01

The carbohydrate-rich cell walls of land plants and algae have been the focus of much interest given the value of cell wall-based products to our current and future economies. Hydroxyproline-rich glycoproteins (HRGPs), a major group of wall glycoproteins, play important roles in plant growth and development, yet little is known about how they have evolved in parallel with the polysaccharide components of walls. We investigate the origins and evolution of the HRGP superfamily, which is commonly divided into three major multigene families: the arabinogalactan proteins (AGPs), extensins (EXTs), and proline-rich proteins. Using motif and amino acid bias, a newly developed bioinformatics pipeline, we identified HRGPs in sequences from the 1000 Plants transcriptome project (www.onekp.com). Our analyses provide new insights into the evolution of HRGPs across major evolutionary milestones, including the transition to land and the early radiation of angiosperms. Significantly, data mining reveals the origin of glycosylphosphatidylinositol (GPI)-anchored AGPs in green algae and a 3- to 4-fold increase in GPI-AGPs in liverworts and mosses. The first detection of cross-linking (CL)-EXTs is observed in bryophytes, which suggests that CL-EXTs arose though the juxtaposition of preexisting SP n EXT glycomotifs with refined Y-based motifs. We also detected the loss of CL-EXT in a few lineages, including the grass family (Poaceae), that have a cell wall composition distinct from other monocots and eudicots. A key challenge in HRGP research is tracking individual HRGPs throughout evolution. Using the 1000 Plants output, we were able to find putative orthologs of Arabidopsis pollen-specific GPI-AGPs in basal eudicots. © 2017 American Society of Plant Biologists. All Rights Reserved.

Defining glycoprotein cancer biomarkers by MS in conjunction with glycoprotein enrichment.

PubMed

Song, Ehwang; Mechref, Yehia

2015-01-01

Protein glycosylation is an important and common post-translational modification. More than 50% of human proteins are believed to be glycosylated to modulate the functionality of proteins. Aberrant glycosylation has been correlated to several diseases, such as inflammatory skin diseases, diabetes mellitus, cardiovascular disorders, rheumatoid arthritis, Alzheimer's and prion diseases, and cancer. Many approved cancer biomarkers are glycoproteins which are not highly abundant proteins. Therefore, effective qualitative and quantitative assessment of glycoproteins entails enrichment methods. This chapter summarizes glycoprotein enrichment methods, including lectin affinity, immunoaffinity, hydrazide chemistry, hydrophilic interaction liquid chromatography, and click chemistry. The use of these enrichment approaches in assessing the qualitative and quantitative changes of glycoproteins in different types of cancers are presented and discussed. This chapter highlights the importance of glycoprotein enrichment techniques for the identification and characterization of new reliable cancer biomarkers.

Generation of the heterodimeric precursor GP3 of the Chlamydomonas cell wall.

PubMed

Voigt, Jürgen; Kiess, Michael; Getzlaff, Rita; Wöstemeyer, Johannes; Frank, Ronald

2010-09-01

The cell wall of the unicellular green alga Chlamydomonas reinhardtii exclusively consists of hydroxyproline-containing glycoproteins. Protein chemical analysis of its polypeptide constituents was hindered by their cross-linking via peroxidase-catalysed intermolecular isodityrosine formation and transaminase-dependent processes. To overcome this problem, we have identified putative soluble precursors using polyclonal antibodies raised against deglycosylation products of the highly purified insoluble wall fraction and analysed their amino acid sequence. The occurrence of the corresponding polypeptide in the insoluble glycoprotein framework was finally probed by epitope mapping of the polyclonal antibodies using overlapping scan peptides which, together, cover the whole amino acid sequence of the putative precursor. As a control, peptide fragments released from the insoluble wall fraction by trypsin treatment were analysed by mass spectroscopy. By this approach, the heterodimeric, chaotrope-soluble glycoprotein GP3 proved to be a constituent of the insoluble extracellular matrix of Chlamydomonas reinhardtii. Furthermore, we have shown that the polypeptide backbones of both GP3 subunits are encoded by the same gene and differ by a C-terminal truncation in the case of GP3A. © 2010 Blackwell Publishing Ltd.

First report of bacterial blight of carrot in Indiana caused by Xanthomonas hortorum pv. carotae

USDA-ARS?s Scientific Manuscript database

In summer 2012, bacterial blight symptoms were observed on leaves of carrot plants in 7 out of 70 plots of carrot breeding lines at the Purdue University Meig Horticulture Research Farm, Lafayette, IN. Symptoms included small to large, variably shaped, water soaked to dry, necrotic lesions, with or ...

The carrot genome provides insights into crop origins and a foundation for future crop improvement

USDA-ARS?s Scientific Manuscript database

The sequencing of the carrot genome was an effort that formally began in 2012 and culminated with the publication and release of the genome in 2016. A full genome sequence provides the ultimate foundation to study genetics, gene function, and evolution of a species. The primary goal of the carrot ge...

Assessment of allelopathic properties of Aloe ferox Mill. on turnip, beetroot and carrot.

PubMed

Arowosegbe, Sunday; Afolayan, Anthony J

2012-01-01

Turnip (Brassica rapa var. rapa L.), beetroot (Beta vulgaris L.) and carrot (Daucus carota L.) are common vegetables in South Africa. The allelopathic potential of aqueous leaf and root extracts of Aloe ferox Mill.- a highly valued medicinal plant- was evaluated against seed germination and seedling growth of the three vegetables in Petri dish experiments. The extracts were tested at concentrations of 2, 4, 6, 8, and 10 mg/mL. Leaf extract concentrations above 4 mg/mL inhibited the germination of all the crops, while the root extract had no significant effect on germination irrespective of concentration. Interestingly, the lowest concentration of leaf extract stimulated root length elongation of beetroot by 31.71%. Other concentrations significantly inhibited both root and shoot growth of the vegetable crops except the turnip shoot. The most sensitive crop was carrot, with percentage inhibition ranging from 29.15 to 100% for root and shoot lengths. Lower percentage inhibition was observed for the root extract than the leaf extract against shoot growth of beetroot and carrot. The results from this study suggested the presence of allelochemicals mostly in the leaves of A. ferox that could inhibit the growth of the turnip, beetroot and carrot.

Forcible destruction of severely misfolded mammalian glycoproteins by the non-glycoprotein ERAD pathway

PubMed Central

Ninagawa, Satoshi; Okada, Tetsuya; Sumitomo, Yoshiki; Horimoto, Satoshi; Sugimoto, Takehiro; Ishikawa, Tokiro; Takeda, Shunichi; Yamamoto, Takashi; Suzuki, Tadashi; Kamiya, Yukiko

2015-01-01

Glycoproteins and non-glycoproteins possessing unfolded/misfolded parts in their luminal regions are cleared from the endoplasmic reticulum (ER) by ER-associated degradation (ERAD)-L with distinct mechanisms. Two-step mannose trimming from Man9GlcNAc2 is crucial in the ERAD-L of glycoproteins. We recently showed that this process is initiated by EDEM2 and completed by EDEM3/EDEM1. Here, we constructed chicken and human cells simultaneously deficient in EDEM1/2/3 and analyzed the fates of four ERAD-L substrates containing three potential N-glycosylation sites. We found that native but unstable or somewhat unfolded glycoproteins, such as ATF6α, ATF6α(C), CD3-δ–ΔTM, and EMC1, were stabilized in EDEM1/2/3 triple knockout cells. In marked contrast, degradation of severely misfolded glycoproteins, such as null Hong Kong (NHK) and deletion or insertion mutants of ATF6α(C), CD3-δ–ΔTM, and EMC1, was delayed only at early chase periods, but they were eventually degraded as in wild-type cells. Thus, higher eukaryotes are able to extract severely misfolded glycoproteins from glycoprotein ERAD and target them to the non-glycoprotein ERAD pathway to maintain the homeostasis of the ER. PMID:26572623

Growth and development of cultured carrot cells and embryos under spaceflight conditions

NASA Technical Reports Server (NTRS)

Krikorian, A. D.; Dutcher, F. R.; Quinn, C. E.; Steward, F. C.

1981-01-01

Morphogenetically competent proembryonic cells and well-developed somatic embryos of carrot at two levels of organization were exposed for 18.5 days to a hypogravity environment aboard the Soviet Biosatellite Cosmos 1129. It was confirmed that cultured totipotent cells of carrot can give rise to embryos with well-developed roots and minimally developed shoots. It was also shown that the space hypogravity environment could support the further growth of already-organized, later somatic embryonic stages and give rise to fully developed embryo-plantlets with roots and shoots.

Carrot juice fermented with Lactobacillus plantarum NCU116 ameliorates type 2 diabetes in rats.

PubMed

Li, Chuan; Ding, Qiao; Nie, Shao-Ping; Zhang, Yan-Song; Xiong, Tao; Xie, Ming-Yong

2014-12-10

The effect of carrot juice fermented with Lactobacillus plantarum NCU116 on high-fat and low-dose streptozotocin (STZ)-induced type 2 diabetes in rats was studied. Rats were randomly divided into five groups: non-diabetes mellitus (NDM), untreated diabetes mellitus (DM), DM plus L. plantarum NCU116 (NCU), DM plus fermented carrot juice with L. plantarum NCU116 (FCJ), and DM plus non-fermented carrot juice (NFCJ). Treatments of NCU and FCJ for 5 weeks were found to favorably regulate blood glucose, hormones, and lipid metabolism in the diabetic rats, accompanied by an increase in short-chain fatty acid (SCFA) in the colon. In addition, NCU and FCJ had restored the antioxidant capacity and morphology of the pancreas and kidney and upregulated mRNA of low-density lipoprotein (LDL) receptor, cholesterol 7α-hydroxylase (CYP7A1), glucose transporter-4 (GLUT-4), peroxisome proliferator-activated receptor-α (PPAR-α), and peroxisome proliferator-activated receptor-γ (PPAR-γ). These results have for the first time demonstrated that L. plantarum NCU116 and the fermented carrot juice had the potential ability to ameliorate type 2 diabetes in rats.

ß-Carotene from Red Carrot Maintains Vitamin A Status, but Lycopene Bioavailability Is Lower Relative to Tomato Paste in Mongolian Gerbils

USDA-ARS?s Scientific Manuscript database

Red carrots contain lycopene in addition to ß-Carotene. The utility of red carrot as a functional food depends in part on the bioavailability of its constituent carotenoids. Lycopene bioavailability was compared in Mongolian gerbils (Meriones unguiculatus) fed freeze-dried red carrot and tomato pa...

Improving Classroom Behavior: The Carrot and the Stick.

ERIC Educational Resources Information Center

Talent, Barbara K.; Busch, Suzanne G.

A set of practical behavior change techniques for improving young children's classroom behavior are briefly discussed. Techniques are classified and discussed under two general categories: those that reduce frequency of behaviors ("sticks") and those that increase their frequency ("carrots"). Included under "sticks" are techniques such as ignoring…

Morphological Characteristics, Anatomical Structure, and Gene Expression: Novel Insights into Cytokinin Accumulation during Carrot Growth and Development.

PubMed

Wang, Guang-Long; Sun, Sheng; Xing, Guo-Ming; Wu, Xue-Jun; Wang, Feng; Xiong, Ai-Sheng

2015-01-01

Cytokinins have been implicated in normal plant growth and development. These bioactive molecules are essential for cell production and expansion in higher plants. Carrot is an Apiaceae vegetable with great value and undergoes significant size changes over the process of plant growth. However, cytokinin accumulation and its potential roles in carrot growth have not been elucidated. To address this problem, carrot plants at five stages were collected, and morphological and anatomical characteristics and expression profiles of cytokinin-related genes were determined. During carrot growth and development, cytokinin levels were the highest at the second stage in the roots, whereas relatively stable levels were observed in the petioles and leaves. DcCYP735A2 showed high expression at stage 2 in the roots, which may contribute largely to the higher cytokinin level at this stage. However, expression of most metabolic genes did not follow a pattern similar to that of cytokinin accumulation, indicating that cytokinin biosynthesis was regulated through a complex network. Genes involved in cytokinin signal perception and transduction were also integrated to normal plant growth and development. The results from the present work suggested that cytokinins may regulate plant growth in a stage-dependent manner. Our work would shed novel insights into cytokinin accumulation and its potential roles during carrot growth. Further studies regarding carrot cytokinins may be achieved by modification of the genes involved in cytokinin biosynthesis, inactivation, and perception.

Morphological Characteristics, Anatomical Structure, and Gene Expression: Novel Insights into Cytokinin Accumulation during Carrot Growth and Development

PubMed Central

Wang, Guang-Long; Sun, Sheng; Xing, Guo-Ming; Wu, Xue-Jun; Wang, Feng; Xiong, Ai-Sheng

2015-01-01

Cytokinins have been implicated in normal plant growth and development. These bioactive molecules are essential for cell production and expansion in higher plants. Carrot is an Apiaceae vegetable with great value and undergoes significant size changes over the process of plant growth. However, cytokinin accumulation and its potential roles in carrot growth have not been elucidated. To address this problem, carrot plants at five stages were collected, and morphological and anatomical characteristics and expression profiles of cytokinin-related genes were determined. During carrot growth and development, cytokinin levels were the highest at the second stage in the roots, whereas relatively stable levels were observed in the petioles and leaves. DcCYP735A2 showed high expression at stage 2 in the roots, which may contribute largely to the higher cytokinin level at this stage. However, expression of most metabolic genes did not follow a pattern similar to that of cytokinin accumulation, indicating that cytokinin biosynthesis was regulated through a complex network. Genes involved in cytokinin signal perception and transduction were also integrated to normal plant growth and development. The results from the present work suggested that cytokinins may regulate plant growth in a stage-dependent manner. Our work would shed novel insights into cytokinin accumulation and its potential roles during carrot growth. Further studies regarding carrot cytokinins may be achieved by modification of the genes involved in cytokinin biosynthesis, inactivation, and perception. PMID:26218147

Dissecting the Genetic Architecture of Shoot Growth in Carrot (Daucus carota L.) Using a Diallel Mating Design

PubMed Central

Turner, Sarah D.; Maurizio, Paul L.; Valdar, William; Yandell, Brian S.; Simon, Philipp W.

2017-01-01

Crop establishment in carrot (Daucus carota L.) is limited by slow seedling growth and delayed canopy closure, resulting in high management costs for weed control. Varieties with improved growth habit (i.e., larger canopy and increased shoot biomass) may help mitigate weed control, but the underlying genetics of these traits in carrot is unknown. This project used a diallel mating design coupled with recent Bayesian analytical methods to determine the genetic basis of carrot shoot growth. Six diverse carrot inbred lines with variable shoot size were crossed in WI in 2014. F1 hybrids, reciprocal crosses, and parental selfs were grown in a randomized complete block design with two blocks in WI (2015) and CA (2015, 2016). Measurements included canopy height, canopy width, shoot biomass, and root biomass. General and specific combining abilities were estimated using Griffing’s Model I, which is a common analysis for plant breeding experiments. In parallel, additive, inbred, cross-specific, and maternal effects were estimated from a Bayesian mixed model, which is robust to dealing with data imbalance and outliers. Both additive and nonadditive effects significantly influenced shoot traits, with nonadditive effects playing a larger role early in the growing season, when weed control is most critical. Results suggest the presence of heritable variation and thus potential for improvement of these phenotypes in carrot. In addition, results present evidence of heterosis for root biomass, which is a major component of carrot yield. PMID:29187419

Carrot cells: a pioneering platform for biopharmaceuticals production.

PubMed

Rosales-Mendoza, Sergio; Tello-Olea, Marlene Anahí

2015-03-01

Carrot (Daucus carota L.) is of importance in the molecular farming field as it constitutes the first plant species approved to produce biopharmaceuticals for human use. In this review, features that make carrot an advantageous species in the molecular farming field are analyzed and a description of the developments achieved with this crop thus far is presented. A guide for genetic transformation procedures is also included. The state of the art comprises ten vaccine prototypes against Measles virus, Hepatitis B virus, Human immunodeficiency virus, Yersinia pestis, Chlamydia trachomatis, Mycobacterium tuberculosis, enterotoxigenic Escherichia coli, Corynebacterium diphtheria/Clostridium tetani/Bordetella pertussis, and Helicobacter pylori; as well as the case of the glucocerebrosidase, an enzyme used for replacement therapy, and other therapeutics. Perspectives for these developments are envisioned and innovations are proposed such as the use of transplastomic technologies-, hairy roots-, and viral expression-based systems to improve yields and develop new products derived from this advantageous plant species.

Conversion of a diversity arrays technology marker differentiating wild and cultivated carrots to a co-dominant cleaved amplified polymorphic site marker

USDA-ARS?s Scientific Manuscript database

Cultivated carrot and its wild ancestor co-occur in most temperate regions of the world and can easily hybridize. The genetic basis of the process of domestication in carrot is not well recognized. Recent results of an investigation on genetic diversity structure of cultivated and wild carrot and si...

Potential of Cinnamon Oil Emulsions as Alternative Washing Solutions of Carrots.

PubMed

Zhang, Yue; Chen, Huaiqiong; Critzer, Faith; Davidson, P Michael; Zhong, Qixin

2017-06-01

The objective of this study was to evaluate the potential of cinnamon oil emulsions as alternative washing solutions to improve the microbial safety of carrots. Whey protein concentrate (WPC), gum arabic (GA), lecithin, and their combinations were used to prepare cinnamon oil emulsions. The emulsions were characterized for their hydrodynamic diameter (D h ) during 7 days of storage and their antimicrobial activity against cocktails of Salmonella enterica , Escherichia coli O157:H7, and Listeria monocytogenes . The D h of the emulsion prepared with the GA+WPC blend did not change significantly (195.0 to 184.1 nm), whereas all other emulsions showed varying degrees of increases in D h . Compared with free cinnamon oil dissolved in 5% ethanol, all emulsions showed similar or lower MICs and MBCs. Emulsions prepared with GA and equal masses of GA and WPC were chosen and diluted to 0.2 and 0.5% cinnamon oil to wash carrots that were surface inoculated with bacterial cocktails because of their lower MICs and MBCs than free oil. Emulsions resulted in significantly higher reductions of pathogens on carrots than free cinnamon oil, 3.0 to 3.7 versus 2.1 to 2.3 log CFU/g at 0.5% cinnamon oil and 2.0 to 3.0 versus 1.0 to 1.7 log CFU/g at 0.2% cinnamon oil. No transfer of bacteria from inoculated carrots to wash solutions and no effects of organic load on log reductions were only observed for wash treatments with 0.5% emulsified cinnamon oil. Thus, the cinnamon oil emulsions are potential alternative postharvest washing solutions for fresh produce production.

Genome-wide identification, expansion, and evolution analysis of homeobox genes and their expression profiles during root development in carrot.

PubMed

Que, Feng; Wang, Guang-Long; Li, Tong; Wang, Ya-Hui; Xu, Zhi-Sheng; Xiong, Ai-Sheng

2018-06-16

The homeobox gene family, a large family represented by transcription factors, has been implicated in secondary growth, early embryo patterning, and hormone response pathways in plants. However, reports about the information and evolutionary history of the homeobox gene family in carrot are limited. In the present study, a total of 130 homeobox family genes were identified in the carrot genome. Specific codomain and phylogenetic analyses revealed that the genes were classified into 14 subgroups. Whole genome and proximal duplication participated in the homeobox gene family expansion in carrot. Purifying selection also contributed to the evolution of carrot homeobox genes. In Gene Ontology (GO) analysis, most members of the HD-ZIP III and IV subfamilies were found to have a lipid binding (GO:0008289) term. Most HD-ZIP III and IV genes also harbored a steroidogenic acute regulatory protein-related lipid transfer (START) domain. These results suggested that the HD-ZIP III and IV subfamilies might be related to lipid transfer. Transcriptome and quantitative real-time PCR (RT-qPCR) data indicated that members of the WOX and KNOX subfamilies were likely implicated in carrot root development. Our study provided a useful basis for further studies on the complexity and function of the homeobox gene family in carrot.

Deposition of carotenoids in egg yolk by short-term supplement of coloured carrot (Daucus carota) varieties as forage material for egg-laying hens.

PubMed

Hammershøj, Marianne; Kidmose, Ulla; Steenfeldt, Sanna

2010-05-01

Supplying egg-laying hens with different forage materials may influence egg production and quality. The aim of this study was to examine the short-term effects of standard feed plus 70 g day(-1) per hen of three coloured carrot varieties (orange, yellow and purple) as forage material in comparison with a standard feed control on egg production, egg yolk colour and deposition of carotenoids in the yolk. Carrot supplementation reduced feed intakes significantly, but not on a dry matter basis. Orange carrot treatment significantly reduced egg mass production, whereas yellow and purple carrot treatments did not differ from the control. Egg and yolk weights of all carrot-supplemented treatments were significantly lower than those of the control, but yolk percentages were similar. Yolk redness increased significantly in the order control < yellow < orange < purple. A similar trend was seen for yolk yellowness, but yellow and orange carrots reached the same level. Yolk colour and carotenoid contents correlated positively and significantly. In particular, purple carrot treatment increased the yolk content of lutein (>1.5-fold) and beta-carotene (>100-fold) compared with the control. Supplementing the feed of egg-laying hens with coloured carrots efficiently increased yolk colour parameters and carotenoid contents, which gives opportunities for improved nutritional value of eggs from forage material-supplemented hens.

Molecular detection of aster yellows phytoplasma and 'Candidatus Liberibacter solanacearum' in carrots affected by the psyllid Trioza apicalis (Hemiptera: triozidae) in Finland

USDA-ARS?s Scientific Manuscript database

Carrot psyllid (Trioza apicalis Förster) causes considerable damage to carrot (Daucus carota L.) in many parts of Europe. It was recently established that the new bacterium “Candidatus Liberibacter solanacearum” is associated with carrot psyllid and plants affected by this insect pest. No other path...

Characterization of inositol phosphates in carrot (Daucus carota L. ) cells

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

Rincon, M.; Chen, Q.; Boss, W.F.

1989-01-01

We have shown previously that inositol-1,4,5-trisphosphate (IP{sub 3}) stimulates an efflux of {sup 45}Ca{sup 2+} from fusogenic carrot protoplasts. In light of these results, we suggested that IP{sub 3} might serve as a second messenger for the mobilization of intracellular Ca{sup 2+} in higher plant cells. To determine whether or not IP{sub 3} and other inositol phosphates were present in the carrot cells, the cells were labeled with myo-(2-{sup 3}H)inositol for 18 hours and extracted with ice-cold 10% trichloroacetic acid. The inositol metabolites were separated by anion exchange chromatography and by paper electrophoresis. We found that ({sup 3}H)inositol metabolites coelutedmore » with inositol bisphosphate (IP{sub 2}) and IP{sub 3} when separated by anion exchange chromatography. However, we could not detect IP{sub 2} or IP{sub 3} when the inositol metabolites were analyzed by paper electrophoresis even though the polyphosphoinositides, which are the source of IP{sub 2} and IP{sub 3}, were present in these cells. Thus, ({sup 3}H)inositol metabolites other than IP{sub 2} and IP{sub 3} had coeluted on the anion exchange columns. The data indicate that either IP{sub 3} is rapidly metabolized or that it is not present at a detectable level in the carrot cells.« less

Phytotoxicity and accumulation of chromium in carrot plants and the derivation of soil thresholds for Chinese soils.

PubMed

Ding, Changfeng; Li, Xiaogang; Zhang, Taolin; Ma, Yibing; Wang, Xingxiang

2014-10-01

Soil environmental quality standards in respect of heavy metals for farmlands should be established considering both their effects on crop yield and their accumulation in the edible part. A greenhouse experiment was conducted to investigate the effects of chromium (Cr) on biomass production and Cr accumulation in carrot plants grown in a wide range of soils. The results revealed that carrot yield significantly decreased in 18 of the total 20 soils with Cr addition being the soil environmental quality standard of China. The Cr content of carrot grown in the five soils with pH>8.0 exceeded the maximum allowable level (0.5mgkg(-1)) according to the Chinese General Standard for Contaminants in Foods. The relationship between carrot Cr concentration and soil pH could be well fitted (R(2)=0.70, P<0.0001) by a linear-linear segmented regression model. The addition of Cr to soil influenced carrot yield firstly rather than the food quality. The major soil factors controlling Cr phytotoxicity and the prediction models were further identified and developed using path analysis and stepwise multiple linear regression analysis. Soil Cr thresholds for phytotoxicity meanwhile ensuring food safety were then derived on the condition of 10 percent yield reduction. Copyright © 2014 Elsevier Inc. All rights reserved.

Profiling of the Terpene Metabolome in Carrot Fruits of Wild ( Daucus carota L. ssp. carota) Accessions and Characterization of a Geraniol Synthase.

PubMed

Yahyaa, Mosaab; Ibdah, Muhammad; Marzouk, Sally; Ibdah, Mwafaq

2018-03-14

Fruits from wild carrot ( Daucus carota L. ssp. carota) have been used for medicinal purposes since ancient times. The oil of its seeds, with their abundant monoterpenes and sesquiterpenes, has drawn attention in recent years because of its potential pharmaceutical application. A combined chemical, biochemical, and molecular study was conducted to evaluate the differential accumulation of terpene volatiles in carrot fruits of wild accessions. This work reports a similarity-based cloning strategy identification and functional characterization of one carrot monoterpene terpene synthase, WtDcTPS1. Recombinant WtDcTPS1 protein produces mainly geraniol, the predominant monoterpene in carrot seeds of wild accession 23727. The results suggest a role for the WtDcTPS1 gene in the biosynthesis of carrot fruit aroma and flavor compounds.

The effect of aldicarb on nematode population and its persistence in carrots, soil and hydroponic solution.

PubMed

Lue, L P; Lewis, C C; Melchor, V E

1984-04-01

Aldicarb, Temik 15 G, was incorporated in furrows at 3.37 and 6.73 kg ai (active ingredient)/ha and carrots (Daucus carota L.) were directly seeded on the same day. The numbers of nematode larvae were significantly suppressed in the treated plots; averages were 249, 74, and 51/50 cc soil samples for control (0), 3.37 and 6.73 kg ai/ha, respectively. Aldicarb treatment resulted in a 28% yield increase as compared to the untreated. Aldicarb residue in carrots was 28 ppb for the low treatment and 46 ppb for the high. Residual levels in soil of high treatment declined from 61 to 31 ppb during two weeks prior to harvest, meanwhile, those in the low decreased slightly from 13 to 12 ppb. Carrots placed in hydroponic solution containing aldicarb 14.5 ppm for 6 days, had an aldicarb residue of 10.26 ppb and the hydroponic solution, 2.7 ppb. Persistence of aldicarb residue was in carrot greater than in soil greater than in hydroponic solution.

Insights into the Evolution of Hydroxyproline-Rich Glycoproteins from 1000 Plant Transcriptomes1[OPEN

PubMed Central

Cassin, Andrew M.; Soltis, Douglas E.; Miles, Nicholas W.; Melkonian, Michael; Melkonian, Barbara; Wu, Shuangxiu; Edger, Patrick P.; Carpenter, Eric J.

2017-01-01

The carbohydrate-rich cell walls of land plants and algae have been the focus of much interest given the value of cell wall-based products to our current and future economies. Hydroxyproline-rich glycoproteins (HRGPs), a major group of wall glycoproteins, play important roles in plant growth and development, yet little is known about how they have evolved in parallel with the polysaccharide components of walls. We investigate the origins and evolution of the HRGP superfamily, which is commonly divided into three major multigene families: the arabinogalactan proteins (AGPs), extensins (EXTs), and proline-rich proteins. Using motif and amino acid bias, a newly developed bioinformatics pipeline, we identified HRGPs in sequences from the 1000 Plants transcriptome project (www.onekp.com). Our analyses provide new insights into the evolution of HRGPs across major evolutionary milestones, including the transition to land and the early radiation of angiosperms. Significantly, data mining reveals the origin of glycosylphosphatidylinositol (GPI)-anchored AGPs in green algae and a 3- to 4-fold increase in GPI-AGPs in liverworts and mosses. The first detection of cross-linking (CL)-EXTs is observed in bryophytes, which suggests that CL-EXTs arose though the juxtaposition of preexisting SPn EXT glycomotifs with refined Y-based motifs. We also detected the loss of CL-EXT in a few lineages, including the grass family (Poaceae), that have a cell wall composition distinct from other monocots and eudicots. A key challenge in HRGP research is tracking individual HRGPs throughout evolution. Using the 1000 Plants output, we were able to find putative orthologs of Arabidopsis pollen-specific GPI-AGPs in basal eudicots. PMID:28446636

Diversity, genetic mapping, and signatures of domestication in the carrot (Daucus carota L.) genome, as revealed by Diversity Arrays Technology (DArT) markers

USDA-ARS?s Scientific Manuscript database

Carrot is one of the most economically important vegetables worldwide, however, genetic and genomic resources supporting carrot breeding remain limited. We developed a Diversity Arrays Technology (DArT) platform for wild and cultivated carrot and used it to investigate genetic diversity and to devel...

The Impact of Carrot Enriched in Iodine through Soil Fertilization on Iodine Concentration and Selected Biochemical Parameters in Wistar Rats.

PubMed

Piątkowska, Ewa; Kopeć, Aneta; Bieżanowska-Kopeć, Renata; Pysz, Mirosław; Kapusta-Duch, Joanna; Koronowicz, Aneta Agnieszka; Smoleń, Sylwester; Skoczylas, Łukasz; Ledwożyw-Smoleń, Iwona; Rakoczy, Roksana; Maślak, Edyta

2016-01-01

Iodine is one of the trace elements which are essential for mammalian life. The major objective of iodine biofortification of plants is to obtain food rich in this trace element, which may increase its consumption by various populations. Additionally, it may reduce the risk of iodine deficiency diseases. In this research for the first time we have assessed the bioavailability of iodine from raw or cooked carrot biofortified with this trace element on iodine concentration in selected tissues and various biochemical parameters as well as mRNA expression of some genes involved in iodine metabolism in Wistar rats. Statistically, a significantly higher iodine level was determined in urine, faeces and selected tissues of rats fed a diet containing biofortified raw carrot as compared to a diet without iodine and a diet containing control cooked carrot. Biofortified raw carrot significantly increased triiodothyronine concentration as compared to animals from other experimental groups. The highest thyroid stimulating hormone level was determined in rats fed control cooked carrots. mRNA expression of selected genes was affected by different dietary treatment in rats' hearts. Biofortified raw and cooked carrot could be taken into account as a potential source of iodine in daily diets to prevent iodine deficiency in various populations.

The Impact of Carrot Enriched in Iodine through Soil Fertilization on Iodine Concentration and Selected Biochemical Parameters in Wistar Rats

PubMed Central

Piątkowska, Ewa; Kopeć, Aneta; Bieżanowska-Kopeć, Renata; Pysz, Mirosław; Kapusta-Duch, Joanna; Koronowicz, Aneta Agnieszka; Smoleń, Sylwester; Skoczylas, Łukasz; Ledwożyw-Smoleń, Iwona; Rakoczy, Roksana; Maślak, Edyta

2016-01-01

Iodine is one of the trace elements which are essential for mammalian life. The major objective of iodine biofortification of plants is to obtain food rich in this trace element, which may increase its consumption by various populations. Additionally, it may reduce the risk of iodine deficiency diseases. In this research for the first time we have assessed the bioavailability of iodine from raw or cooked carrot biofortified with this trace element on iodine concentration in selected tissues and various biochemical parameters as well as mRNA expression of some genes involved in iodine metabolism in Wistar rats. Statistically, a significantly higher iodine level was determined in urine, faeces and selected tissues of rats fed a diet containing biofortified raw carrot as compared to a diet without iodine and a diet containing control cooked carrot. Biofortified raw carrot significantly increased triiodothyronine concentration as compared to animals from other experimental groups. The highest thyroid stimulating hormone level was determined in rats fed control cooked carrots. mRNA expression of selected genes was affected by different dietary treatment in rats’ hearts. Biofortified raw and cooked carrot could be taken into account as a potential source of iodine in daily diets to prevent iodine deficiency in various populations. PMID:27043135

Cooked carrots volatiles. AEDA and odor activity comparisons. Identification of Linden Ether as an important aroma component

USDA-ARS?s Scientific Manuscript database

MS with GC-RI evidence was found for the presence of Linden ether in cooked carrot. Evaluation of the GC effluent from cooked carrot volatiles using Aroma Extract Dilution Analysis (AEDA) found Linden ether with the highest Flavor Dilution (FD) factor. Others with 10 fold lower FD factors were B-i...

Thioredoxin and NADP-thioredoxin reductase from cultured carrot cells

NASA Technical Reports Server (NTRS)

Johnson, T. C.; Cao, R. Q.; Kung, J. E.; Buchanan, B. B.

1987-01-01

Dark-grown carrot (Daucus carota L.) tissue cultures were found to contain both protein components of the NADP/thioredoxin system--NADP-thioredoxin reductase and the thioredoxin characteristic of heterotrophic systems, thioredoxin h. Thioredoxin h was purified to apparent homogeneity and, like typical bacterial counterparts, was a 12-kdalton (kDa) acidic protein capable of activating chloroplast NADP-malate dehydrogenase (EC 1.1.1.82) more effectively than fructose-1,6-bisphosphatase (EC 3.1.3.11). NADP-thioredoxin reductase (EC 1.6.4.5) was partially purified and found to be an arsenite-sensitive enzyme composed of two 34-kDa subunits. Carrot NADP-thioredoxin reductase resembled more closely its counterpart from bacteria rather than animal cells in acceptor (thioredoxin) specificity. Upon greening of the cells, the content of NADP-thioredoxin-reductase activity, and, to a lesser extent, thioredoxin h decreased. The results confirm the presence of a heterotrophic-type thioredoxin system in plant cells and raise the question of its physiological function.

Uptake of polybrominated diphenyl ethers by carrot and lettuce crops grown in compost-amended soils.

PubMed

Bizkarguenaga, E; Iparraguirre, A; Oliva, E; Quintana, J B; Rodil, R; Fernández, L A; Zuloaga, O; Prieto, A

2016-02-01

The uptake of polybrominated diphenyl ethers (PBDEs) by carrot and lettuce was investigated. Degradation of PBDEs in soil in the absence of the plants was discarded. Different carrot (Nantesa and Chantenay) and lettuce (Batavia Golden Spring and Summer Queen) varieties were grown in fortified or contaminated compost-amended soil mixtures under greenhouse conditions. After plant harvesting, roots (core and peel) and leaves were analyzed separately for carrot, while for lettuce, leaves and hearts were analyzed together. The corresponding bioconcentration factors (BCFs) were calculated. In carrots, a concentration gradient of 2,2',3,4,4',5'-hexabromodiphenyl ether (BDE-138) became evident that decreased from the root peel via root core to the leaves. For decabromodiphenyl ether (BDE-209) at the low concentration level (7 and 20 ng g(-1)), the leaves incorporated the highest concentration of the target substance. For lettuce, a decrease in the BCF value (from 0.24 to 0.02) was observed the higher the octanol-water partition coefficient, except in the case of BDE-183 (BCF = 0.51) and BDE-209 (BCF values from 0.41 to 0.74). Significant influence of the soils and crop varieties on the uptake could not be supported. Metabolic debromination, hydroxylation or methylation of the target PBDEs in the soil-plant system was not observed.

Polyphosphoinositides are present in plasma membranes isolated from fusogenic carrot cells

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

Wheeler, J.J.; Boss, W.F.

1987-10-01

Fusogenic carrot cells grown in suspension culture were labeled 12 hours with myo-(2-/sup 3/H)inositol. Plasma membranes were isolated from the prelabeled fusogenic carrot cells by both aqueous polymer two-phase partitioning and Renografin density gradients. With both methods, the plasma membrane-enriched fractions, as identified by marker enzymes, were enriched in (/sup 3/H)inositol-labeled phosphatidylinositol monophosphate (PIP) and phosphatidylinositol bisphosphate (PIP/sub 2/). An additional (/sup 3/H)inositol-labeled lipid, lysophosphatidylinositol monophosphate, which migrated between PIP and PIP/sub 2/ on thin layer plates, was found primarily in the plasma membrane-rich fraction of the fusogenic cells. This was in contrast to lysophosphatidylinositol which is found primarily inmore » the lower phase, microsomal/mitchrondrial-rich fraction.« less

Ingestion of transgenic carrots expressing the Escherichia coli heat-labile enterotoxin B subunit protects mice against cholera toxin challenge.

PubMed

Rosales-Mendoza, Sergio; Soria-Guerra, Ruth Elena; López-Revilla, Rubén; Moreno-Fierros, Leticia; Alpuche-Solís, Angel Gabriel

2008-01-01

Diarrheal diseases caused by Vibrio cholerae and enterotoxigenic Escherichia coli (ETEC) are worldwide health problems that might be prevented with vaccines based on edible plants expressing the B subunit from either the cholera toxin (CTB) or the E. coli heat labile toxin (LTB). In this work we analyzed the immunity induced in Balb/c mice by ingestion of three weekly doses of 10 mug of LTB derived from transgenic carrot material. Although the anti-LTB serum immunoglobulin G (IgG) and intestinal IgA antibody responses were higher with 10 mug-doses of pure bacterial recombinant LTB (rLTB), the transgenic carrot material also elicited significant serum and intestinal antibody responses. Serum anti-LTB IgG1 antibodies predominated over IgG2a antibodies, suggesting that mainly Th2 responses were induced. A decrease of intestinal fluid accumulation after cholera toxin challenge was observed in mice immunized with either rLTB or LTB-containing carrot material. These results demonstrate that ingestion of carrot-derived LTB induces antitoxin systemic and intestinal immunity in mice and suggest that transgenic carrots expressing LTB may be used as an effective edible vaccine against cholera and ETEC diarrhea in humans.

The effect of boron on plasma membrane electron transport and associated proton secretion by cultured carrot cells.

PubMed

Barr, R; Böttger, M; Crane, F L

1993-09-01

Plasma membrane electron transport reactions and associated proton secretion were studied in boron-deficient carrot cells. It was found that the hormone-sensitive plasma membrane NADH oxidase was inhibited by boron deficiency and that under such conditions activity could be restored by exogenous boric acid with or without 2,4-dichlorophenoxy acetic acid. Gramicidin, a channel-forming protonophore, further stimulated NADH oxidase by carrot cells. Proton secretion, associated with plasma membrane H(+)-ATPase, was also affected by boron deficiency, but not as severely as ferricyanide-generated proton secretion, reflecting plasma membrane electron transport. The addition of 1 mM boric acid and 1 microM 2,4-dichlorophenoxy acetic acid to carrot cells fully restored the H+ secretion in presence of ferricyanide. The effect of boron deficiency in cultured carrot cells can, therefore, be directly associated with cell growth through its effect on the plasma membrane NADH oxidase and H+ secretion. Ferricyanide provides a probe which activates transmembrane electron transport that is only coupled to proton release when boron is present.

Changes in Cell Wall Properties Coincide with Overexpression of Extensin Fusion Proteins in Suspension Cultured Tobacco Cells

DOE PAGES

Tan, Li; Pu, Yunqiao; Pattathil, Sivakumar; ...

2014-12-23

Extensins are one subfamily of the cell wall hydroxyproline-rich glycoproteins, containing characteristic SerHyp4 glycosylation motifs and intermolecular cross-linking motifs such as the TyrXaaTyr sequence. Extensins are believed to form a cross-linked network in the plant cell wall through the tyrosine-derivatives isodityrosine, pulcherosine, and di-isodityrosine. Overexpression of three synthetic genes encoding different elastin-arabinogalactan protein-extensin hybrids in tobacco suspension cultured cells yielded novel cross-linking glycoproteins that shared features of the extensins, arabinogalactan proteins and elastin. The cell wall properties of the three transgenic cell lines were all changed, but in different ways. One transgenic cell line showed decreased cellulose crystallinity and increasedmore » wall xyloglucan content; the second transgenic cell line contained dramatically increased hydration capacity and notably increased cell wall biomass, increased di-isodityrosine, and increased protein content; the third transgenic cell line displayed wall phenotypes similar to wild type cells, except changed xyloglucan epitope extractability. In conclusion, these data indicate that overexpression of modified extensins may be a route to engineer plants for bioenergy and biomaterial production.« less

Changes in cell wall properties coincide with overexpression of extensin fusion proteins in suspension cultured tobacco cells.

PubMed

Tan, Li; Pu, Yunqiao; Pattathil, Sivakumar; Avci, Utku; Qian, Jin; Arter, Allison; Chen, Liwei; Hahn, Michael G; Ragauskas, Arthur J; Kieliszewski, Marcia J

2014-01-01

Extensins are one subfamily of the cell wall hydroxyproline-rich glycoproteins, containing characteristic SerHyp4 glycosylation motifs and intermolecular cross-linking motifs such as the TyrXaaTyr sequence. Extensins are believed to form a cross-linked network in the plant cell wall through the tyrosine-derivatives isodityrosine, pulcherosine, and di-isodityrosine. Overexpression of three synthetic genes encoding different elastin-arabinogalactan protein-extensin hybrids in tobacco suspension cultured cells yielded novel cross-linking glycoproteins that shared features of the extensins, arabinogalactan proteins and elastin. The cell wall properties of the three transgenic cell lines were all changed, but in different ways. One transgenic cell line showed decreased cellulose crystallinity and increased wall xyloglucan content; the second transgenic cell line contained dramatically increased hydration capacity and notably increased cell wall biomass, increased di-isodityrosine, and increased protein content; the third transgenic cell line displayed wall phenotypes similar to wild type cells, except changed xyloglucan epitope extractability. These data indicate that overexpression of modified extensins may be a route to engineer plants for bioenergy and biomaterial production.

[Hydroxyproline: Rich glycoproteins of the plant and cell wall

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

Varner, J.E.

1993-01-01

Since xylem tissue includes the main cell types which are lignified, we are interested in gene expression of glycine-rich proteins and proline-rich proteins, and other proteins which are involved in secondary cell wall thickening during xylogenesis. Since the main feature of xylogenesis is the deposition of additional wall components, study of the mechanism of xylogenesis will greatly advance our knowledge of the synthesis and assembly of wall macromolecules. We are using the in vitro xylogenesis system from isolated Zinnia mesophyll cells to isolate genes which are specifically expressed during xylogenesis. We have used subtractive hybridization methods to isolate a numbermore » of cDNA clones for differentially regulated genes from the cells after hormonal induction. So far, we have partially characterized 18 different cDNA clones from 239 positive clones. These differentially regulated genes can be divided into three sets according to the characteristics of gene expression in the induction medium and the control medium. The first set is induced in both the induction medium and the control medium without hormones. The second set is induced mainly in the induction medium and in the control medium with the addition of NAA alone. Two of thesegenes are exclusively induced by auxin. The third set of genes is induced mainly in the induction medium. Since these genes are not induced by either auxin or cytokinin alone, they may be directly involved in the process of xylogenesis. Our experiments on the localization of H[sub 2]O[sub 2] production reinforce the earlier ideas of others that H[sub 2]O[sub 2] is involved in normal lignification.« less

Expression of Pneumocystis jirovecii Major Surface Glycoprotein in Saccharomyces cerevisiae

PubMed Central

Kutty, Geetha; England, Katherine J.; Kovacs, Joseph A.

2013-01-01

The major surface glycoprotein (Msg), which is the most abundant protein expressed on the cell surface of Pneumocystis organisms, plays an important role in the attachment of this organism to epithelial cells and macrophages. In the present study, we expressed Pneumocystis jirovecii Msg in Saccharomyces cerevisiae, a phylogenetically related organism. Full-length P. jirovecii Msg was expressed with a DNA construct that used codons optimized for expression in yeast. Unlike in Pneumocystis organisms, recombinant Msg localized to the plasma membrane of yeast rather than to the cell wall. Msg expression was targeted to the yeast cell wall by replacing its signal peptide, serine-threonine–rich region, and glycophosphatidylinositol anchor signal region with the signal peptide of cell wall protein α-agglutinin of S. cerevisiae, the serine-threonine–rich region of epithelial adhesin (Epa1) of Candida glabrata, and the carboxyl region of the cell wall protein (Cwp2) of S. cerevisiae, respectively. Immunofluorescence analysis and treatment with β-1,3 glucanase demonstrated that the expressed Msg fusion protein localized to the yeast cell wall. Surface expression of Msg protein resulted in increased adherence of yeast to A549 alveolar epithelial cells. Heterologous expression of Msg in yeast will facilitate studies of the biologic properties of Pneumocystis Msg. PMID:23532098

Experimental study and modelling of water sorption/desorption isotherms on two agricultural products: Apple and carrot

NASA Astrophysics Data System (ADS)

Timoumi, S.; Zagrouba, F.; Mihoubi, D.; Tlili, M. M.

2004-12-01

This work is focused on some properties of dried apple (Red Chief) and carrot (Misky). Water sorption isotherms of carrot and apple were investigated at three temperatures: 30, 40 and 60°C, corresponding to drying temperatures, by the static method consisting of the use of different sulphuric acid solutions. Guggenheim-Anderson-de Boer (G.A.B) model is found to describe the experimental curves better than Henderson, Hasley and Oswin models with a correlation coefficient superior to 0.97 for both products. The hysteresis phenomenon was clearly observed in the case of apple isotherms. The experimental data were also used to determine the isosteric enthalpy of desorption of apple and carrot. The isosteric enthalpy of desorption decreased with increase in moisture content and the trend became asymptotic.

Growth of Carrot and Tomato from Oxamyl-coated Seed and Control of Meloidogyne hapla.

PubMed

Townshend, J L

1990-04-01

Oxamyl was coated on carrot (Daucus carota L. cv. Spartan Fancy-80) and tomato (Lycopersicon esculentum Mill. cv. Glamour) seeds with a polymer sticker for the control of Meloidogyne hapla. The sticker diluted in water 1:1 delayed carrot seedling emergence. Oxamyl at 40 mg/ml in a 1:5 dilution of sticker lowered the rate of carrot seedling emergence until day 13 and plant growth until day 28. Oxamyl at 20 or 40 mg/ml in a 1:5 dilution of sticker on carrot seeds planted in M. hapla-infested muck soil resulted in fewer galled tap roots and fewer galls per root system 4 weeks after planting. Tap root lengths were greater than those of the control. Tomato seedling emergence was delayed and top and root weights were reduced, relative to the control, at 25 days by the sticker diluted 1:1 to 1:3. Oxamyl at 20 or 40 mg/ml in a 1:5 diluted sticker delayed tomato seedling emergence. Top weights of tomato seedlings from seeds coated with 20 mg/ml of oxamyl in a 1:5 diluted sticker planted in a silt loam were greater than control top weights at 4 and 6 weeks. Root weights were greater than those of the control only at 4 weeks. There were fewer galls per gram of root on seedlings from oxamyl-coated seeds and fewer juveniles per pot of soil, relative to the controls, only at 4 weeks.

Polyamine Uptake in Carrot Cell Cultures 1

PubMed Central

Pistocchi, Rossella; Bagni, Nello; Creus, José A.

1987-01-01

Putrescine and spermidine uptake into carrot (Daucus carota L.) cells in culture was studied. The time course of uptake showed that the two polyamines were very quickly transported into the cells, reaching a maximum absorption within 1 minute. Increasing external polyamine concentrations up to 100 millimolar showed the existence of a biphasic system with different affinities at low and high polyamine concentrations. The cellular localization of absorbed polyamines was such that a greater amount of putrescine was present in the cytoplasmic soluble fraction, while spermidine was mostly present in cell walls. The absorbed polyamines were released into the medium in the presence of increasing external concentrations of the corresponding polyamine or Ca2+. The effects of Ca2+ were different for putrescine and spermidine; putrescine uptake was slightly stimulated by 10 micromolar Ca2+ and inhibited by higher concentrations, while for spermidine uptake there was an increasing stimulation in the Ca2+ concentration range between 10 micromolar and 1 millimolar. La3+ nullified the stimulatory effect of 10 micromolar Ca2+ on putrescine uptake and that of 1 millimolar Ca2+ on spermidine uptake. La3+ at 0.5 to 1 millimolar markedly inhibited the uptake of both polyamines, suggesting that it interferes with the sites of polyamine uptake. Putrescine uptake was affected to a lesser extent by metabolic inhibitors than was spermidine uptake. It is proposed that the entry of polyamines into the cells is driven by the transmembrane electrical gradient, with a possible antiport mechanism between external and internal polyamine molecule. PMID:16665446

The Effect of Temperature and Concentration of Foaming Agent to the β-Carotene Content in Product Derived from Carrots

NASA Astrophysics Data System (ADS)

Fardiyah, Qonitah; Rumhayati, Barlah; Husnul Khotimah, Yuniesti

2018-01-01

Carrot (Daucus carota L) is vegetable that contain body essetial vitamins, especially β-carotene. In this research, the essense of fresh carrots are taken and processed to carrot powder using foam mat drying method. This research aims to study the effect of temperature and concentration of foaming agent to the β-carotene content in product derived from carrots. The temperature variation that used in this research are 40°C, 50°C, 60°C and 70°C, while he variation of foaming agent (tween 80) are 0,01% (v/v); 0,1%(v/v); 0,2%(v/v) and 0,3%(v/v). The results shows that the maximum drying temperature is 50°C with β-carotene content 10.55 mg/kg and the maximum concentration of foaming agent (tween 80) is 0.2% (v/v) with β-carotene content 10.36 mg/kg.

The 1064 nm laser-induced breakdown spectroscopy (LIBS) inspection to detect the nutrient elements in freshly cut carrot samples

NASA Astrophysics Data System (ADS)

Yudasari, N.; Prasetyo, S.; Suliyanti, M. M.

2018-03-01

The laser-induced breakdown spectroscopy (LIBS) technique was applied to detect the nutrient elements contained in fresh carrot. Nd:YAG laser the wavelength of 1064 nm was employed in the experiments for ablation. Employing simple set-up of LIBS and preparing the sample with less step method, we are able to detect 18 chemical elements including some fundamental element of carrot, i.e Mg, Al, Fe, Mn, Ti, Ca, and Mn. By applying normalized profiles calculation on some of the element, we are able to compare the concentration level of each element of the outer and inner part of carrot.

Apparent Inhibition of β-Fructosidase Secretion by Tunicamycin May Be Explained by Breakdown of the Unglycosylated Protein during Secretion 1

PubMed Central

Faye, Loïc; Chrispeels, Maarten J.

1989-01-01

Suspension-cultured carrot (Daucus carota) cells synthesize and secrete β-fructosidase, a glycoprotein with asparagine-linked glycans. Treatment of the cells with tunicamycin completely inhibits the apparent secretion of β-fructosidase as measured by the accumulation of the radioactive protein in the cell wall or the culture medium. In the past, such a result has been interpreted as an inhibition of secretion by tunicamycin, but we suggest another explanation based on the following results. In the presence of tunicamycin, unglycosylated β-fructosidase is synthesized and is associated with an endoplasmic-reticulum-rich microsomal fraction. Pulse-chase experiments show that the unglycosylated β-fructosidase does not remain in the cells and appears to be secreted in the same way as glycosylated β-fructosidase; however, no radioactive, unglycosylated β-fructosidase accumulates extracellularly (cell wall or medium). Protoplasts obtained from carrot cells secrete β-fructosidase protein and activity, and treatment of the protoplasts with tunicamycin results in the synthesis of unglycosylated β-fructosidase. In the presence of tunicamycin, there is no accumulation of β-fructosidase activity or unglycosylated β-fructosidase polypeptide in the protoplast incubation medium. These results are consistent with the interpretation that the glycans of β-fructosidase are necessary for its stability, and that in these suspension-cultured cells, the unglycosylated enzyme is degraded during the last stage(s) of secretion, or immediately after its arrival in the wall. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 PMID:16666631

Apparent inhibition of. beta. -fructosidase secretion by tunicamycin may be explained by breakdown of the unglycosylated protein during secretion. [Daucus carota

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

Faye, L.; Chrispeels, M.J.

1989-03-01

Suspension-cultured carrot (Daucus carota) cells synthesize and secrete {beta}-fructosidase, a glycoprotein with asparagine-linked glycans. Treatment of the cells with tunicamycin completely inhibits the apparent secretion of {beta}-fructosidase as measured by the accumulation of the {sup 35}S-labelled protein in the cell wall or the culture medium. In the past, such a result has been interpreted as an inhibition of secretion by tunicamycin, but we suggest another explanation based on the following results. In the presence of tunicamycin, unglycosylated {beta}-fructosidase is synthesized and is associated with an endoplasmic-reticulum-rich microsomal fraction. Pulse-chase experiments show that the unglycosylated {beta}-fructosidase does not remain in themore » cells and appears to be secreted in the same way as glycosylated {beta}-fructosidase; however, no radioactive, unglycosylated {beta}-fructosidase accumulates extracellularly (cell wall or medium). Protoplasts obtained from carrot cells secrete {beta}-fructosidase protein and activity, and treatment of the protoplasts with tunicamycin results in the synthesis of unglycosylated {beta}-fructosidase. In the presence of tunicamycin, there is no accumulation of {beta}-fructosidase activity or unglycosylated {beta}-fructosidase polypeptide in the protoplast incubation medium. These results are consistent with the interpretation that the glycans of {beta}-fructosidase are necessary for its stability, and that in these suspension-cultured cells, the unglycosylated enzyme is degraded during the last stage(s) of secretion, or immediately after its arrival in the wall.« less

Genomic identification of WRKY transcription factors in carrot (Daucus carota) and analysis of evolution and homologous groups for plants.

PubMed

Li, Meng-Yao; Xu, Zhi-Sheng; Tian, Chang; Huang, Ying; Wang, Feng; Xiong, Ai-Sheng

2016-03-15

WRKY transcription factors belong to one of the largest transcription factor families. These factors possess functions in plant growth and development, signal transduction, and stress response. Here, we identified 95 DcWRKY genes in carrot based on the carrot genomic and transcriptomic data, and divided them into three groups. Phylogenetic analysis of WRKY proteins from carrot and Arabidopsis divided these proteins into seven subgroups. To elucidate the evolution and distribution of WRKY transcription factors in different species, we constructed a schematic of the phylogenetic tree and compared the WRKY family factors among 22 species, which including plants, slime mold and protozoan. An in-depth study was performed to clarify the homologous factor groups of nine divergent taxa in lower and higher plants. Based on the orthologous factors between carrot and Arabidopsis, 38 DcWRKY proteins were calculated to interact with other proteins in the carrot genome. Yeast two-hybrid assay showed that DcWRKY20 can interact with DcMAPK1 and DcMAPK4. The expression patterns of the selected DcWRKY genes based on transcriptome data and qRT-PCR suggested that those selected DcWRKY genes are involved in root development, biotic and abiotic stress response. This comprehensive analysis provides a basis for investigating the evolution and function of WRKY genes.

Characterization of a genomic region under selection in cultivated carrot (Daucus carota subsp. sativus) reveals a candidate domestication gene

USDA-ARS?s Scientific Manuscript database

Carrot is one of the most important vegetables worldwide, owing to its capability to develop fleshy, highly nutritious storage roots. It was domesticated ca. 1,100 years ago in Central Asia. No systematic knowledge about the molecular mechanisms involved in the domestication syndrome in carrot are a...

Architecture and Biosynthesis of the Saccharomyces cerevisiae Cell Wall

PubMed Central

Orlean, Peter

2012-01-01

The wall gives a Saccharomyces cerevisiae cell its osmotic integrity; defines cell shape during budding growth, mating, sporulation, and pseudohypha formation; and presents adhesive glycoproteins to other yeast cells. The wall consists of β1,3- and β1,6-glucans, a small amount of chitin, and many different proteins that may bear N- and O-linked glycans and a glycolipid anchor. These components become cross-linked in various ways to form higher-order complexes. Wall composition and degree of cross-linking vary during growth and development and change in response to cell wall stress. This article reviews wall biogenesis in vegetative cells, covering the structure of wall components and how they are cross-linked; the biosynthesis of N- and O-linked glycans, glycosylphosphatidylinositol membrane anchors, β1,3- and β1,6-linked glucans, and chitin; the reactions that cross-link wall components; and the possible functions of enzymatic and nonenzymatic cell wall proteins. PMID:23135325

Mating-Induced Shedding of Cell Walls, Removal of Walls from Vegetative Cells, and Osmotic Stress Induce Presumed Cell Wall Genes in Chlamydomonas1

PubMed Central

Hoffmann, Xenia-Katharina; Beck, Christoph F.

2005-01-01

The first step in sexual differentiation of the unicellular green alga Chlamydomonas reinhardtii is the formation of gametes. Three genes, GAS28, GAS30, and GAS31, encoding Hyp-rich glycoproteins that presumably are cell wall constituents, are expressed in the late phase of gametogenesis. These genes, in addition, are activated by zygote formation and cell wall removal and by the application of osmotic stress. The induction by zygote formation could be traced to cell wall shedding prior to gamete fusion since it was seen in mutants defective in cell fusion. However, it was absent in mutants defective in the initial steps of mating, i.e. in flagellar agglutination and in accumulation of adenosine 3′,5′-cyclic monophosphate in response to this agglutination. Induction of the three GAS genes was also observed when cultures were exposed to hypoosmotic or hyperosmotic stress. To address the question whether the induction seen upon cell wall removal from both gametes and vegetative cells was elicited by osmotic stress, cell wall removal was performed under isosmotic conditions. Also under such conditions an activation of the genes was observed, suggesting that the signaling pathway(s) is (are) activated by wall removal itself. PMID:16183845

Effects of ethylene on gene expression in carrot roots

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

Nichols, S.E.

1984-01-01

To investigate ethylene effects on expression of genetic information, cDNA clones corresponding to ethylene-induced carrot root mRNAs were constructed and isolated. RNA dot blot analysis showed that for the three clones studied peak cytosolic mRNA prevalence occurred at 21 hours of treatment followed thereafter by rapid messenger decay. DNA filter excess hybridization to in vitro synthesized nuclear RNA showed that the ethylene-induced mRNA increase is engendered by transcription of previously quiescent genes. The kinetics and magnitude of changes in mRNA prevalence parallel changes in transcriptional activity; therefore, the ethylene effect is primarily at the level of the transcription. In vivomore » pulse labelling with (/sup 35/S)-methionine showed that between 18 and 27 hours of ethylene treatment a 2.5 fold increase in translational efficiency occurred for one message studied. The resulting protein is the predominant protein synthesized in carrots treated with ethylene for 27 hours. Thus, ethylene exerts multiple regulatory controls on the expression of genetic information.« less

Development of Glycoprotein Capture-Based Label-Free Method for the High-throughput Screening of Differential Glycoproteins in Hepatocellular Carcinoma*

PubMed Central

Chen, Rui; Tan, Yexiong; Wang, Min; Wang, Fangjun; Yao, Zhenzhen; Dong, Liwei; Ye, Mingliang; Wang, Hongyang; Zou, Hanfa

2011-01-01

A robust, reproducible, and high throughput method was developed for the relative quantitative analysis of glycoprotein abundances in human serum. Instead of quantifying glycoproteins by glycopeptides in conventional quantitative glycoproteomics, glycoproteins were quantified by nonglycosylated peptides derived from the glycoprotein digest, which consists of the capture of glycoproteins in serum samples and the release of nonglycopeptides by trypsin digestion of captured glycoproteins followed by two-dimensional liquid chromatography-tandem MS analysis of released peptides. Protein quantification was achieved by comparing the spectrum counts of identified nonglycosylated peptides of glycoproteins between different samples. This method was demonstrated to have almost the same specificity and sensitivity in glycoproteins quantification as capture at glycopeptides level. The differential abundance of proteins present at as low as nanogram per milliliter levels was quantified with high confidence. The established method was applied to the analysis of human serum samples from healthy people and patients with hepatocellular carcinoma (HCC) to screen differential glycoproteins in HCC. Thirty eight glycoproteins were found with substantial concentration changes between normal and HCC serum samples, including α-fetoprotein, the only clinically used marker for HCC diagnosis. The abundance changes of three glycoproteins, i.e. galectin-3 binding protein, insulin-like growth factor binding protein 3, and thrombospondin 1, which were associated with the development of HCC, were further confirmed by enzyme-linked immunosorbent assay. In conclusion, the developed method was an effective approach to quantitatively analyze glycoproteins in human serum and could be further applied in the biomarker discovery for HCC and other cancers. PMID:21474793

First report of 'Candidatus Liberibacter solanacearum' on carrot in Africa

USDA-ARS?s Scientific Manuscript database

In March of 2014, carrot plants (Daucus carota L. var. Mascot) exhibiting symptoms of yellowing, purpling, and curling of leaves, proliferation of shoots, formation of hairy secondary roots, general stunting and plant decline were observed in commercial fields in the Gharb region of Morocco. The sym...

Chromoplasts ultrastructure and estimated carotene content in root secondary phloem of different carrot varieties.

PubMed

Kim, Ji Eun; Rensing, Kim H; Douglas, Carl J; Cheng, Kimberly M

2010-02-01

There have been few studies on quantifying carotenoid accumulation in carrots, and none have taken the comparative approach. The abundance and distribution of carotenes in carrot roots of three varieties, white, orange, and high carotene mass (HCM) were compared using light and transmission electron microscopy (TEM). Light microscopy has indicated that, in all three varieties, carotenes were most abundant in the secondary phloem and this area was selected for further TEM analysis. While carotenes were extracted during the fixation process for TEM, the high-pressure freezing technique we employed preserved the spaces (CS) left behind by the extracted carotene crystals. Chromoplasts from the HCM variety contained significantly (P < 0.05) more CS than chromoplasts from the orange variety. Chromoplasts from the white variety had few or no CS. There was no significant difference between the HCM and orange varieties in the number of chromoplasts per unit area, but the white variety had significantly (P < 0.05) fewer chromoplasts than the other two varieties. A large number of starch-filled amyloplasts was observed in secondary phloem of the white variety but these were not found in the other two varieties. The results from this comparative approach clearly define the subcellular localization of carotenoids in carrot roots and suggest that while the HCM genotype was selectively bred for increased carotene content, this selection did not lead to increased numbers of carotene-containing chromoplasts but rather greater accumulation of carotene per chromoplast. Furthermore, the results confirm that roots of the white carrot variety retain residual amounts of carotene.

Combined effect of water loss and wounding stress on gene activation of metabolic pathways associated with phenolic biosynthesis in carrot

PubMed Central

Becerra-Moreno, Alejandro; Redondo-Gil, Mónica; Benavides, Jorge; Nair, Vimal; Cisneros-Zevallos, Luis; Jacobo-Velázquez, Daniel A.

2015-01-01

The application of postharvest abiotic stresses is an effective strategy to activate the primary and secondary metabolism of plants inducing the accumulation of antioxidant phenolic compounds. In the present study, the effect of water stress applied alone and in combination with wounding stress on the activation of primary (shikimic acid) and secondary (phenylpropanoid) metabolic pathways related with the accumulation of phenolic compound in plants was evaluated. Carrot (Daucus carota) was used as model system for this study, and the effect of abiotic stresses was evaluated at the gene expression level and on the accumulation of metabolites. As control of the study, whole carrots were stored under the same conditions. Results demonstrated that water stress activated the primary and secondary metabolism of carrots, favoring the lignification process. Likewise, wounding stress induced higher activation of the primary and secondary metabolism of carrots as compared to water stress alone, leading to higher accumulation of shikimic acid, phenolic compounds, and lignin. Additional water stress applied on wounded carrots exerted a synergistic effect on the wound-response at the gene expression level. For instance, when wounded carrots were treated with water stress, the tissue showed 20- and 14-fold increases in the relative expression of 3-deoxy-D-arabino-heptulosanate synthase and phenylalanine ammonia-lyase genes, respectively. However, since lignification was increased, lower accumulation of phenolic compounds was detected. Indicatively, at 48 h of storage, wounded carrots treated with water stress showed ~31% lower levels of phenolic compounds and ~23% higher lignin content as compared with wounded controls. In the present study, it was demonstrated that water stress is one of the pivotal mechanism of the wound-response in carrot. Results allowed the elucidation of strategies to induce the accumulation of specific primary or secondary metabolites when plants are

Deglycosylation systematically improves N-glycoprotein identification in liquid chromatography-tandem mass spectrometry proteomics for analysis of cell wall stress responses in Saccharomyces cerevisiae lacking Alg3p.

PubMed

Bailey, Ulla-Maja; Schulz, Benjamin L

2013-04-01

Post-translational modification of proteins with glycosylation is of key importance in many biological systems in eukaryotes, influencing fundamental biological processes and regulating protein function. Changes in glycosylation are therefore of interest in understanding these processes and are also useful as clinical biomarkers of disease. The presence of glycosylation can also inhibit protease digestion and lower the quality and confidence of protein identification by mass spectrometry. While deglycosylation can improve the efficiency of subsequent protease digest and increase protein coverage, this step is often excluded from proteomic workflows. Here, we performed a systematic analysis that showed that deglycosylation with peptide-N-glycosidase F (PNGase F) prior to protease digestion with AspN or trypsin improved the quality of identification of the yeast cell wall proteome. The improvement in the confidence of identification of glycoproteins following PNGase F deglycosylation correlated with a higher density of glycosylation sites. Optimal identification across the proteome was achieved with PNGase F deglycosylation and complementary proteolysis with either AspN or trypsin. We used this combination of deglycosylation and complementary protease digest to identify changes in the yeast cell wall proteome caused by lack of the Alg3p protein, a key component of the biosynthetic pathway of protein N-glycosylation. The cell wall of yeast lacking Alg3p showed specifically increased levels of Cis3p, a protein important for cell wall integrity. Our results showed that deglycosylation prior to protease digestion improved the quality of proteomic analyses even if protein glycosylation is not of direct relevance to the study at hand. Copyright © 2013 Elsevier B.V. All rights reserved.

Cooked carrot volatiles. AEDA and odor activity comparisons. Identification of linden ether as an important aroma component.

PubMed

Buttery, Ron G; Takeoka, Gary R

2013-09-25

MS with GC-RI evidence was found for the presence of linden ether in cooked carrot (Daucus carota). Evaluation of the GC effluent from cooked carrot volatiles using aroma extract dilution analysis (AEDA) found linden ether with the highest flavor dilution (FD) factor. Others with 10-fold lower FD factors were β-ionone, eugenol, the previously unidentified β-damascenone, (E)-2-nonenal, octanal (+ myrcene), and heptanal. All other previously identified volatiles showed lower FD factors. Odor thresholds, concentrations, and odor activity values of previously identified compounds are reviewed. This indicated that at least 20 compounds occur in cooked carrots above their odor thresholds (in water). Compounds showing the highest odor activity values included β-damascenone, (E)-2-nonenal, (E,E)-2,4-decadienal, β-ionone, octanal, (E)-2-decenal, eugenol, and p-vinylguaiacol.

KRE5 Suppression Induces Cell Wall Stress and Alternative ER Stress Response Required for Maintaining Cell Wall Integrity in Candida glabrata

PubMed Central

Sasaki, Masato; Ito, Fumie; Aoyama, Toshio; Sato-Okamoto, Michiyo; Takahashi-Nakaguchi, Azusa; Chibana, Hiroji; Shibata, Nobuyuki

2016-01-01

The maintenance of cell wall integrity in fungi is required for normal cell growth, division, hyphae formation, and antifungal tolerance. We observed that endoplasmic reticulum stress regulated cell wall integrity in Candida glabrata, which possesses uniquely evolved mechanisms for unfolded protein response mechanisms. Tetracycline-mediated suppression of KRE5, which encodes a predicted UDP-glucose:glycoprotein glucosyltransferase localized in the endoplasmic reticulum, significantly increased cell wall chitin content and decreased cell wall β-1,6-glucan content. KRE5 repression induced endoplasmic reticulum stress-related gene expression and MAP kinase pathway activation, including Slt2p and Hog1p phosphorylation, through the cell wall integrity signaling pathway. Moreover, the calcineurin pathway negatively regulated cell wall integrity, but not the reduction of β-1,6-glucan content. These results indicate that KRE5 is required for maintaining both endoplasmic reticulum homeostasis and cell wall integrity, and that the calcineurin pathway acts as a regulator of chitin-glucan balance in the cell wall and as an alternative mediator of endoplasmic reticulum stress in C. glabrata. PMID:27548283

Genomic identification of WRKY transcription factors in carrot (Daucus carota) and analysis of evolution and homologous groups for plants

PubMed Central

Li, Meng-Yao; Xu, Zhi-Sheng; Tian, Chang; Huang, Ying; Wang, Feng; Xiong, Ai-Sheng

2016-01-01

WRKY transcription factors belong to one of the largest transcription factor families. These factors possess functions in plant growth and development, signal transduction, and stress response. Here, we identified 95 DcWRKY genes in carrot based on the carrot genomic and transcriptomic data, and divided them into three groups. Phylogenetic analysis of WRKY proteins from carrot and Arabidopsis divided these proteins into seven subgroups. To elucidate the evolution and distribution of WRKY transcription factors in different species, we constructed a schematic of the phylogenetic tree and compared the WRKY family factors among 22 species, which including plants, slime mold and protozoan. An in-depth study was performed to clarify the homologous factor groups of nine divergent taxa in lower and higher plants. Based on the orthologous factors between carrot and Arabidopsis, 38 DcWRKY proteins were calculated to interact with other proteins in the carrot genome. Yeast two-hybrid assay showed that DcWRKY20 can interact with DcMAPK1 and DcMAPK4. The expression patterns of the selected DcWRKY genes based on transcriptome data and qRT-PCR suggested that those selected DcWRKY genes are involved in root development, biotic and abiotic stress response. This comprehensive analysis provides a basis for investigating the evolution and function of WRKY genes. PMID:26975939

Carrot juice ingestion attenuates high fructose-induced circulatory pro-inflammatory mediators in weanling Wistar rats.

PubMed

Mahesh, Malleswarapu; Bharathi, Munugala; Raja Gopal Reddy, Mooli; Pappu, Pranati; Putcha, Uday Kumar; Vajreswari, Ayyalasomayajula; Jeyakumar, Shanmugam M

2017-03-01

Adipose tissue, an endocrine organ, plays a vital role not only in energy homeostasis, but also in the development and/or progression of various metabolic diseases, such as insulin resistance, type 2 diabetes and non-alcoholic fatty liver disease (NAFLD), via several factors and mechanisms, including inflammation. This study tested, whether carrot juice administration affected the adipose tissue development and its inflammatory status in a high fructose diet-induced rat model. For this purpose, male weanling Wistar rats were divided into four groups and fed either control or high fructose diet of AIN-93G composition with or without carrot juice ingestion for an 8 week period. Administration of carrot juice did not affect the adiposity and cell size of visceral fat depot; retroperitoneal white adipose tissue (RPWAT), which was corroborated with unaltered expression of genes involved in adipogenic and lipogenic pathways. However, it significantly reduced the high fructose diet-induced elevation of plasma free fatty acid (FFA) (P ≤ 0.05), macrophage chemoattractant protein 1 (MCP1) (P ≤ 0.01) and high sensitive C-reactive protein (hsCRP) (P ≤ 0.05) levels. Carrot juice administration attenuated the high fructose diet-induced elevation of levels of circulatory FFA and pro-inflammatory mediators; MCP1 and hsCRP without affecting the adiposity and cell size of visceral fat depot; RPWAT. © 2016 Society of Chemical Industry. © 2016 Society of Chemical Industry.

The Terpene Synthase Gene Family of Carrot (Daucus carota L.): Identification of QTLs and Candidate Genes Associated with Terpenoid Volatile Compounds

PubMed Central

Keilwagen, Jens; Lehnert, Heike; Berner, Thomas; Budahn, Holger; Nothnagel, Thomas; Ulrich, Detlef; Dunemann, Frank

2017-01-01

Terpenes are an important group of secondary metabolites in carrots influencing taste and flavor, and some of them might also play a role as bioactive substances with an impact on human physiology and health. Understanding the genetic and molecular basis of terpene synthases (TPS) involved in the biosynthesis of volatile terpenoids will provide insights for improving breeding strategies aimed at quality traits and for developing specific carrot chemotypes possibly useful for pharmaceutical applications. Hence, a combination of terpene metabolite profiling, genotyping-by-sequencing (GBS), and genome-wide association study (GWAS) was used in this work to get insights into the genetic control of terpene biosynthesis in carrots and to identify several TPS candidate genes that might be involved in the production of specific monoterpenes. In a panel of 85 carrot cultivars and accessions, metabolite profiling was used to identify 31 terpenoid volatile organic compounds (VOCs) in carrot leaves and roots, and a GBS approach was used to provide dense genome-wide marker coverage (>168,000 SNPs). Based on this data, a total of 30 quantitative trait loci (QTLs) was identified for 15 terpenoid volatiles. Most QTLs were detected for the monoterpene compounds ocimene, sabinene, β-pinene, borneol and bornyl acetate. We identified four genomic regions on three different carrot chromosomes by GWAS which are both associated with high significance (LOD ≥ 5.91) to distinct monoterpenes and to TPS candidate genes, which have been identified by homology-based gene prediction utilizing RNA-seq data. In total, 65 TPS candidate gene models in carrot were identified and assigned to known plant TPS subfamilies with the exception of TPS-d and TPS-h. TPS-b was identified as largest subfamily with 32 TPS candidate genes. PMID:29170675

Effect of gamma irradiation on microbial quality of minimally processed carrot and lettuce: A case study in Greater Accra region of Ghana

NASA Astrophysics Data System (ADS)

Frimpong, G. K.; Kottoh, I. D.; Ofosu, D. O.; Larbi, D.

2015-05-01

The effect of ionizing radiation on the microbiological quality on minimally processed carrot and lettuce was studied. The aim was to investigate the effect of irradiation as a sanitizing agent on the bacteriological quality of some raw eaten salad vegetables obtained from retailers in Accra, Ghana. Minimally processed carrot and lettuce were analysed for total viable count, total coliform count and pathogenic organisms. The samples collected were treated and analysed for a 15 day period. The total viable count for carrot ranged from 1.49 to 14.01 log10 cfu/10 g while that of lettuce was 0.70 to 8.5 7 log10 cfu/10 g. It was also observed that total coliform count for carrot was 1.46-7.53 log10 cfu/10 g and 0.14-7.35 log10 cfu/10 g for lettuce. The predominant pathogenic organisms identified were Bacillus cereus, Cronobacter sakazakii, Staphylococcus aureus, and Klebsiella spp. It was concluded that 2 kGy was most effective for medium dose treatment of minimally processed carrot and lettuce.

Morphogenetic responses of cultured totipotent cells of carrot /Daucus carota var. carota/ at zero gravity

NASA Technical Reports Server (NTRS)

Krikorian, A. D.; Steward, F. C.

1978-01-01

An experiment designed to test whether embryos capable of developing from isolated somatic carrot cells could do so under conditions of weightlessness in space was performed aboard the unmanned Soviet biosatellite Kosmos 782 under the auspices of the joint United States-Soviet Biological Satellite Mission. Space flight and weightlessness seem to have had no adverse effects on the induction of embryoids or on the development of their organs. A portion of the crop of carrot plantlets originated in space and grown to maturity were not morphologically different from controls.

Dye-sensitized solar cells fabricated with black raspberry, black carrot and rosella juice

NASA Astrophysics Data System (ADS)

Tekerek, S.; Kudret, A.; Alver, Ü.

2011-10-01

In this work, dye sensitized solar cells (DSSC's) were constructed from black raspberry ( Rubus Ideaus), black carrot ( Daucuscarota L.) and rosella juice ( Hibiscus Sabdariffa L.). In order to fabricate a DSSC the fluorine-doped tin (IV) oxide (FTO) thin films obtained by using spray pyrolysis technique were used as a substrate. TiO2 films on FTO layers were prepared by doctor-blading technique. Platinum-coated counter electrode and liquid Iodide/Iodine electrolyte solution were used to fabricate DSSC's. The efficiencies of solar cells produced with black carrot, rosella and black raspberry juice were calculated as 0.25%, 0.16% and 0.16% respectively, under a sunny day in Kahramanmaraş-Turkey.

Genetic and phenological variation of tocochromanol (vitamin E) content in wild (Daucus carota L. var. carota) and domesticated carrot (D. carota L. var. sativa)

PubMed Central

Luby, Claire H; Maeda, Hiroshi A; Goldman, Irwin L

2014-01-01

Carrot roots (Daucus carota L. var. sativa) produce tocochromanol compounds, collectively known as vitamin E. However, little is known about their types and amounts. Here we determined the range and variation in types and amounts of tocochromanols in a variety of cultivated carrot accessions throughout carrot postharvest storage and reproductive stages and in wild-type roots (Daucus carota L. var. carota). Of eight possible tocochromanol compounds, we detected and quantified α-, and the combined peak for β- and γ- forms of tocopherols and tocotrienols. Significant variation in amounts of tocochromanol compounds was observed across accessions and over time. Large increases in α-tocopherol were noted during both reproductive growth and the postharvest stages. The variation of tocochromanols in carrot root tissue provides useful information for future research seeking to understand the role of these compounds in carrot root tissue or to breed varieties with increased levels of these compounds. PMID:26504534

Aldaulactone - An Original Phytotoxic Secondary Metabolite Involved in the Aggressiveness of Alternaria dauci on Carrot.

PubMed

Courtial, Julia; Hamama, Latifa; Helesbeux, Jean-Jacques; Lecomte, Mickaël; Renaux, Yann; Guichard, Esteban; Voisine, Linda; Yovanopoulos, Claire; Hamon, Bruno; Ogé, Laurent; Richomme, Pascal; Briard, Mathilde; Boureau, Tristan; Gagné, Séverine; Poupard, Pascal; Berruyer, Romain

2018-01-01

Qualitative plant resistance mechanisms and pathogen virulence have been extensively studied since the formulation of the gene-for-gene hypothesis. The mechanisms involved in the quantitative traits of aggressiveness and plant partial resistance are less well-known. Nevertheless, they are prevalent in most plant-necrotrophic pathogen interactions, including the Daucus carota - Alternaria dauci interaction. Phytotoxic metabolite production by the pathogen plays a key role in aggressiveness in these interactions. The aim of the present study was to explore the link between A. dauci aggressiveness and toxin production. We challenged carrot embryogenic cell cultures from a susceptible genotype (H1) and two partially resistant genotypes (I2 and K3) with exudates from A. dauci strains with various aggressiveness levels. Interestingly, A. dauci -resistant carrot genotypes were only affected by exudates from the most aggressive strain in our study (ITA002). Our results highlight a positive link between A. dauci aggressiveness and the fungal exudate cell toxicity. We hypothesize that the fungal exudate toxicity was linked with the amount of toxic compounds produced by the fungus. Interestingly, organic exudate production by the fungus was correlated with aggressiveness. Hence, we further analyzed the fungal organic extract using HPLC, and correlations between the observed peak intensities and fungal aggressiveness were measured. One observed peak was closely correlated with fungal aggressiveness. We succeeded in purifying this peak and NMR analysis revealed that the purified compound was a novel 10-membered benzenediol lactone, a polyketid that we named 'aldaulactone'. We used a new automated image analysis method and found that aldaulactone was toxic to in vitro cultured plant cells at those concentrations. The effects of both aldaulactone and fungal organic extracts were weaker on I2-resistant carrot cells compared to H1 carrot cells. Taken together, our results suggest

Oligosaccharides Released from Milk Glycoproteins Are Selective Growth Substrates for Infant-Associated Bifidobacteria

PubMed Central

Karav, Sercan; Le Parc, Annabelle; Leite Nobrega de Moura Bell, Juliana Maria; Frese, Steven A.; Kirmiz, Nina; Block, David E.; Barile, Daniela

2016-01-01

ABSTRACT Milk, in addition to nourishing the neonate, provides a range of complex glycans whose construction ensures a specific enrichment of key members of the gut microbiota in the nursing infant, a consortium known as the milk-oriented microbiome. Milk glycoproteins are thought to function similarly, as specific growth substrates for bifidobacteria common to the breast-fed infant gut. Recently, a cell wall-associated endo-β-N-acetylglucosaminidase (EndoBI-1) found in various infant-borne bifidobacteria was shown to remove a range of intact N-linked glycans. We hypothesized that these released oligosaccharide structures can serve as a sole source for the selective growth of bifidobacteria. We demonstrated that EndoBI-1 released N-glycans from concentrated bovine colostrum at the pilot scale. EndoBI-1-released N-glycans supported the rapid growth of Bifidobacterium longum subsp. infantis (B. infantis), a species that grows well on human milk oligosaccharides, but did not support growth of Bifidobacterium animalis subsp. lactis (B. lactis), a species which does not. Conversely, B. infantis ATCC 15697 did not grow on the deglycosylated milk protein fraction, clearly demonstrating that the glycan portion of milk glycoproteins provided the key substrate for growth. Mass spectrometry-based profiling revealed that B. infantis consumed 73% of neutral and 92% of sialylated N-glycans, while B. lactis degraded only 11% of neutral and virtually no (<1%) sialylated N-glycans. These results provide mechanistic support that N-linked glycoproteins from milk serve as selective substrates for the enrichment of infant-associated bifidobacteria capable of carrying out the initial deglycosylation. Moreover, released N-glycans were better growth substrates than the intact milk glycoproteins, suggesting that EndoBI-1 cleavage is a key initial step in consumption of glycoproteins. Finally, the variety of N-glycans released from bovine milk glycoproteins suggests that they may serve as

Selenium biofortification of broccoli and carrots grown in soil amended with Se-enriched hyperaccumulator Stanleya pinnata.

PubMed

Bañuelos, Gary S; Arroyo, Irvin; Pickering, Ingrid J; Yang, Soo In; Freeman, John L

2015-01-01

Amending soils with Se-hyperaccumulator plant derived sources of selenium (Se) may be useful for increasing the Se content in food crops in Se-deficient regions of the world. In this study we evaluated total Se and the different chemical species of Se in broccoli and carrots grown in soils amended with ground shoots of the Se-hyperaccumulator Stanleyapinnata. With increasing application rates of S. pinnata, total plant Se concentrations increased to nutritionally ideal levels inside edible parts. Selenium compounds in aqueous extracts were analyzed by SAX-HPLC-ICPMS and identified as a variety of mainly organic-Se forms. Together with bulk Se K-edge X-ray absorption near-edge structure (XANES) analysis performed on broccoli florets, carrot roots and shoots, dried ground S. pinnata, and the amended soil at post-plant, we demonstrate that Se-enriched S. pinnata is valuable as a soil amendment for enriching broccoli and carrots with healthful forms of organic-Se. Published by Elsevier Ltd.

UV-C and hyperoxia abiotic stresses to improve healthiness of carrots: study of combined effects.

PubMed

Formica-Oliveira, Anna Carolina; Martínez-Hernández, Ginés Benito; Aguayo, Encarna; Gómez, Perla A; Artés, Francisco; Artés-Hernández, Francisco

2016-09-01

Phenolic compounds are phytochemicals with high health-promoting properties. Carrot is a vegetable highly worldwide consumed although its phenolic content is low compared to other plant products. The aim of this work was to evaluate changes in phenolic compounds in carrots caused by abiotic stresses. The phenylalanine ammonia-lyase (PAL) activity, phenolic compounds and total antioxidant capacity (TAC) changes during storage up to 72 h at 15 °C after wounding (shredding), 9 kJ UV-C m -2 pretreatment and hyperoxia (80 kPa) conditions of carrots were studied. Shredding and hyperoxia storage induced the highest phenolic compounds and TAC enhancements. Accumulation of phenolic compounds in shredded carrots could be structured in the following phases: 1st phase (<24 h): unchanged phenolic compounds levels with minimum PAL activity; 2nd phase (24-48 h): moderate phenolic increases (≈600-700 mg CAE kg -1 accumulated in 24 h) concurring with the greatest increase of PAL activity; 3nd phase (48-72 h): high phenolic increases (≈1600-2700 mg CAE kg -1 , accumulated in 24 h) while a moderate increment of PAL activity was registered. Although UV-C pretreatment of shreds reduced phenolic accumulation, 600 % increments were still registered in those samples stored under hyperoxia conditions for 72 h. However, the contents of chlorogenic acid at 72 h were 1.4-fold higher in irradiated shreds under hyperoxia compared to the same samples under air conditions.

Raman spectroscopy application in frozen carrot cooked in different ways and the relationship with carotenoids.

PubMed

Camorani, Paolo; Chiavaro, Emma; Cristofolini, Luigi; Paciulli, Maria; Zaupa, Maria; Visconti, Attilio; Fogliano, Vincenzo; Pellegrini, Nicoletta

2015-08-30

Raman spectroscopy, in its confocal micro-Raman variation, has been recently proposed as a spatially resolved method to identify carotenoids in various food matrices, being faster, non-destructive, and avoiding sample extraction, but no data are present in the literature concerning its application to the evaluation of carotenoid pattern changes after thermal treatment of carrots. The effect of three cooking methods (i.e. boiling, steaming and microwaving) was evaluated on frozen carrot, comparing changes on carotenoid profiles measured by means of Raman spectroscopy with their high-performance liquid chromatographic determination and colour. A more pronounced detrimental effect on carotenoids was detected in steamed carrots, in accordance with colour data. Conversely, boiling and, to a lesser extent, microwaving caused an increase in carotenoid concentration. Cooking procedures affected the Raman spectral features of carotenoids, causing a shift of vibration frequencies towards a higher energy, increase in the spectral baseline and peak intensities as well as a broadening of their width, probably in relation to the thermal degradation of longer carotenoids (i.e. the all-trans form) and the isomerization process. In particular, steamed samples showed a significantly higher increase of centre frequency, in accordance with a more pronounced isomerization and changes in colour parameters. This work showed that the evolution of Raman spectral parameters could provide information on carotenoid bioaccessibility for carrots cooked using various methods. This paves the way for a future use of this technique to monitor and optimize cooking processes aimed at maximizing carotenoid bioaccessibility and bioavailability. © 2014 Society of Chemical Industry.

Irradiation treatment of minimally processed carrots for ensuring microbiological safety

NASA Astrophysics Data System (ADS)

Ashraf Chaudry, Muhammad; Bibi, Nizakat; Khan, Misal; Khan, Maazullah; Badshah, Amal; Jamil Qureshi, Muhammad

2004-09-01

Minimally processed fruits and vegetables are very common in developed countries and are gaining popularity in developing countries due to their convenience and freshness. However, minimally processing may result in undesirable changes in colour, taste and appearance due to the transfer of microbes from skin to the flesh. Irradiation is a well-known technology for elimination of microbial contamination. Food irradiation has been approved by 50 countries and is being applied commercially in USA. The purpose of this study was to evaluate the effect of irradiation on the quality of minimally processed carrots. Fresh carrots were peeled, sliced and PE packaged. The samples were irradiated (0, 0.5, 1.0, 2.0, 2.5, 3.0 kGy) and stored at 5°C for 2 weeks. The samples were analyzed for hardness, organoleptic acceptance and microbial load at 0, 7th and 15th day. The mean firmness of the control and all irradiated samples remained between 4.31 and 4.42 kg of force, showing no adverse effect of radiation dose. The effect of storage (2 weeks) was significant ( P< 0.05) with values ranging between 4.28 and 4.39 kg of force. The total bacterial counts at 5°C for non-irradiated and 0.5 kGy irradiated samples were 6.3×10 5 cfu/g, 3.0×10 2 and few colonies(>10) in all other irradiated samples(1.0, 2.0, 2.5 and 3.0 kGy) after 2 weeks storage. No coliform or E. coli were detected in any of the samples (radiated or control) immediately after irradiation and during the entire storage period in minimally processed carrots. A dose of 2.0 kGy completely controlled the fungal and bacterial counts. The irradiated samples (2.0 kGy) were also acceptable sensorially.

Effect of pectin methylesterase on carrot (Daucus carota) juice cloud stability.

PubMed

Schultz, Alison K; Anthon, Gordon E; Dungan, Stephanie R; Barrett, Diane M

2014-02-05

To determine the effect of residual enzyme activity on carrot juice cloud, 0 to 1 U/g pectin methylesterase (PME) was added to pasteurized carrot juice. Cloud stability and particle diameters were measured to quantify juice cloud stability and clarification for 56 days of storage. All levels of PME addition resulted in clarification; higher amounts had a modest effect in causing more rapid clarification, due to a faster increase in particle size. The cloud initially exhibited a trimodal distribution of particle sizes. For enzyme-containing samples, particles in the smallest-sized mode initially aggregated to merge with the second peak over 5-10 days. This larger population then continued to aggregate more slowly over longer times. This observation of a more rapid destabilization process initially, followed by slower subsequent changes in the cloud, was also manifested in measurements of sedimentation extent and in turbidity tests. Optical microscopy showed that aggregation created elongated, fractal particle structures over time.

Identification and characterization of bZIP-type transcription factors involved in carrot (Daucus carota L.) somatic embryogenesis.

PubMed

Guan, Yucheng; Ren, Haibo; Xie, He; Ma, Zeyang; Chen, Fan

2009-10-01

Seed dormancy is an important adaptive trait that enables seeds of many species to remain quiescent until conditions become favorable for germination. Abscisic acid (ABA) plays an important role in these developmental processes. Like dormancy and germination, the elongation of carrot somatic embryo radicles is retarded by sucrose concentrations at or above 6%, and normal growth resumes at sucrose concentrations below 3%. Using a yeast one-hybrid screening system, we isolated two bZIP-type transcription factors, CAREB1 and CAREB2, from a cDNA library prepared from carrot somatic embryos cultured in a high-sucrose medium. Both CAREB1 and CAREB2 were localized to the nucleus, and specifically bound to the ABA response element (ABRE) in the Dc3 promoter. Expression of CAREB2 was induced in seedlings by drought and exogenous ABA application; whereas expression of CAREB1 increased during late embryogenesis, and reduced dramatically when somatic embryos were treated with fluridone, an inhibitor of ABA synthesis. Overexpression of CAREB1 caused somatic embryos to develop slowly when cultured in low-sucrose medium, and retarded the elongation of the radicles. These results indicate that CAREB1 and CAREB2 have similar DNA-binding activities, but play different roles during carrot development. Our results indicate that CAREB1 functions as an important trans-acting factor in the ABA signal transduction pathway during carrot somatic embryogenesis.

Characterization of Centromeric Histone H3 (CENH3) Variants in Cultivated and Wild Carrots (Daucus sp.)

PubMed Central

Dunemann, Frank; Schrader, Otto; Budahn, Holger; Houben, Andreas

2014-01-01

In eukaryotes, centromeres are the assembly sites for the kinetochore, a multi-protein complex to which spindle microtubules are attached at mitosis and meiosis, thereby ensuring segregation of chromosomes during cell division. They are specified by incorporation of CENH3, a centromere specific histone H3 variant which replaces canonical histone H3 in the nucleosomes of functional centromeres. To lay a first foundation of a putative alternative haploidization strategy based on centromere-mediated genome elimination in cultivated carrots, in the presented research we aimed at the identification and cloning of functional CENH3 genes in Daucus carota and three distantly related wild species of genus Daucus varying in basic chromosome numbers. Based on mining the carrot transcriptome followed by a subsequent PCR-based cloning, homologous coding sequences for CENH3s of the four Daucus species were identified. The ORFs of the CENH3 variants were very similar, and an amino acid sequence length of 146 aa was found in three out of the four species. Comparison of Daucus CENH3 amino acid sequences with those of other plant CENH3s as well as their phylogenetic arrangement among other dicot CENH3s suggest that the identified genes are authentic CENH3 homologs. To verify the location of the CENH3 protein in the kinetochore regions of the Daucus chromosomes, a polyclonal antibody based on a peptide corresponding to the N-terminus of DcCENH3 was developed and used for anti-CENH3 immunostaining of mitotic root cells. The chromosomal location of CENH3 proteins in the centromere regions of the chromosomes could be confirmed. For genetic localization of the CENH3 gene in the carrot genome, a previously constructed linkage map for carrot was used for mapping a CENH3-specific simple sequence repeat (SSR) marker, and the CENH3 locus was mapped on the carrot chromosome 9. PMID:24887084

Aldaulactone – An Original Phytotoxic Secondary Metabolite Involved in the Aggressiveness of Alternaria dauci on Carrot

PubMed Central

Courtial, Julia; Hamama, Latifa; Helesbeux, Jean-Jacques; Lecomte, Mickaël; Renaux, Yann; Guichard, Esteban; Voisine, Linda; Yovanopoulos, Claire; Hamon, Bruno; Ogé, Laurent; Richomme, Pascal; Briard, Mathilde; Boureau, Tristan; Gagné, Séverine; Poupard, Pascal; Berruyer, Romain

2018-01-01

Qualitative plant resistance mechanisms and pathogen virulence have been extensively studied since the formulation of the gene-for-gene hypothesis. The mechanisms involved in the quantitative traits of aggressiveness and plant partial resistance are less well-known. Nevertheless, they are prevalent in most plant-necrotrophic pathogen interactions, including the Daucus carota–Alternaria dauci interaction. Phytotoxic metabolite production by the pathogen plays a key role in aggressiveness in these interactions. The aim of the present study was to explore the link between A. dauci aggressiveness and toxin production. We challenged carrot embryogenic cell cultures from a susceptible genotype (H1) and two partially resistant genotypes (I2 and K3) with exudates from A. dauci strains with various aggressiveness levels. Interestingly, A. dauci-resistant carrot genotypes were only affected by exudates from the most aggressive strain in our study (ITA002). Our results highlight a positive link between A. dauci aggressiveness and the fungal exudate cell toxicity. We hypothesize that the fungal exudate toxicity was linked with the amount of toxic compounds produced by the fungus. Interestingly, organic exudate production by the fungus was correlated with aggressiveness. Hence, we further analyzed the fungal organic extract using HPLC, and correlations between the observed peak intensities and fungal aggressiveness were measured. One observed peak was closely correlated with fungal aggressiveness. We succeeded in purifying this peak and NMR analysis revealed that the purified compound was a novel 10-membered benzenediol lactone, a polyketid that we named ‘aldaulactone’. We used a new automated image analysis method and found that aldaulactone was toxic to in vitro cultured plant cells at those concentrations. The effects of both aldaulactone and fungal organic extracts were weaker on I2-resistant carrot cells compared to H1 carrot cells. Taken together, our results

Tunicamycins: translocase-I inhibitors that target bacterial cell wall and mammalian N-glycoproteins. The potential for selective inhibitors

USDA-ARS?s Scientific Manuscript database

Tunicamycins are a heterologous family of nucleoside antibiotics that target the biosynthesis of bacterial peptidoglycan and eukaryotic N-glycoproteins. The mechanism of action is known, with the tunicamycin-Mg2+ complex established as a transition state analog for hexosamine-1-phosphate: prenol pho...

Development and Characterization of Novel SSR Markers in Carrot (Daucus Carota L.) and Their Application for Mapping and Diversity Analysis in Apiaceae

USDA-ARS?s Scientific Manuscript database

Genomic resources in carrot and other Apiaceae are relatively underdeveloped. The availability of a large set of pcr-based codominant markers, such as simple sequence repeats (SSR), would allow integration of the different carrot genetic maps constructed to date (mainly using anonymous dominant mark...

Essential-oil composition of Daucus carota ssp. major (Pastinocello Carrot) and nine different commercial varieties of Daucus carota ssp. sativus fruits.

PubMed

Flamini, Guido; Cosimi, Elena; Cioni, Pier Luigi; Molfetta, Ilaria; Braca, Alessandra

2014-07-01

The chemical composition of the essential oils obtained by hydrodistillation from the pastinocello carrot, Daucus carota ssp. major (Vis.) Arcang. (flowers and achenes), and from nine different commercial varieties of D. carota L. ssp. sativus (achenes) was investigated by GC/MS analyses. Selective breeding over centuries of a naturally occurring subspecies of the wild carrot, D. carota L. ssp. sativus, has produced the common garden vegetable with reduced bitterness, increased sweetness, and minimized woody core. On the other hand, the cultivation of the pastinocello carrot has been abandoned, even if, recently, there has been renewed interest in the development of this species, which risks genetic erosion. The cultivated carrot (D. carota ssp. sativus) and the pastinocello carrot (D. carota ssp. major) were classified as different subspecies of the same species. This close relationship between the two subspecies urged us to compare the chemical composition of their essential oils, to evaluate the differences. The main essential-oil constituents isolated from the pastinocello fruits were geranyl acetate (34.2%), α-pinene (12.9%), geraniol (6.9%), myrcene (4.7%), epi-α-bisabolol (4.5%), sabinene (3.3%), and limonene (3.0%). The fruit essential oils of the nine commercial varieties of D. carota ssp. sativus were very different from that of pastinocello, as also confirmed by multivariate statistical analyses. Copyright © 2014 Verlag Helvetica Chimica Acta AG, Zürich.

Impact of liposomal encapsulation on degradation of anthocyanins of black carrot extract by adding ascorbic acid.

PubMed

Guldiken, Burcu; Gibis, Monika; Boyacioglu, Dilek; Capanoglu, Esra; Weiss, Jochen

2017-03-22

Black carrot anthocyanins are known to be relatively stable because they contain acylated anthocyanins. The degradation of vitamin C (l-ascorbic acid) on anthocyanins is a known fact in beverage systems. In this study, the effects of various liposomal systems, including black carrot extract (0.1%, 0.2%, 0.4% w/w) and lecithin (1%, 2%, 4% w/w), on the color and degradation of anthocyanin in different ascorbic acid (0.01%, 0.025%, 0.05%, 0.1% w/w) concentrations were examined via UV/VIS spectroscopy and visual control of the color. The physical characteristics of the liposomal systems resulted in particle diameters of 41-46 nm and zeta-potentials of (-23)-(-20) mV. The encapsulation efficiencies of the liposomal systems increased up to 50% with increasing lecithin concentrations. The encapsulation of black carrot extract in liposomes enhanced the color and stability of the anthocyanins during storage. This study showed that the degradation of anthocyanins due to ascorbic acid can be reduced by liposomes in aqueous solutions.

[Influence of arbuscular mycorrhizal inoculation on growth and phoxim residue of carrot (Daucus carota L.)].

PubMed

Wang, Fa-Yuan; Chen, Xin; Sun, Xian-Ming; Shi, Zhao-Yong

2010-12-01

A pot culture experiment was carried out to study the influence of arbuscular mycorrhizal (AM) fungi on the growth and phoxim residue of carrot (Daucus carota L). Four levels of phoxim (0, 200, 400, 800 mg x L(-1)) and two AM fungal inocula, Glomus intraradices BEG 141(141), Glomus mosseae BEG 167 (167),and one nonmycorrhizal inoculum (CK), were applied to the sterilized soil. The plants were harvested after 5 months of growth and phoxim was irrigated into the root zone 14 d before plant harvest. Although decreasing with the increase of phoxim dosage, root infection rates of all the mycorrhizal plants were higher than 70%. Phoxim showed no significant dose effect on shoot wet weights and root yields, which were all increased by AM inoculation at four phoxim dosages. Phoxim residues in shoots and roots increased with the increase of phoxim dosage, but decreased by AM inoculation. In general, Glomus intraradices BEG 141 showed more pronounced effects on the growth and phoxim residue of carrot than Glomus mosseae BEG 167 did. Our results show a promising potential of AM fungi in carrot production and controlling pesticide residues.

Improvement of cloud stability, yield and β-carotene content of carrot juice by process modification.

PubMed

Yu, Li Juan; Rupasinghe, H P Vasantha

2013-10-01

This study investigated the effects of three processing factors, acid blanching, centrifugation and dynamic high pressure homogenization, on cloud stability of carrot juice. Results indicated that the optimum processing condition for stabilized carrot juice were with dynamic high pressure homogenization at 100 MPa combined with 2% citric acid blanching at 95-100  for 2 min followed by 2000 r/min centrifugation for 10 min. The improvement of juice yield was also investigated using a pre-treatment of three commercial enzymes: Pectinex 3XL® (pectinase), Celluclast 1.5 L® (cellulase) and Novozyme 188™ (β-glucosidase). The combination of 0.1 g/kg of Pectinex 3XL®, 0.1 g/kg of Celluclast 1.5 L® and 0.1 g/kg of Novozyme 188™ at 50  and pH 4.0 for 90 min was the most effective condition to improve carrot juice yield from 49% to 67%. The enzymatic treatment increased juice total soluble solids from 7.5 to 8.9°Brix and β-carotene content from 21.4 to 33.7 mg/kg.

Developmental localization and the role of hydroxyproline rich glycoproteins during somatic embryogenesis of banana (Musa spp. AAA)

PubMed Central

2011-01-01

Background Hydroxyproline rich glycoproteins (HRGPs) are implicated to have a role in many aspects of plant growth and development but there is limited knowledge about their localization and function during somatic embryogenesis of higher plants. In this study, the localization and function of hydroxyproline rich glycoproteins in embryogenic cells (ECs) and somatic embryos of banana were investigated by using immunobloting and immunocytochemistry with monoclonal JIM11 and JIM20 antibodies as well as by treatment with 3,4-dehydro-L-proline (3,4-DHP, an inhibitor of extensin biosynthesis), and by immunomodulation with the JIM11 antibody. Results Immunofluorescence labelling of JIM11 and JIM20 hydroxyproline rich glycoprotein epitopes was relatively weak in non-embryogenic cells (NECs), mainly on the edge of small cell aggregates. On the other hand, hydroxyproline rich glycoprotein epitopes were found to be enriched in early embryogenic cells as well as in various developmental stages of somatic embryos. Embryogenic cells (ECs), proembryos and globular embryos showed strong labelling of hydroxyproline rich glycoprotein epitopes, especially in their cell walls and outer surface layer, so-called extracellular matrix (ECM). This hydroxyproline rich glycoprotein signal at embryo surfaces decreased and/or fully disappeared during later developmental stages (e.g. pear-shaped and cotyledonary stages) of embryos. In these later developmental embryogenic stages, however, new prominent hydroxyproline rich glycoprotein labelling appeared in tri-cellular junctions among parenchymatic cells inside these embryos. Overall immunofluorescence labelling of late stage embryos with JIM20 antibody was weaker than that of JIM11. Western blot analysis supported the above immunolocalization data. The treatment with 3,4-DHP inhibited the development of embryogenic cells and decreased the rate of embryo germination. Embryo-like structures, which developed after 3,4-DHP treatment showed

A Comparison of 2,4-Dichlorophenoxyacetic Acid Metabolism in Cultured Soybean Cells and in Embryogenic Carrot Cells

PubMed Central

Montague, Michael J.; Enns, Russell K.; Siegel, Ned R.; Jaworski, Ernest G.

1981-01-01

The removal or reduction in concentration of auxin is often a successful method for obtaining morphogenesis in cell cultures of higher plants, such as carrot, but not for soybean. For this reason, the metabolism of one auxin, 2,4-dichlorophenoxyacetic acid (2,4-D), was compared in both carrot and soybean cells. Whereas soybean cells conjugated a high percentage of their 2,4-D to amino acids, carrot cells contained primarily free 2,4-D. Moreover, after long-term exposure to 2,4-D, carrot cells released much more 2,4-D upon transfer to 2,4-D-free (embryogenic) medium than did soybean cells. It appears that the retention of 2,4-D by soybean cells might interfere with subsequent morphogenesis. Because no impairment of 2,4-D efflux was found with short-term exposure to radiolabeled 2,4-D, it was concluded that 2,4-D retention in soybean cells might be due to a time-dependent, metabolic process. The conjugation of 2,4-D to amino acids was shown to be one such time-dependent process. Additionally, the release of 2,4-D from the cells was shown to be due primarily to a loss of free 2,4-D and not 2,4-D-amino acid conjugates. It seems that the greater retention of 2,4-D by soybean cells upon transfer to 2,4-D-free medium is due to greater formation of 2,4-D-amino acid conjugates. PMID:16661722

Genetic structure and domestication of carrot (Daucus carota subsp. sativus L.) (Apiaceae)

USDA-ARS?s Scientific Manuscript database

Analyses of genetic structure and phylogenetic relationships illuminate the origin and domestication of modern crops. Despite being an important world-wide vegetable, the genetic structure and domestication of carrot (Daucus carota L.) is poorly understood. We provide the first such study using a la...

Enhancing Nutraceutical Bioavailability from Raw and Cooked Vegetables Using Excipient Emulsions: Influence of Lipid Type on Carotenoid Bioaccessibility from Carrots.

PubMed

Zhang, Ruojie; Zhang, Zipei; Zou, Liqiang; Xiao, Hang; Zhang, Guodong; Decker, Eric Andrew; McClements, David Julian

2015-12-09

The influence of the nature of the lipid phase in excipient emulsions on the bioaccessibility and transformation of carotenoid from carrots was investigated using a gastrointestinal tract (GIT) model. Excipient emulsions were fabricated using whey protein as an emulsifier and medium-chain triglycerides (MCT), fish oil, or corn oil as the oil phase. Changes in particle size, charge, and microstructure were measured as the carrot-emulsion mixtures were passed through simulated mouth, stomach, and small intestine regions. Carotenoid bioaccessibility depended on the type of lipids used to form the excipient emulsions (corn oil > fish oil ≫ MCT), which was attributed to differences in the solubilization capacity of mixed micelles formed from different lipid digestion products. The transformation of carotenoids was greater for fish oil and corn oil than for MCT, which may have been due to greater oxidation or isomerization. The bioaccessibility of the carotenoids was higher from boiled than raw carrots, which was attributed to greater disruption of the plant tissue facilitating carotenoid release. In conclusion, excipient emulsions are highly effective at increasing carotenoid bioaccessibility from carrots, but lipid type must be optimized to ensure high efficacy.

Fine mapping, transcriptome analysis, and marker development for Y2, the gene that conditions beta-carotene accumulation in carrot (Daucus carota L.)

USDA-ARS?s Scientific Manuscript database

Domesticated carrots, Daucus carota subsp. sativus, are the richest source of beta-carotene in the US diet, which when consumed is converted into vitamin A, an essential component of eye health and immunity. The Y2 locus plays a significant role in beta-carotene accumulation in carrot roots, but a c...

Carrot, Corn, Lettuce and Soybean Nutrient Contents are ...

EPA Pesticide Factsheets

Biochar, the carbon-rich material remaining after pyrolysis of cellulosic and manure feedstocks, has the potential as a soil amendment to sequester carbon and to improve soil water-holding and nutrient properties- thereby enhancing plant growth. However, biochar produced from some feedstocks also could adversely affect crop quality by changing soil pH and reducing nutrients (e.g., Ca, K, Mg, N, Na, and P) in plant tissues. To evaluate effects of biochar on the nutrient quality of four crops, we conducted a greenhouse study using pots with: carrot (Daucus carota cv. Tendersweet), corn (Zea mays, cv. Golden Bantam), lettuce (Lactuca sativa, cv. Black-Seeded Simpson) and soybean (Glycine max cv. Viking 2265). Plants were grown in one of two South Carolina sandy Coastal Plain soils (Norfolk and Coxville Soil Series), along with biochar (1% by weight) produced from pine chips (PC), poultry litter (PL), swine solids (SS), switchgrass (SG), and two blends of pine chips plus poultry litter (PC/PL, 50/50% and 80/20%). Each of the feedstocks and feedstock blends was pyrolyzed at 350, 500, and 700 ̊ C to produce the biochar used to amend the Norfolk and Coxville soils. Effects of biochar on leaf nutrients (% dry weight) statistically varied with species, soil, feedstock and temperature and nutrient. For carrot and lettuce, the PL, PL/PC, and SS biochars generally decreased leaf N, Ca, Mg, and P; while PL and PL/PC increased K and Na. Biochars had little effect on lea

Elimination of Listeria inoculated in ready-to-eat carrots by combination of antimicrobial coating and γ-irradiation

NASA Astrophysics Data System (ADS)

Turgis, Mélanie; Millette, Mathieu; Salmieri, Stéphane; Lacroix, Monique

2012-08-01

A combined treatment of an edible coating composed of trans-cinnamaldehyde (TCN; 0.5% p/p) with γ-irradiation was investigated against Listeria inoculated in peeled mini-carrots. First, the D10 value (γ-irradiation dose required to eliminate 90% of the bacterial population) of TCN was evaluated under air. This treatment resulted in a 3.66-fold increase in relative bacterial radiosensitivity (RBR) as compared to the control without antimicrobial coating. Secondly, the shelf life of mini-carrots during 21 day of storage at 4 °C was studied. Antimicrobial coating containing TCN was assayed in combination with two irradiation doses (0.25 and 0.5 kGy). Results suggested that the inactive coating did not have any antimicrobial effect against Listeria while the coating containing TCN resulted in a 1.29 log reduction in carrots packed under air after 21 days of storage. Hence, these observations indicated that the combination of irradiation with antimicrobial coating played an important role in enhancing the radiosensitization of Listeria to γ-irradiation.

Antioxidants and Antioxidant Capacity of Biofortified Carrots (Daucus Carota, L.) of Various Colors

USDA-ARS?s Scientific Manuscript database

Antioxidants and antioxidant capacity of seven colored carrots were determined. Five anthocyanins, chlorogenic acid, caffeic acid, and four carotenoids, were quantified by HPLC. Total phenolic content was determined by the Folin-Ciocalteau method. Antioxidant capacities of the hydrophilic and hyd...

DcSto: carrot Stowaway-like elements are abundant, diverse, and polymorphic

USDA-ARS?s Scientific Manuscript database

We investigated nine families of Stowaway-like MITEs in the carrot genome, named DcSto1 to DcSto9. All of them were AT-rich and shared a highly conserved 6 bp-long TIR typical for Stowaways. The copy number of DcSto1 elements was estimated as ca. 5,000 per diploid genome. We observed preference for ...

COSII-Based Mapping and Diversity in Potato, Tomato, Sweet Potato and Carrot

USDA-ARS?s Scientific Manuscript database

This poster presentation reports progress on a USDA NRI grant to enhance understanding of and access to the genetic diversity in wild and landrace relatives of tomato, potato, sweet potato, and carrot, and contribute to the same for other euasterid plant species. This is being done with Conserved Or...

Glycoprotein Disease Markers and Single Protein-omics*

PubMed Central

Chandler, Kevin; Goldman, Radoslav

2013-01-01

Glycoproteins are well represented among biomarkers for inflammatory and cancer diseases. Secreted and membrane-associated glycoproteins make excellent targets for noninvasive detection. In this review, we discuss clinically applicable markers of cancer diseases and methods for their analysis. High throughput discovery continues to supply marker candidates with unusual glycan structures, altered glycoprotein abundance, or distribution of site-specific glycoforms. Improved analytical methods are needed to unlock the potential of these discoveries in validated clinical assays. A new generation of targeted quantitative assays is expected to advance the use of glycoproteins in early detection of diseases, molecular disease classification, and monitoring of therapeutic interventions. PMID:23399550

Ammonia transport in the kidney by Rhesus glycoproteins

PubMed Central

Verlander, Jill W.

2014-01-01

Renal ammonia metabolism is a fundamental element of acid-base homeostasis, comprising a major component of both basal and physiologically altered renal net acid excretion. Over the past several years, a fundamental change in our understanding of the mechanisms of renal epithelial cell ammonia transport has occurred, replacing the previous model which was based upon diffusion equilibrium for NH3 and trapping of NH4+ with a new model in which specific and regulated transport of both NH3 and NH4+ across renal epithelial cell membranes via specific membrane proteins is required for normal ammonia metabolism. A major advance has been the recognition that members of a recently recognized transporter family, the Rhesus glycoprotein family, mediate critical roles in renal and extrarenal ammonia transport. The erythroid-specific Rhesus glycoprotein, Rh A Glycoprotein (Rhag), was the first Rhesus glycoprotein recognized as an ammonia-specific transporter. Subsequently, the nonerythroid Rh glycoproteins, Rh B Glycoprotein (Rhbg) and Rh C Glycoprotein (Rhcg), were cloned and identified as ammonia transporters. They are expressed in specific cell populations and membrane domains in distal renal epithelial cells, where they facilitate ammonia secretion. In this review, we discuss the distribution of Rhbg and Rhcg in the kidney, the regulation of their expression and activity in physiological disturbances, the effects of genetic deletion on renal ammonia metabolism, and the molecular mechanisms of Rh glycoprotein-mediated ammonia transport. PMID:24647713

Impact of Cell Wall Composition on Maize Resistance to Pests and Diseases

PubMed Central

Santiago, Rogelio; Barros-Rios, Jaime; Malvar, Rosa A.

2013-01-01

In cereals, the primary cell wall is built of a skeleton of cellulosic microfibrils embedded in a matrix of hemicelluloses and smaller amounts of pectins, glycoproteins and hydroxycinnamates. Later, during secondary wall development, p-coumaryl, coniferyl and sinapyl alcohols are copolymerized to form mixed lignins. Several of these cell wall components show a determinative role in maize resistance to pest and diseases. However, defense mechanisms are very complex and vary among the same plant species, different tissues or even the same tissue at different developmental stages. Thus, it is important to highlight that the role of the cell wall components needs to be tested in diverse genotypes and specific tissues where the feeding or attacking by the pathogen takes place. Understanding the role of cell wall constituents as defense mechanisms may allow modifications of crops to withstand pests and diseases. PMID:23535334

Improving microbiological safety and maintaining sensory and nutritional quality of pre-cut tomato and carrot by gamma irradiation

NASA Astrophysics Data System (ADS)

Mohácsi-Farkas, Cs.; Nyirő-Fekete, B.; Daood, H.; Dalmadi, I.; Kiskó, G.

2014-06-01

Pre-cut tomato and carrot were irradiated with doses of 1.0, 1.5 and 2 kGy. Unirradiated control and irradiated samples were compared organoleptically by a sensory panel. Microbiological analyses were performed directly after irradiation and during post-irradiation storage for 8 days at 5 °C. Ascorbic acid contents, composition of carotenoids and tocopherols were determined. Statistically significant differences of sensory scores between unirradiated and irradiated samples were observed only in the texture of sliced carrots. Total aerobic viable cell counts have been reduced by about two log cycles with 1.5 kGy dose. Total coliforms and moulds were below the detection limit of 15 CFU/g in the irradiated samples during the refrigerated storage. Yeasts were relatively resistant part of the microbiota of pre-cut tomatoes, but 2 kGy dose reduced them below the detection limit. In pre-cut tomatoes, alpha-tocopherol and some carotenoids seemed to be the most radio-sensitive losing approximately one-third of their original concentrations at the dose of 2 kGy. At this dose tocopherols and the level of ascorbic acid decreased also one-third of the initial level in sliced carrots. Additional experiments were conducted to study the effect of irradiation and storage on the population of Listeria monocytogenes and Listeria innocua artificially inoculated on cut tomato and carrot. Cell numbers of both test organisms decreased by at least two log-cycles as an effect of 1 kGy dose. Our studies confirmed earlier findings on a temporary antilisterial effect of freshly cut carrot tissue. No re-growth of Listeria was observed during the studied storage period. The results of these studies suggest that irradiation with 1 kGy gamma rays could improve sufficiently the microbiological safety of the investigated pre-cut produce to satisfy the requirement of low microbial raw diets with acceptable nutritional quality and without diminishing significantly the organoleptic parameters of the

Serum a- and b-carotene concentrations qualitatively respond to sustained carrot feeding

USDA-ARS?s Scientific Manuscript database

b-Carotene is a predominant source of vitamin A in developing countries. Genetically selected ‘‘high carotene’’ carrots could have an impact on the vitamin A and antioxidant status of people if widely adopted. A 3 3 3 crossover study in humans (n = 10) evaluated the difference in uptake and clearanc...

Solid-State Fermentation of Carrot Pomace for the Production of Inulinase by Penicillium oxalicum BGPUP-4.

PubMed

Singh, Ram Sarup; Chauhan, Kanika; Singh, Jagroop; Pandey, Ashok; Larroche, Christian

2018-03-01

Inulinases are an important class of industrial enzymes which are used for the production of high-fructose syrup and fructooligosaccharides. Inulin, a polyfructan, is generally employed for the production of inulinase, which is a very expensive substrate. A number of agroindustrial residues have been used for cost-effective production of inulinases. In the present study, carrot pomace was selected as a substrate for the production of inulinase by Penicillium oxalicum BGPUP-4 in solid-state fermentation. Carrot pomace is one of the good substrates for bioprocesses, because it is rich in soluble and insoluble carbohydrates. A central composite rotatable design (CCRD) used in response surface methodology was employed for the optimal production of inulinase from carrot pomace. Using CCRD, 15 runs were practiced to optimize the range of three independent variables: moisture content (70-90%), incubation time (4-6 days) and pH (5.0-7.0) for inulinase production. Carrot pomace supplemented with 0.5% inulin as an inducer, 0.2% NH 4 H 2 PO 4 , 0.2% NaNO 3 , 0.2% KH 2 PO 4 , 0.05% MgSO 4 ·7H 2 O and 0.001% FeSO 4 ·7H 2 O was used for the production of inulinase in solid-state fermentation at 30 °C. Inulinase production (322.10 IU per g of dry substrate) was obtained under the optimized conditions, i.e . moisture content of 90%, incubation time 4 days and pH=7.0. The corresponding inulinase/invertase (I/S) ratio (3.38) was also high, which indicates the inulolytic nature of the enzyme. Multiple correlation coefficients R for inulinase production and I/S ratio were 0.9995 and 0.9947, respectively. The R value very close to one indicates an excellent correlation between experimental and predicted results.

Solid-State Fermentation of Carrot Pomace for the Production of Inulinase by Penicillium oxalicum BGPUP-4

PubMed Central

2018-01-01

Summary Inulinases are an important class of industrial enzymes which are used for the production of high-fructose syrup and fructooligosaccharides. Inulin, a polyfructan, is generally employed for the production of inulinase, which is a very expensive substrate. A number of agroindustrial residues have been used for cost-effective production of inulinases. In the present study, carrot pomace was selected as a substrate for the production of inulinase by Penicillium oxalicum BGPUP-4 in solid-state fermentation. Carrot pomace is one of the good substrates for bioprocesses, because it is rich in soluble and insoluble carbohydrates. A central composite rotatable design (CCRD) used in response surface methodology was employed for the optimal production of inulinase from carrot pomace. Using CCRD, 15 runs were practiced to optimize the range of three independent variables: moisture content (70-90%), incubation time (4-6 days) and pH (5.0-7.0) for inulinase production. Carrot pomace supplemented with 0.5% inulin as an inducer, 0.2% NH4H2PO4, 0.2% NaNO3, 0.2% KH2PO4, 0.05% MgSO4·7H2O and 0.001% FeSO4·7H2O was used for the production of inulinase in solid-state fermentation at 30 °C. Inulinase production (322.10 IU per g of dry substrate) was obtained under the optimized conditions, i.e. moisture content of 90%, incubation time 4 days and pH=7.0. The corresponding inulinase/invertase (I/S) ratio (3.38) was also high, which indicates the inulolytic nature of the enzyme. Multiple correlation coefficients R for inulinase production and I/S ratio were 0.9995 and 0.9947, respectively. The R value very close to one indicates an excellent correlation between experimental and predicted results. PMID:29795994

Abscisic acid regulation of DC8, a carrot embryonic gene. [Daucus carota

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

Hatzopoulos, P.; Fong, F.; Sung, Z.R.

1990-10-01

DC8 encodes a hydrophylic 66 kilodalton protein located in the cytoplasm and cell walls of carrot (Daucus carota) embryo and endosperm. During somatic embryogenesis, the levels of DC8 mRNA and protein begin to increase 5 days after removal of auxin. To study the role of abscisic acid (ABA) in the regulation of DC8 gene, fluridone, 1-methyl-3-phenyl,-5(3-trifluoro-methyl-phenyl)-4(1H)-pyridinone, was used to inhibit the endogenous ABA content of the embryos. Fluridone, 50 micrograms per milliliter, effectively inhibits the accumulation of ABA in globular-tage embryos. Western and Northern analysis show that when fluridone is added to the culture medium DC8 protein and mRNA decreasemore » to very low levels. ABA added to fluridone supplemented culture media restores the DC8 protein and mRNA to control levels. Globular-stage embryos contain 0.9 to 1.4 {times} 10{sup {minus}7} molar ABA while 10{sup {minus}6} molar exogenously supplied ABA is the optimal concentration for restoration of DC8 protein accumulation in fluridone-treated embryos. The mRNA level is increased after 15 minutes of ABA addition and reaches maximal levels by 60 minutes. Evidence is presented that, unlike other ABA-regulated genes, DC8 is not induced in nonembryonic tissues via desiccation nor addition of ABA.« less

Characterization of chemical, biological and antiproliferative properties of fermented black carrot juice,shalgam

USDA-ARS?s Scientific Manuscript database

Shalgam juice is a dark red-colored and sour fermented beverage produced and consumed in Turkey. The main ingredient of shalgam juice is black carrot, which is rich in anthocyanins. In this study, commercially available shalgam juice was characterized by determining its chemical composition and anti...

Genotyping-by-sequencing reveals the origin of the Tunisian relatives of cultivated carrot

USDA-ARS?s Scientific Manuscript database

Knowledge of the morphological diversity of a germplasm collection is fundamental for genebank managers and plant breeders. The main objective of this work was to characterize 33 landraces of carrot from 13 different regions of Tunisia, based on 34 agro-morphological characters related to leaves and...

Effect of simultaneous infrared dry-blanching and dehydration on quality characteristics of carrot slices

USDA-ARS?s Scientific Manuscript database

This study investigated the effects of various processing parameters on carrot slices exposed to infrared (IR) radiation heating for achieving simultaneous infrared dry-blanching and dehydration (SIRDBD). The investigated parameters were product surface temperature, slice thickness and processing ti...

Carrot injury and yield response to ozone

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

Bennett, J.P.; Oshima, R.J.

1976-11-01

Container-grown plants of carrot (Daucus carota L.) exposed intermittently to 0.19 or 0.25 ppm ozone throughout their growth increased in plant height and total number of leaves in spite of the development of chlorotic leaves. Leaf dry weight was unaffected by ozone, but root dry matter decreased 32 to 46%. As a result, the root weight/total dry weight ration and root/shoot ratio declined significantly in the presence of ozone. A regression of root dry weight on chlorotic lead dry weight explained 35% of the root loss and predicted that 1.5 g of root tissue is lost for every g ofmore » chlorotic leaf dry weight casued by ozone injury.« less

Hyperostotic polyarthropathy in a rabbit--a suspected case of chronic hypervitaminosis A from a diet of carrots.

PubMed

Frater, J

2001-09-01

Chronic hypervitaminosis A can occur in many species after excessive dietary intake of Vitamin A (retinol). The most common presentation of chronic hypervitaminosis A is a polyarthropathy with hyperostosis and ankylosis of various joints. This case report describes a probable case of naturally occurring hypervitaminosis A-induced polyarthropathy in a rabbit after chronic ingestion of a diet made up almost exclusively of carrots. Carrots do not contain retinol, but are rich in provitamin A (or beta-carotene). Rabbits are unique in that they can convert 100% of dietary beta-carotene into retinol. A syndrome of naturally occurring hypervitaminosis A-induced polyarthropathy has not been described in a rabbit before.

21 CFR 866.5420 - Alpha-1-glycoproteins immunological test system.

Code of Federal Regulations, 2010 CFR

2010-04-01

... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Alpha-1-glycoproteins immunological test system....5420 Alpha-1-glycoproteins immunological test system. (a) Identification. An alpha-1-glycoproteins... alpha-1-glycoproteins (a group of plasma proteins found in the alpha-1 group when subjected to...

The inside and outside: topological issues in plant cell wall biosynthesis and the roles of nucleotide sugar transporters.

PubMed

Temple, Henry; Saez-Aguayo, Susana; Reyes, Francisca C; Orellana, Ariel

2016-09-01

The cell wall is a complex extracellular matrix composed primarily of polysaccharides. Noncellulosic polysaccharides, glycoproteins and proteoglycans are synthesized in the Golgi apparatus by glycosyltransferases (GTs), which use nucleotide sugars as donors to glycosylate nascent glycan and glycoprotein acceptors that are subsequently exported to the extracellular space. Many nucleotide sugars are synthesized in the cytosol, leading to a topological issue because the active sites of most GTs are located in the Golgi lumen. Nucleotide sugar transporters (NSTs) overcome this problem by translocating nucleoside diphosphate sugars from the cytosol into the lumen of the organelle. The structures of the cell wall components synthesized in the Golgi are diverse and complex; therefore, transporter activities are necessary so that the nucleotide sugars can provide substrates for the GTs. In this review, we describe the topology of reactions involved in polysaccharide biosynthesis in the Golgi and focus on the roles of NSTs as well as their impacts on cell wall structure when they are altered. © The Author 2016. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Solubilization of glycoproteins of envelope viruses by detergents

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

Berezin, V.E.; Zaides, V.M.; Artamsnov, A.F.

1986-11-20

The action of a number of known ionic and nonionic detergents, as well as the new nonionic detergent MESK, on envelope viruses was investigated. It was shown that the nonionic detergents MESK, Triton X-100, and octyl-..beta..-D-glucopyranoside selectively solubilize the outer glycoproteins of the virus particles. The nonionic detergent MESK has the mildest action. Using MESK, purified glycoproteins of influenza, parainfluenza, Venezuelan equine encephalomyelitis, vesicular stomatitis, rabies, and herpes viruses were obtained. The procedure for obtaining glycoproteins includes incubation of the virus suspension with the detergent MESK, removal of subvirus structures by centrifuging, and purification of glycoproteins from detergents by dialysis.more » Isolated glycoproteins retain a native structure and biological activity and possess high immunogenicity. The detergent MESK is promising for laboratory tests and with respect to the production of subunit vaccines.« less

Antioxidant characterization and sensory evaluation during storage of ultraviolet-B light exposed baby carrots (abstract)

USDA-ARS?s Scientific Manuscript database

Baby carrot processing induces wounding stress activation of phenylalanine ammonia-lyase (PAL), enhancing its nutrient content by increasing synthesis of secondary metabolites. Ultraviolet-B (UV-B) exposure further promotes the formation of soluble phenolic compounds, significantly increasing antiox...

Influence of unit operations on the levels of polyacetylenes in minimally processed carrots and parsnips: An industrial trial.

PubMed

Koidis, Anastasios; Rawson, Ashish; Tuohy, Maria; Brunton, Nigel

2012-06-01

Carrots and parsnips are often consumed as minimally processed ready-to-eat convenient foods and contain in minor quantities, bioactive aliphatic C17-polyacetylenes (falcarinol, falcarindiol, falcarindiol-3-acetate). Their retention during minimal processing in an industrial trial was evaluated. Carrot and parsnips were prepared in four different forms (disc cutting, baton cutting, cubing and shredding) and samples were taken in every point of their processing line. The unit operations were: peeling, cutting and washing with chlorinated water and also retention during 7days storage was evaluated. The results showed that the initial unit operations (mainly peeling) influence the polyacetylene retention. This was attributed to the high polyacetylene content of their peels. In most cases, when washing was performed after cutting, less retention was observed possibly due to leakage during tissue damage occurred in the cutting step. The relatively high retention during storage indicates high plant matrix stability. Comparing the behaviour of polyacetylenes in the two vegetables during storage, the results showed that they were slightly more retained in parsnips than in carrots. Unit operations and especially abrasive peeling might need further optimisation to make them gentler and minimise bioactive losses. Copyright © 2011 Elsevier Ltd. All rights reserved.

Occurrence and molecular detection of Spiroplasma citri in carrots and Circulifer tenellus in Mexico

USDA-ARS?s Scientific Manuscript database

In the fall of 2014, carrot plants in Zacatecas, Mexico, were found with yellow, brown (chlorotic), and/or purple-colored leaves, small and/or rolled leaves, and hairy, deformed, and/or small roots. Molecular diagnostics of these symptomatic plants failed to detect phytoplasmas in these samples, bu...

Ultraviolet-C light effect on physicochemical, bioactive, microbiological, and sensorial characteristics of carrot (Daucus carota) beverages.

PubMed

Hernández-Carranza, Paola; Ruiz-López, Irving Israel; Pacheco-Aguirre, Francisco Manuel; Guerrero-Beltrán, José Ángel; Ávila-Sosa, Raúl; Ochoa-Velasco, Carlos Enrique

2016-09-01

The aim of this research was to evaluate the effect of ultraviolet-C light on physicochemical, bioactive, microbial, and sensory characteristics of carrot beverages. Beverages were formulated with different concentrations of carrot juice (60, 80, and 100% [v/v]) and treated with ultraviolet-C light at different flow rates (0, 0.5, 3.9, and 7.9 mL s(-1)) and times (5, 10, 15, 20, and 30 min), equivalent to ultraviolet-C dosages of 13.2, 26.4, 39.6, 52.8, and 79.2 J cm(-2) Total soluble solids, pH, and titratable acidity were not affected by the ultraviolet-C light treatment. Ultraviolet-C light significantly affected (p < 0.05) color parameters of pure juice; however, at low concentration of juice, total color change was slightly affected (ΔE = 2.0 ± 0.7). Phenolic compounds (4.1 ± 0.1, 5.2 ± 0.2, and 8.6 ± 0.3 mg of GAE 100 mL(-1) of beverage with 60, 80, and 100% of juice, respectively) and antioxidant capacity (6.1 ± 0.4, 8.5 ± 0.4, and 9.4 ± 0.3 mg of Trolox 100 mL(-1) of beverage with 60, 80, and 100% of juice, respectively) of carrot beverages were not affected by ultraviolet-C light treatment. Microbial kinetics showed that mesophiles were mostly reduced at high flow rates in carrot beverages with 60% of juice. Maximum logarithmic reductions for mesophiles and total coliforms were 3.2 ± 0.1 and 2.6 ± 0.1, respectively, after 30 min of ultraviolet-C light processing. Beverages were well accepted (6-7) by judges who did not perceive the difference between untreated and Ultraviolet-C light treated beverages. © The Author(s) 2016.

Transgenic carrot expressing fusion protein comprising M. tuberculosis antigens induces immune response in mice.

PubMed

Permyakova, Natalia V; Zagorskaya, Alla A; Belavin, Pavel A; Uvarova, Elena A; Nosareva, Olesya V; Nesterov, Andrey E; Novikovskaya, Anna A; Zav'yalov, Evgeniy L; Moshkin, Mikhail P; Deineko, Elena V

2015-01-01

Tuberculosis remains one of the major infectious diseases, which continues to pose a major global health problem. Transgenic plants may serve as bioreactors to produce heterologous proteins including antibodies, antigens, and hormones. In the present study, a genetic construct has been designed that comprises the Mycobacterium tuberculosis genes cfp10, esat6 and dIFN gene, which encode deltaferon, a recombinant analog of the human γ-interferon designed for expression in plant tissues. This construct was transferred to the carrot (Daucus carota L.) genome by Agrobacterium-mediated transformation. This study demonstrates that the fusion protein CFP10-ESAT6-dIFN is synthesized in the transgenic carrot storage roots. The protein is able to induce both humoral and cell-mediated immune responses in laboratory animals (mice) when administered either orally or by injection. It should be emphasized that M. tuberculosis antigens contained in the fusion protein have no cytotoxic effect on peripheral blood mononuclear cells.

Home-based preparation approaches altered the availability of health beneficial components from carrots and blueberries

USDA-ARS?s Scientific Manuscript database

This study investigated the effects of different home food preparation methods on the availability of the total phenolic contents (TPC) and radical scavenging components, as well as the selected health beneficial compounds from fresh blueberries and carrots. High performance liquid chromatography (...

Gulose as a constituent of a glycoprotein.

PubMed

Mengele, R; Sumper, M

1992-02-17

The aldohexose gulose was identified as a constituent of a hydroxyproline-rich glycopeptide derived from the glycoprotein SSG 185. This glycoprotein is part of the extracellular matrix of the green alga Volvox carteri. The gulose residue occupies a terminal position in the corresponding saccharide.

Formation of norisoprenoid flavor compounds in carrot (Daucus carota L.) roots: characterization of a cyclic-specific carotenoid cleavage dioxygenase 1 gene.

PubMed

Yahyaa, Mosaab; Bar, Einat; Dubey, Neeraj Kumar; Meir, Ayala; Davidovich-Rikanati, Rachel; Hirschberg, Joseph; Aly, Radi; Tholl, Dorothea; Simon, Philipp W; Tadmor, Yaakov; Lewinsohn, Efraim; Ibdah, Mwafaq

2013-12-18

Carotenoids are isoprenoid pigments that upon oxidative cleavage lead to the production of norisoprenoids that have profound effect on flavor and aromas of agricultural products. The biosynthetic pathway to norisoprenoids in carrots (Daucus carota L.) is still largely unknown. We found the volatile norisoprenoids farnesylacetone, α-ionone, and β-ionone accumulated in Nairobi, Rothild, and Purple Haze cultivars but not in Yellowstone and Creme de Lite in a pattern reflecting their carotenoid content. A cDNA encoding a protein with carotenoid cleavage dioxygenase activity, DcCCD1, was identified in carrot and was overexpressed in Escherichia coli strains previously engineered to produce different carotenoids. The recombinant DcCCD1 enzyme cleaves cyclic carotenes to generate α- and β-ionone. No cleavage products were found when DcCCD1 was co-expressed in E. coli strains accumulating non-cyclic carotenoids, such as phytoene or lycopene. Our results suggest a role for DcCCD1 in carrot flavor biosynthesis.

Retention of glucose units added by the UDP-GLC:glycoprotein glucosyltransferase delays exit of glycoproteins from the endoplasmic reticulum

PubMed Central

1995-01-01

It has been proposed that the UDP-Glc:glycoprotein glucosyltransferase, an endoplasmic reticulum enzyme that only glucosylates improperly folded glycoproteins forming protein-linked Glc1Man7-9-GlcNAc2 from the corresponding unglucosylated species, participates together with lectin- like chaperones that recognize monoglucosylated oligosaccharides in the control mechanism by which cells only allow passage of properly folded glycoproteins to the Golgi apparatus. Trypanosoma cruzi cells were used to test this model as in trypanosomatids addition of glucosidase inhibitors leads to the accumulation of only monoglucosylated oligosaccharides, their formation being catalyzed by the UDP- Glc:glycoprotein glucosyltransferase. In all other eukaryotic cells the inhibitors produce underglycosylation of proteins and/or accumulation of oliogosaccharides containing two or three glucose units. Cruzipain, a lysosomal proteinase having three potential N-glycosylation sites, two at the catalytic domain and one at the COOH-terminal domain, was isolated in a glucosylated form from cells grown in the presence of the glucosidase II inhibitor 1-deoxynojirimycin. The oligosaccharides present at the single glycosylation site of the COOH-terminal domain were glucosylated in some cruzipain molecules but not in others, this result being consistent with an asynchronous folding of glycoproteins in the endoplasmic reticulum. In spite of not affecting cell growth rate or the cellular general metabolism in short and long term incubations, 1-deoxynojirimycin caused a marked delay in the arrival of cruzipain to lysosomes. These results are compatible with the model proposed by which monoglucosylated glycoproteins may be transiently retained in the endoplasmic reticulum by lectin-like anchors recognizing monoglucosylated oligosaccharides. PMID:7642696

Carotenoids are more bioavailable from papaya than from tomato and carrot in humans: a randomised cross-over study

PubMed Central

Schweiggert, Ralf M.; Kopec, Rachel E.; Villalobos-Gutierrez, Maria G.; Högel, Josef; Quesada, Silvia; Esquivel, Patricia; Schwartz, Steven J.; Carle, Reinhold

2014-01-01

Carrot, tomato and papaya represent important dietary sources of β-carotene and lycopene. The main objective of the present study was to compare the bioavailability of carotenoids from these food sources in healthy human subjects. A total of sixteen participants were recruited for a randomised cross-over study. Test meals containing raw carrots, tomatoes and papayas were adjusted to deliver an equal amount of β-carotene and lycopene. For the evaluation of bioavailability, TAG-rich lipoprotein (TRL) fractions containing newly absorbed carotenoids were analysed over 9.5 h after test meal consumption. The bioavailability of β-carotene from papayas was approximately three times higher than that from carrots and tomatoes, whereas differences in the bioavailability of β-carotene from carrots and tomatoes were insignificant. Retinyl esters appeared in the TRL fractions at a significantly higher concentration after the consumption of the papaya test meal. Similarly, lycopene was approximately 2.6 times more bioavailable from papayas than from tomatoes. Furthermore, the bioavailability of β-cryptoxanthin from papayas was shown to be 2.9 and 2.3 times higher than that of the other papaya carotenoids β-carotene and lycopene, respectively. The morphology of chromoplasts and the physical deposition form of carotenoids were hypothesised to play a major role in the differences observed in the bioavailability of carotenoids from the foods investigated. Particularly, the liquid-crystalline deposition of β-carotene and the storage of lycopene in very small crystalloids in papayas were found to be associated with their high bioavailability. In conclusion, papaya was shown to provide highly bioavailable β-carotene, β-cryptoxanthin and lycopene and may represent a readily available dietary source of provitamin A for reducing the incidence of vitamin A deficiencies in many subtropical and tropical developing countries. PMID:23931131

Analgesic effects of glycoproteins from Panax ginseng root in mice.

PubMed

Wang, Ying; Chen, Yinghong; Xu, Hong; Luo, Haoming; Jiang, Ruizhi

2013-07-30

The root of Panax ginseng C.A. Mey has various beneficial pharmacological effects. The present study aimed to evaluate the analgesic activities of glycoproteins from the root of Panax ginseng C.A. Mey in mice. Glycoproteins were isolated and purified from the root of Panax ginseng C.A. Mey. Physicochemical properties and molecular mass were determined by chemical assay and HPLC. Acetic acid-induced writhing and hot-plate tests were employed to study the analgesic effect of glycoproteins and compared with that of aspirin or morphine. The locomotor activity was tested in mice by using actophometer. Four glycoproteins were obtained. The glycoproteins which protein content was the highest (73.04%) displayed dose-dependent analgesic effect. In writhing test, the glycoproteins significantly inhibited writhes (P<0.001) at the dose of 20 mg/kg by intraperitoneal injection. In hot-plate test, only at the dose of 20 mg/kg prolong the hot-plate latency (P<0.05, at 30 min). In the locomotor activity test, the glycoproteins were significant decrease of motility counts at the dose of 20 and 40 mg/kg. These findings collectively indicate that the glycoproteins from the root of Panax ginseng C.A. Mey exhibited significant analgesic activities and the proteins were the active site, providing evidence for its pharmacal use. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

Determining P-glycoprotein-drug interactions: evaluation of reconstituted P-glycoprotein in a liposomal system and LLC-MDR1 polarized cell monolayers.

PubMed

Melchior, Donald L; Sharom, Frances J; Evers, Raymond; Wright, George E; Chu, Joseph W K; Wright, Stephen E; Chu, Xiaoyan; Yabut, Jocelyn

2012-03-01

P-Glycoprotein (ABCB1, MDR1) is a multidrug efflux pump that is a member of the ATP-binding cassette (ABC) superfamily. Many drugs in common clinical use are either substrates or inhibitors of this transporter. Quantitative details of P-glycoprotein inhibition by pharmaceutical agents are essential for assessment of their pharmacokinetic behavior and prevention of negative patient reactions. Cell-based systems have been widely used for determination of drug interactions with P-glycoprotein, but they suffer from several disadvantages, and results are often widely variable between laboratories. We aimed to demonstrate that a novel liposomal system employing contemporary biochemical methodologies could measure the ability of clinically used drugs to inhibit the P-glycoprotein pump. To accomplish this we compared results with those of cell-based approaches. Purified transport-competent hamster Abcb1a P-glycoprotein was reconstituted into a unilamellar liposomal system, Fluorosome-trans-pgp, whose aqueous interior contains fluorescent drug sensors. This provides a well-defined system for measuring P-glycoprotein transport inhibition by test drugs in real time using rapid fluorescence-based technology. Inhibition of ATP-driven transport by Fluorosome-trans-pgp employed a panel of 46 representative drugs. Resulting IC50 values correlated well (r2=0.80) with Kd values for drug binding to purified P-glycoprotein. They also showed a similar trend to transport inhibition data obtained using LLC-MDR1 cell monolayers. Fluorosome-trans-pgp IC50 values were in agreement with published results of digoxin drug-drug interaction studies in humans. This novel approach using a liposomal system and fluorescence-based technology is shown to be suitable to study whether marketed drugs and drug candidates are P-glycoprotein inhibitors. The assay is rapid, allowing a 7-point IC50 determination in <6 min, and requires minimal quantities of test drug. The method is amenable to robotics and

Determining P-glycoprotein-drug interactions: evaluation of reconstituted P-glycoprotein in a liposomal system and LLC-MDR1 polarized cell monolayers

PubMed Central

Melchior, Donald L.; Sharom, Frances J.; Evers, Raymond; Wright, George E.; Chu, Joseph W.K.; Wright, Stephen E.; Chu, Xiaoyan; Yabut, Jocelyn

2012-01-01

Introduction P-Glycoprotein (ABCB1, MDR1) is a multidrug efflux pump that is a member of the ATP-binding cassette (ABC) superfamily. Many drugs in common clinical use are either substrates or inhibitors of this transporter. Quantitative details of P-glycoprotein inhibition by pharmaceutical agents are essential for assessment of their pharmacokinetic behavior and prevention of negative patient reactions. Cell-based systems have been widely used for determination of drug interactions with P-glycoprotein, but they suffer from several disadvantages, and results are often widely variable between laboratories. We aimed to demonstrate that a novel liposomal system employing contemporary biochemical methodologies could measure the ability of clinically used drugs to inhibit the P-glycoprotein pump. To accomplish this we compared results with those of cell-based approaches. Methods Purified transport-competent hamster Abcb1a P-glycoprotein was reconstituted into a unilamellar liposomal system, Fluorosome-trans-pgp, whose aqueous interior contains fluorescent drug sensors. This provides a well-defined system for measuring P-glycoprotein transport inhibition by test drugs in real time using rapid fluorescence-based technology. Results Inhibition of ATP-driven transport by Fluorosome-trans-pgp employed a panel of 46 representative drugs. Resulting IC50 values correlated well (r2 = 0.80) with Kd values for drug binding to purified P-glycoprotein. They also showed a similar trend to transport inhibition data obtained using LLC-MDR1 cell monolayers. Fluorosome-trans-pgp IC50 values were in agreement with published results of digoxin drug-drug interaction studies in humans. Discussion This novel approach using a liposomal system and fluorescence-based technology is shown to be suitable to study whether marketed drugs and drug candidates are P-glycoprotein inhibitors. The assay is rapid, allowing a 7-point IC50 determination in <6 minutes, and requires minimal quantities of test

Transcriptional regulation of chlorogenic acid biosynthesis in carrot root slices exposed to UV-B light

USDA-ARS?s Scientific Manuscript database

Orange carrots are well known for their nutritional value as producers of ß-carotene, a Vitamin A precursor. Lesser known, is their ability to accumulate antioxidants such as chlorogenic acid. Chlorogenic acid is produced through the same biosynthetic pathway that produces lignins, anthocyanins, f...

Glycoprotein synthesis

DOEpatents

Schultz, Peter G.; Wang, Lei; Zhang, Zhiwen

2005-08-09

Methods for making glycoproteins, both in vitro and in vivo, are provided. One method involves incorporating an unnatural amino acid into a protein and attaching one or more saccharide moieties to the unnatural amino acid. Another method involves incorporating an unnatural amino acid that includes a saccharide moiety into a protein. Proteins made by both methods can be further modified with additional sugars.

Glycoprotein synthesis

DOEpatents

Schultz, Peter G.; Wang, Lei; Zhang, Zhiwen

2010-11-16

Methods for making glycoproteins, both in vitro and in vivo, are provided. One method involves incorporating an unnatural amino acid into a protein and attaching one or more saccharide moieties to the unnatural amino acid. Another method involves incorporating an unnatural amino acid that includes a saccharide moiety into a protein. Proteins made by both methods can be further modified with additional sugars.

Glycoprotein synthesis

DOEpatents

Methods for making glycoproteins, both in vitro and in vivo, are provided. One method involves incorporating an unnatural amino acid into a protein and attaching one or more saccharide moieties to the unnatural amino acid. Another method involves incorporating an unnatural amino acid that includes a saccharide moiety into a protein. Proteins made by both methods can be further modified with additional sugars.

2009-07-14

Methods for making glycoproteins, both in vitro and in vivo, are provided. One method involves incorporating an unnatural amino acid into a protein and attaching one or more saccharide moieties to the unnatural amino acid. Another method involves incorporating an unnatural amino acid that includes a saccharide moiety into a protein. Proteins made by both methods can be further modified with additional sugars.

Glycoprotein synthesis

DOEpatents

Schultz, Peter G.; Wang, Lei; Zhang, Zhiwen

2006-10-31

Methods for making glycoproteins, both in vitro and in vivo, are provided. One method involves incorporating an unnatural amino acid into a protein and attaching one or more saccharide moieties to the unnatural amino acid. Another method involves incorporating an unnatural amino acid that includes a saccharide moiety into a protein. Proteins made by both methods can be further modified with additional sugars.

Glycoprotein synthesis

DOEpatents

Schultz, Peter G [La Jolla, CA; Wang, Lei [San Diego, CA; Zhang, Zhiwen [San Diego, CA

2007-08-28

Methods for making glycoproteins, both in vitro and in vivo, are provided. One method involves incorporating an unnatural amino acid into a protein and attaching one or more saccharide moieties to the unnatural amino acid. Another method involves incorporating an unnatural amino acid that includes a saccharide moiety into a protein. Proteins made by both methods can be further modified with additional sugars.

Glycoprotein synthesis

DOEpatents

Schultz, Peter G [La Jolla, CA; Wang, Lei [San Diego, CA; Zhang, Zhiwen [San Diego, CA

2007-07-03

Methods for making glycoproteins, both in vitro and in vivo, are provided. One method involves incorporating an unnatural amino acid into a protein and attaching one or more saccharide moieties to the unnatural amino acid. Another method involves incorporating an unnatural amino acid that includes a saccharide moiety into a protein. Proteins made by both methods can be further modified with additional sugars.

Glycoprotein synthesis

DOEpatents

Schultz, Peter G.; Wang, Lei; Zhang, Zhiwen

2010-11-02

Methods for making glycoproteins, both in vitro and in vivo, are provided. One method involves incorporating an unnatural amino acid into a protein and attaching one or more saccharide moieties to the unnatural amino acid. Another method involves incorporating an unnatural amino acid that includes a saccharide moiety into a protein. Proteins made by both methods can be further modified with additional sugars.

Glycoprotein synthesis

DOEpatents

Schultz, Peter G [La Jolla, CA; Wang, Lei [San Diego, CA; Zhang, Zhiwen [San Diego, CA

2007-05-15

Methods for making glycoproteins, both in vitro and in vivo, are provided. One method involves incorporating an unnatural amino acid into a protein and attaching one or more saccharide moieties to the unnatural amino acid. Another method involves incorporating an unnatural amino acid that includes a saccharide moiety into a protein. Proteins made by both methods can be further modified with additional sugars.

Glycoprotein synthesis

DOEpatents

Schultz, Peter G.; Wang, Lei; Zhang, Zhiwen

2007-02-27

Methods for making glycoproteins, both in vitro and in vivo, are provided. One method involves incorporating an unnatural amino acid into a protein and attaching one or more saccharide moieties to the unnatural amino acid. Another method involves incorporating an unnatural amino acid that includes a saccharide moiety into a protein. Proteins made by both methods can be further modified with additional sugars.

Glycoprotein synthesis

DOEpatents

Shultz, Peter G [La Jolla, CA; Wang, Lei [San Diego, CA; Zhang, Zhiwen [San Diego, CA

2007-04-03

Methods for making glycoproteins, both in vitro and in vivo, are provided. One method involves incorporating an unnatural amino acid into a protein and attaching one or more saccharide moieties to the unnatural amino acid. Another method involves incorporating an unnatural amino acid that includes a saccharide moiety into a protein. Proteins made by both methods can be further modified with additional sugars.

A novel β-carotene-associated carrot (Daucus carota L.) pectic polysaccharide.

PubMed

Natarajmurthy, Sindhuja H; Askari, Marjan; Pullabhatla, Srinivas; Dharmesh, Shylaja M

2016-01-01

Carrot (Daucus carota L.) is a nutritional source with enriched β-carotene (0.01%) and pectins (0.8%). Although studies have highlighted the association between β-carotene and carrot pectic polysaccharide (CRPP), the precise binding of β-carotene to CRPP and its biological implications were not yet clearly understood. The aim of the present study is to report for the first time a natural carbohydrate polymer-CRPP-with associated β-carotene. The presence of β-carotene in CRPP and its binding is demonstrated by various studies including differential extraction followed by liquid chromatography (LC), liquid chromatography mass spectrometry (LC MS), and Fourier transform infrared spectroscopy (FTIR). Further, we show the binding via its release from CRPP using LC, fermentation studies and scanning electron microscopy (SEM) studies. The role of β-carotene in induction of apoptosis and the galactic-3 inhibitory property was also studied. The presence of β-carotene in CRPP was demonstrated, as well as an association between β-carotene and CRPP, apoptotic and galectin-3 inhibitory properties of CRPP, and upregulation of connexin 43 by CRPP. The data highlights that with the combination of β-carotene with CRPP, the prooxidant effect of β-carotene may be reduced, an apoptotic effect may be enhanced, and the anticancer potential of CRPP thus may be strengthened via galectin-3 inhibition. Copyright © 2016 Elsevier Inc. All rights reserved.

Immune Response Induced by an Immunodominant 60 kDa Glycoprotein of the Cell Wall of Sporothrix schenckii in Two Mice Strains with Experimental Sporotrichosis.

PubMed

Alba-Fierro, Carlos A; Pérez-Torres, Armando; Toriello, Conchita; Pulido-Camarillo, Evelyn; López-Romero, Everardo; Romo-Lozano, Yolanda; Gutiérrez-Sánchez, Gerardo; Ruiz-Baca, Estela

2016-01-01

Cell wall (CW) components of fungus Sporothrix schenckii are the major inductors antigens of immune responses. The immunodominant 60 kDa glycoprotein (gp60) has been shown to be associated with the virulence of this fungus but its role in experimental sporotrichosis is unknown. In this work, the immunological effects of CW-purified gp60 were investigated in a model of experimental subcutaneous sporotrichosis in normal and gp60-preimmunized C57BL/6 and BALB/c mice strains which were then infected with S. schenckii conidia. Results showed that both mice strains use different cytokine profiles in order to fight S. schenckii infection; C57BL/6 mice seem to use a Th17 response while BALB/c mice tend to depend on a Th1 profile. Preimmunization with gp60 showed a downregulatory effect on the immune response since cytokines levels were diminished in both strains. There were no significant differences in the magnitude of dorsoplantar inflammation between gp60-preimmunized and nonimmunized mice of both strains. However, skin lesions due to the infection in gp60-preimmunized mice were more severe in BALB/c than in C57BL/6 mice, suggesting that the antigen exerts a higher downregulatory effect on the Th1 response.

Immune Response Induced by an Immunodominant 60 kDa Glycoprotein of the Cell Wall of Sporothrix schenckii in Two Mice Strains with Experimental Sporotrichosis

PubMed Central

Alba-Fierro, Carlos A.; Pérez-Torres, Armando; Toriello, Conchita; Pulido-Camarillo, Evelyn; Romo-Lozano, Yolanda; Gutiérrez-Sánchez, Gerardo

2016-01-01

Cell wall (CW) components of fungus Sporothrix schenckii are the major inductors antigens of immune responses. The immunodominant 60 kDa glycoprotein (gp60) has been shown to be associated with the virulence of this fungus but its role in experimental sporotrichosis is unknown. In this work, the immunological effects of CW-purified gp60 were investigated in a model of experimental subcutaneous sporotrichosis in normal and gp60-preimmunized C57BL/6 and BALB/c mice strains which were then infected with S. schenckii conidia. Results showed that both mice strains use different cytokine profiles in order to fight S. schenckii infection; C57BL/6 mice seem to use a Th17 response while BALB/c mice tend to depend on a Th1 profile. Preimmunization with gp60 showed a downregulatory effect on the immune response since cytokines levels were diminished in both strains. There were no significant differences in the magnitude of dorsoplantar inflammation between gp60-preimmunized and nonimmunized mice of both strains. However, skin lesions due to the infection in gp60-preimmunized mice were more severe in BALB/c than in C57BL/6 mice, suggesting that the antigen exerts a higher downregulatory effect on the Th1 response. PMID:27051673

Protein and glycoprotein content of lymphocystis disease virus (LCDV).

PubMed

García-Rosado, Esther; Castro, Dolores; Cano, Irene; Alonso, M Carmen; Pérez-Prieto, Sara I; Borrego, Juan J

2004-06-01

The polypeptide and glycoprotein composition of eight strains of the fish-pathogenic lymphocystis disease virus (LCDV) isolated from gilt-head seabream (Sparus aurata), blackspot seabream (Pagellus bogaraveo), and sole (Solea senegalensis) were determined. The protein electrophoretic patterns of all LCDV isolates were quite similar regardless of the host fish, showing two major proteins (79.9 and 55.6 kDa) and a variable number of minor proteins. Three groups of LCDV isolates were distinguished according to the number and molecular masses of the minor proteins. Eight glycoproteins were detected inside viral particles of LCDV 2, LCDV 3 and LCDV 5 isolates, but only seven glycoproteins were found inside viral particles of LCDV 1, LCDV 4, LCDV 6, LCDV 7, and LCDV 11 isolates and the reference virus ATCC VR 342 by using five lectins. LCDV glycoproteins were mainly composed of mannose and sialic acid. These glycoproteins could be part of an external viral envelope probably derived from the host cell membrane.

Diversity Arrays Technology (DArT) platform for genotyping and mapping in carrot (Daucus carota L.)

USDA-ARS?s Scientific Manuscript database

Carrot is one of the most important root vegetable crops grown worldwide on more than one million hectares. Its progenitor, wild Daucus carota, is a weed commonly occurring across continents in the temperate climatic zone. Diversity Array Technology (DArT) is a microarray-based molecular marker syst...

Targeted Entry via Somatostatin Receptors Using a Novel Modified Retrovirus Glycoprotein That Delivers Genes at Levels Comparable to Those of Wild-Type Viral Glycoproteins

PubMed Central

Li, Fang; Ryu, Byoung Y.; Krueger, Robin L.; Heldt, Scott A.

2012-01-01

Here we report a novel viral glycoprotein created by replacing a natural receptor-binding sequence of the ecotropic Moloney murine leukemia virus envelope glycoprotein with the peptide ligand somatostatin. This new chimeric glycoprotein, which has been named the Sst receptor binding site (Sst-RBS), gives targeted transduction based on three criteria: (i) a gain of the use of a new entry receptor not used by any known virus; (ii) targeted entry at levels comparable to gene delivery by wild-type ecotropic Moloney murine leukemia virus and vesicular stomatitis virus (VSV) G glycoproteins; and (iii) a loss of the use of the natural ecotropic virus receptor. Retroviral vectors coated with Sst-RBS gained the ability to bind and transduce human 293 cells expressing somatostatin receptors. Their infection was specific to target somatostatin receptors, since a synthetic somatostatin peptide inhibited infection in a dose-dependent manner and the ability to transduce mouse cells bearing the natural ecotropic receptor was effectively lost. Importantly, vectors coated with the Sst-RBS glycoprotein gave targeted entry of up to 1 × 106 transducing U/ml, a level comparable to that seen with infection of vectors coated with the parental wild-type ecotropic Moloney murine leukemia virus glycoprotein through the ecotropic receptor and approaching that of infection of VSV G-coated vectors through the VSV receptor. To our knowledge, this is the first example of a glycoprotein that gives targeted entry of retroviral vectors at levels comparable to the natural capacity of viral envelope glycoproteins. PMID:22013043

Fate of Escherichia coli O157:H7 in manure compost-amended soil and on carrots and onions grown in an environmentally controlled growth chamber.

PubMed

Islam, Mahbub; Morgan, Jennie; Doyle, Michael P; Jiang, Xiuping

2004-03-01

Studies were done to determine the fate of Escherichia coli O157:H7 in manure compost-amended soil and on carrots and green onions grown in an environmentally controlled growth chamber. Commercial dairy cattle manure compost was inoculated with a five-strain mixture of green fluorescent protein-labeled E. coli O157:H7 at 10(7) CFU g(-1) and mixed with unsterilized Tifton sandy loam soil at a ratio of 1:5. Baby carrot or green onion seedlings were planted into the manure compost-amended soil in pots, and soil samples surrounding the plant, edible carrot roots and onion bulb samples, and soil immediately beneath the roots were assayed for E. coli O157:H7 in triplicate at weekly intervals for the first 4 weeks, and every 2 weeks for the remainder of the plant growth cycle (up to 3 months). E. coli O157:H7 cell numbers decreased within 64 days by 3 log CFU/g in soil and soil beneath the roots of green onions and by more than 2 log CFU/g on onions. E. coli O157:H7 survived better during the production of carrots, with a 2.3-log CFU/g reduction in soil and a 1.7-log CFU/g reduction on carrots within 84 days. These results indicate that the type of plant grown is an important factor influencing the survival of E. coli O157:H7 both on the vegetable and in the soil in which the vegetable is grown.

Medical care as the carrot: the Red Cross in Indonesia during the war of decolonization, 1945-1950.

PubMed

van Bergen, L

2013-01-01

During the war of decolonisation in Indonesia 1945-1950, the Dutch Red Cross and the Dutch East Indies Red Cross delivered aid to sick and wounded soldiers and civilians. This was supposed to happen in cooperation with organisations including the Indonesian Red Cross, the International Committee of the Red Cross, the military health service and civilian health services. Due to lack of resources, doctors and nurses, and due to differing interests, cooperation went anything but smoothly, severely undermining medical aid. On top of that, the aid that was given turned out be a tool of propaganda for the Dutch cause. Aid was deliberately--and with Red Cross consent--used as a political-military tool in the service of Dutch national interests. In a military strategy of carrot and stick, medical care served as the carrot.

Relationship between moisture content and electrical impedance of carrot slices during drying

NASA Astrophysics Data System (ADS)

Kertész, Ákos; Hlaváčová, Zuzana; Vozáry, Eszter; Staroňová, Lenka

2015-01-01

Electrical properties of food materials can give information about the inner structure and physiological state of biological tissues. Generally, the process of drying of fruits and vegetables is followed by weight loss. The aim of this study was to measure the impedance spectra of carrot slices during drying and to correlate impedance parameters to moisture content in different drying periods. Cylindrical slices were cut out from the carrot root along the axis. The slices were dried in a Venticell 111 air oven at 50°C. The weight of the slices was measured with a Denver SI-603 electronic analytical and precision balance. The weighing of the samples was performed every 30 min at the beginning of drying and every 60 min after the process. The moisture content of the samples was calculated on wet basis. The magnitude and phase angle of electrical impedance of the slices were measured with HP 4284A and 4285A precision LCR meters in the frequency range from 30 Hz to 1 MHz and from 75 kHz to 30 MHz, respectively, at voltage 1 V. The impedance measurement was performed after weighting. The change in the magnitude of impedance during drying showed a good correlation with the change in the moisture content.

The hydroxyapatite-binding regions of a rat salivary glycoprotein.

PubMed

Embery, G; Green, D R

1989-09-01

The regions of a salivary sulphated glycoprotein which are involved in its attachment to hydroxyapatite (Biogel HTP) have been characterised. The sulphated glycoprotein, a 35S-labelled preparation from mixed palatal and buccal minor gland secretions of the rat was bound onto hydroxyapatite and the resultant glycoprotein-hydroxyapatite complex was sequentially digested with pronase E and alpha-L-fucosidase, a treatment which released 86.8% +/- 1.7% of the radioactivity of the initially bound glycoprotein. The fragments which remained attached to the hydroxyapatite after enzymic digestion were fractionated on Sephadex G-25 and analysed for carbohydrate and amino acid components. A range of amino acids were detected which could reflect both glycosylated and non-glycosylated-binding regions. Sialic acid, although considered to be involved in the attachment process was not detected in any of the fragments remaining after enzymic digestion, a finding which provides indirect evidence that the enzymically liberated products do not subsequently re-attach to the hydroxyapatite surface. The notable feature of the fractions with average Mr estimated at 1000 or less is the high proportion of N-acetylhexosamine and N-acetylgalactosamine. It is apparent that the hexosamine residues, which normally bear the ester sulphate moieties of sulphated glycoproteins, play an important role in the attachment of sulphated glycoproteins to hydroxyapatite.

Metabolomics for organic food authentication: Results from a long-term field study in carrots.

PubMed

Cubero-Leon, Elena; De Rudder, Olivier; Maquet, Alain

2018-01-15

Increasing demand for organic products and their premium prices make them an attractive target for fraudulent malpractices. In this study, a large-scale comparative metabolomics approach was applied to investigate the effect of the agronomic production system on the metabolite composition of carrots and to build statistical models for prediction purposes. Orthogonal projections to latent structures-discriminant analysis (OPLS-DA) was applied successfully to predict the origin of the agricultural system of the harvested carrots on the basis of features determined by liquid chromatography-mass spectrometry. When the training set used to build the OPLS-DA models contained samples representative of each harvest year, the models were able to classify unknown samples correctly (100% correct classification). If a harvest year was left out of the training sets and used for predictions, the correct classification rates achieved ranged from 76% to 100%. The results therefore highlight the potential of metabolomic fingerprinting for organic food authentication purposes. Copyright © 2017 The Author(s). Published by Elsevier Ltd.. All rights reserved.

A workflow for large-scale empirical identification of cell wall N-linked glycoproteins of tomato (Solanum lycopersicum) fruit by tandem mass spectrometry

USDA-ARS?s Scientific Manuscript database

Glycosylation is a common post-translational modification of plant proteins that impacts a large number of important biological processes. Nevertheless, the impacts of differential site occupancy and the nature of specific glycoforms are obscure. Historically, characterization of glycoproteins has b...

Comparison of purple carrot juice and β-carotene in a high-carbohydrate, high-fat diet-fed rat model of the metabolic syndrome.

PubMed

Poudyal, Hemant; Panchal, Sunil; Brown, Lindsay

2010-11-01

Anthocyanins, phenolic acids and carotenoids are the predominant phytochemicals present in purple carrots. These phytochemicals could be useful in treatment of the metabolic syndrome since anthocyanins improve dyslipidaemia, glucose tolerance, hypertension and insulin resistance; the phenolic acids may also protect against CVD and β-carotene may protect against oxidative processes. In the present study, we have compared the ability of purple carrot juice and β-carotene to reverse the structural and functional changes in rats fed a high-carbohydrate, high-fat diet as a model of the metabolic syndrome induced by diet. Cardiac structure and function were defined by histology, echocardiography and in isolated hearts and blood vessels; liver structure and function, oxidative stress and inflammation were defined by histology and plasma markers. High-carbohydrate, high-fat diet-fed rats developed hypertension, cardiac fibrosis, increased cardiac stiffness, endothelial dysfunction, impaired glucose tolerance, increased abdominal fat deposition, altered plasma lipid profile, liver fibrosis and increased plasma liver enzymes together with increased plasma markers of oxidative stress and inflammation as well as increased inflammatory cell infiltration. Purple carrot juice attenuated or reversed all changes while β-carotene did not reduce oxidative stress, cardiac stiffness or hepatic fat deposition. As the juice itself contained low concentrations of carotenoids, it is likely that the anthocyanins are responsible for the antioxidant and anti-inflammatory properties of purple carrot juice to improve glucose tolerance as well as cardiovascular and hepatic structure and function.

Involvement of plasma membrane-located calmodulin in the response decay of cyclic nucleotide-gated cation channel of cultured carrot cells.

PubMed

Kurosaki, F; Kaburaki, H; Nishi, A

1994-03-07

Increase in cytoplasmic cyclic AMP concentration stimulates Ca2+ influx through the cyclic AMP-gated cation channel in the plasma membrane of cultured carrot cells. However, the Ca2+ current terminated after a few minutes even in the presence of high concentrations of cyclic AMP indicating that hydrolysis of the nucleotide is not responsible for stop of the Ca2+ influx. Cyclic AMP evoked discharge of Ca2+ from inside-out sealed vesicles of carrot plasma membrane, and it was strongly inhibited when the suspension of the vesicles was supplemented with 1 microM of free Ca2+, while Ca2+ lower than 0.1 microM did not affect the Ca(2+)-release. The Ca2+ flux across plasma membrane was restored from this Ca(2+)-induced inhibition by the addition of calmodulin inhibitors or anti-calmodulin. These results suggest that Ca2+ influx initiated by the increase in intracellular cAMP in cultured carrot cells is terminated when the cytosolic Ca2+ concentration reaches the excitatory level in the cells, and calmodulin located in the plasma membrane plays an important role in the response decay of the cyclic nucleotide-gated Ca2+ channel.

Arenavirus Stable Signal Peptide Is the Keystone Subunit for Glycoprotein Complex Organization

PubMed Central

Bederka, Lydia H.; Bonhomme, Cyrille J.; Ling, Emily L.

2014-01-01

ABSTRACT The rodent arenavirus glycoprotein complex encodes a stable signal peptide (SSP) that is an essential structural component of mature virions. The SSP, GP1, and GP2 subunits of the trimeric glycoprotein complex noncovalently interact to stud the surface of virions and initiate arenavirus infectivity. Nascent glycoprotein production undergoes two proteolytic cleavage events: first within the endoplasmic reticulum (ER) to cleave SSP from the remaining precursor GP1/2 (glycoprotein complex [GPC]) glycoprotein and second within the Golgi stacks by the cellular SKI-1/S1P for GP1/2 processing to yield GP1 and GP2 subunits. Cleaved SSP is not degraded but retained as an essential glycoprotein subunit. Here, we defined functions of the 58-amino-acid lymphocytic choriomeningitis virus (LCMV) SSP in regard to glycoprotein complex processing and maturation. Using molecular biology techniques, confocal microscopy, and flow cytometry, we detected SSP at the plasma membrane of transfected cells. Further, we identified a sorting signal (FLLL) near the carboxyl terminus of SSP that is required for glycoprotein maturation and trafficking. In the absence of SSP, the glycoprotein accumulated within the ER and was unable to undergo processing by SKI-1/S1P. Mutation of this highly conserved FLLL motif showed impaired glycoprotein processing and secretory pathway trafficking, as well as defective surface expression and pH-dependent membrane fusion. Immunoprecipitation of SSP confirmed an interaction between the signal peptide and the GP2 subunit; however, mutations within this FLLL motif disrupted the association of the GP1 subunit with the remaining glycoprotein complex. PMID:25352624

Influence of different processing and storage conditions on in vitro bioaccessibility of polyphenols in black carrot jams and marmalades.

PubMed

Kamiloglu, Senem; Pasli, Ayca Ayfer; Ozcelik, Beraat; Van Camp, John; Capanoglu, Esra

2015-11-01

Black carrot is indicated to play an important role in nutrition, as it comprises a variety of health-promoting components, including polyphenols. The objective of the present study was to monitor the stability of total phenolics, antioxidant capacity and phenolic acids in black carrot jams and marmalades after processing, storage and in vitro gastrointestinal digestion. Total phenolic content and antioxidant capacity were determined using spectrophotometric methods, whereas phenolic acids were identified using HPLC-PDA. Jam and marmalade processing significantly decreased total phenolics (89.2-90.5%), antioxidant capacity (83.3-91.3%) and phenolic acids (49.5-96.7%) (p < 0.05). After 20 weeks of storage, the percent decrease in total phenolics in samples stored at 25 °C (26.4-48.0%) was slightly higher than the samples stored at 4 °C (21.0-42.5%). In addition, jam and marmalade processing led to increases in the percent recovery of bioaccessible total phenolics (7.2-12.6%) and phenolic acids (4.7-31.5%), as well as antioxidant capacity (1.4-8.1%). In conclusion, current study highlighted black carrot jams and marmalades as good sources of polyphenols, with high bioaccessibility levels. Copyright © 2014 Elsevier Ltd. All rights reserved.

Immunomodulatory Effects of Nontoxic Glycoprotein Fraction Isolated from Rice Bran.

PubMed

Park, Ho-Young; Yu, A-Reum; Hong, Hee-Do; Kim, Ha Hyung; Lee, Kwang-Won; Choi, Hee-Don

2016-05-01

Rice bran, a by-product of brown rice milling, is a rich source of dietary fiber and protein, and its usage as a functional food is expected to increase. In this study, immunomodulatory effects of glycoprotein obtained from rice bran were studied in normal mice and mouse models of cyclophosphamide-induced immunosuppression. We prepared glycoprotein from rice bran by using ammonium precipitation and anion chromatography techniques. Different doses of glycoprotein from rice bran (10, 25, and 50 mg/kg) were administered orally for 28 days. On day 21, cyclophosphamide at a dose of 100 mg/kg was administered intraperitoneally. Glycoprotein from rice bran showed a significant dose-dependent restoration of the spleen index and white blood cell count in the immunocompromised mice. Glycoprotein from rice bran affected the immunomodulatory function by inducing the proliferation of splenic lymphocytes, which produce potential T and B cells. Moreover, it prevented cyclophosphamide-induced damage of Th1-type immunomodulatory function through enhanced secretion of Th1-type cytokines (interferon-γ and interleukin-12). These results indicate that glycoprotein from rice bran significantly recovered cyclophosphamide-induced immunosuppression. Based on these data, it was concluded that glycoprotein from rice bran is a potent immunomodulator and can be developed to recover the immunity of immunocompromised individuals. Georg Thieme Verlag KG Stuttgart · New York.

Detection of glycoproteins in the Acanthamoeba plasma membrane

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

Paatero, G.I.L.; Gahmberg, C.G.

1988-11-01

In the present study the authors have shown that glycoproteins are present in the plasma membrane of Acanthamoeba castellanii by utilizing different radioactive labeling techniques. Plasma membrane proteins in the amoeba were iodinated by {sup 125}I-lactoperoxidase labeling and the solubilized radiolabeled glycoproteins were separated by lectin-Sepharose affinity chromatography followed by polyacrylamide gel electrophoresis. The periodate/NaB{sup 3}H{sub 4} and galactose oxidase/NaB{sup 3}H{sub 4} labeling techniques were used for labeling of surface carbohydrates in the amoeba. Several surface-labeled glycoproteins were observed in addition to a diffusely labeled region with M{sub r} of 55,000-75,000 seen on electrophoresis, which could represent glycolipids. The presencemore » of glycoproteins in the plasma membrane of Acanthamoeba castellanii was confirmed by metabolic labeling with ({sup 35}S)methionine followed by lectin-Sepharose affinity chromatography and polyacrylamide gel electrophoresis.« less

Bio-Friendly Alternatives for Xylene – Carrot oil, Olive oil, Pine oil, Rose oil

PubMed Central

Nandan, Surapaneni Rateesh Kumar; Kulkarni, Pavan G.; Rao, Thokala Madhusudan; Palakurthy, Pavan

2015-01-01

Background Xylene is a flammable liquid with characteristic petroleum or aromatic odours, it is miscible with most of the organic solvents and paraffin wax. Xylene clears tissues rapidly and renders transparency, facilitating clearing endpoint determination, this made it to be used as a clearing agent in routine histopathological techniques. Even though it is a good clearing agent, it causes damage to the tissues by its hardening effect particularly those fixed in non-protein coagulant fixatives. Apart from these tissue effects, it has severe, long lasting ill effects on health of technicians and pathologists when exposed to longer duration. Hence in order to overcome these effects and replace xylene with a safe alternative agent, the present study was carried out to assess the clearing ability and bio-friendly nature of four different natural oils i.e., Carrot oil, Olive oil, Pine oil and Rose oil in comparison with that of Xylene. According to Bernoulli’s principle of fluid dynamics, to decrease viscosity of these oils and increase penetration into tissues for rapid clearing hot-air oven technique was used. Aims To assess:1) Clearing ability and bio-friendly nature of four different oils i.e., Carrot oil, Olive oil, Pine oil, Rose oil in comparison with that of xylene, 2) Application of Bernoulli’s principle of fluid dynamics in rapid clearing of tissues by using hot-air oven. Materials and Methods Forty different formalin fixed tissue samples were taken. Each sample of tissue was cut into 5 bits (40x5=200 total bits) which were subjected for dehydration in differential alcohol gradients. Later, each bit is kept in 4 different oils such as Carrot oil, Olive oil, Pine oil, Rose oil and xylene and transferred into hot-air oven. Further routine steps of processing, sectioning and staining were done. Individual sections cleared in four different oils were assessed for cellular architecture, staining quality and a comparison was done between them. Results Results

Three-Dimensionally Functionalized Reverse Phase Glycoprotein Array for Cancer Biomarker Discovery and Validation.

PubMed

Pan, Li; Aguilar, Hillary Andaluz; Wang, Linna; Iliuk, Anton; Tao, W Andy

2016-11-30

Glycoproteins have vast structural diversity that plays an important role in many biological processes and have great potential as disease biomarkers. Here, we report a novel functionalized reverse phase protein array (RPPA), termed polymer-based reverse phase glycoprotein array (polyGPA), to capture and profile glycoproteomes specifically, and validate glycoproteins. Nitrocellulose membrane functionalized with globular hydroxyaminodendrimers was used to covalently capture preoxidized glycans on glycoproteins from complex protein samples such as biofluids. The captured glycoproteins were subsequently detected using the same validated antibodies as in RPPA. We demonstrated the outstanding specificity, sensitivity, and quantitative capabilities of polyGPA by capturing and detecting purified as well as endogenous α-1-acid glycoprotein (AGP) in human plasma. We further applied quantitative N-glycoproteomics and the strategy to validate a panel of glycoproteins identified as potential biomarkers for bladder cancer by analyzing urine glycoproteins from bladder cancer patients or matched healthy individuals.

21 CFR 866.5440 - Beta-2-glycoprotein III immunological test system.

Code of Federal Regulations, 2010 CFR

2010-04-01

... SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Immunological Test Systems § 866.5440 Beta-2-glycoprotein III immunological test system. (a) Identification. A beta-2-glycoprotein III... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Beta-2-glycoprotein III immunological test system...

21 CFR 866.5430 - Beta-2-glycoprotein I immunological test system.

Code of Federal Regulations, 2010 CFR

2010-04-01

... SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Immunological Test Systems § 866.5430 Beta-2-glycoprotein I immunological test system. (a) Identification. A beta-2-glycoprotein I... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Beta-2-glycoprotein I immunological test system...

Glycoprotein Enrichment Analytical Techniques: Advantages and Disadvantages.

PubMed

Zhu, R; Zacharias, L; Wooding, K M; Peng, W; Mechref, Y

2017-01-01

Protein glycosylation is one of the most important posttranslational modifications. Numerous biological functions are related to protein glycosylation. However, analytical challenges remain in the glycoprotein analysis. To overcome the challenges associated with glycoprotein analysis, many analytical techniques were developed in recent years. Enrichment methods were used to improve the sensitivity of detection, while HPLC and mass spectrometry methods were developed to facilitate the separation of glycopeptides/proteins and enhance detection, respectively. Fragmentation techniques applied in modern mass spectrometers allow the structural interpretation of glycopeptides/proteins, while automated software tools started replacing manual processing to improve the reliability and throughput of the analysis. In this chapter, the current methodologies of glycoprotein analysis were discussed. Multiple analytical techniques are compared, and advantages and disadvantages of each technique are highlighted. © 2017 Elsevier Inc. All rights reserved.

Effects of direct and gradual salinity exposure on carrot (Daucus carota L.) seeds and recovery response

USDA-ARS?s Scientific Manuscript database

Salinity is a major cause of abiotic stress in arid and semi-arid climates that substantially reduces crop yield. This study evaluated the effects of salinity on germination and early seedling growth of two carrot cultivars in vitro under varying salinity levels. Salinity was induced by incorporatin...

Utilization of Landsat-8 data for the estimation of carrot and maize crop water footprint under the arid climate of Saudi Arabia.

PubMed

Madugundu, Rangaswamy; Al-Gaadi, Khalid A; Tola, ElKamil; Hassaballa, Abdalhaleem A; Kayad, Ahmed G

2018-01-01

The crop Water Footprint (WF) can provide a comprehensive knowledge of the use of water through the demarcation of the amount of the water consumed by different crops. The WF has three components: green (WFg), blue (WFb) and grey (WFgr) water footprints. The WFg refers to the rainwater stored in the root zone soil layer and is mainly utilized for agricultural, horticultural and forestry production. The WFb, however, is the consumptive use of water from surface or groundwater resources and mainly deals with irrigated agriculture, industry, domestic water use, etc. While the WFgr is the amount of fresh water required to assimilate pollutants resulting from the use of fertilizers/agrochemicals. This study was conducted on six agricultural fields in the Eastern region of Saudi Arabia, during the period from December 2015 to December 2016, to investigate the spatiotemporal variation of the WF of silage maize and carrot crops. The WF of each crop was estimated in two ways, namely agro-meteorological (WFAgro) and remote sensing (WFRS) methods. The blue, green and grey components of WFAgro were computed with the use of weather station/Eddy covariance measurements and field recorded crop yield datasets. The WFRS estimated by applying surface energy balance principles on Landsat-8 imageries. However, due to non-availability of Landsat-8 data on the event of rainy days, this study was limited to blue component (WFRS-b). The WFAgro of silage maize was found to range from 3545 m3 t-1 to 4960 m3 t-1; on an average, the WFAgro-g, WFAgro-b, and WFAgro-gr are composed of < 1%, 77%, and 22%, respectively. In the case of carrot, the WFAgro ranged between 297 m3 t-1 and 502 m3 t-1. The WFAgro-g of carrot crop was estimated at <1%, while WFAgro-b and WFAgro-gr was 67% and 32%, respectively. The WFAgro-b is occupied as a major portion in WF of silage maize (77%) and carrot (68%) crops. This is due to the high crop water demand combined with a very erratic rainfall, the irrigation is

Carrot Juice Administration Decreases Liver Stearoyl-CoA Desaturase 1 and Improves Docosahexaenoic Acid Levels, but Not Steatosis in High Fructose Diet-Fed Weanling Wistar Rats.

PubMed

Mahesh, Malleswarapu; Bharathi, Munugala; Reddy, Mooli Raja Gopal; Kumar, Manchiryala Sravan; Putcha, Uday Kumar; Vajreswari, Ayyalasomayajula; Jeyakumar, Shanmugam M

2016-09-01

Non-alcoholic fatty liver disease (NAFLD) is one of the most prevalent liver diseases associated with an altered lifestyle, besides genetic factors. The control and management of NAFLD mostly depend on lifestyle modifications, due to the lack of a specific therapeutic approach. In this context, we assessed the effect of carrot juice on the development of high fructose-induced hepatic steatosis. For this purpose, male weanling Wistar rats were divided into 4 groups, fed either a control (Con) or high fructose (HFr) diet of AIN93G composition, with or without carrot juice (CJ) for 8 weeks. At the end of the experimental period, plasma biochemical markers, such as triglycerides, alanine aminotransferase, and β-hydroxy butyrate levels were comparable among the 4 groups. Although, the liver injury marker, aspartate aminotransferase, levels in plasma showed a reduction, hepatic triglycerides levels were not significantly reduced by carrot juice ingestion in the HFr diet-fed rats (HFr-CJ). On the other hand, the key triglyceride synthesis pathway enzyme, hepatic stearoyl-CoA desaturase 1 (SCD1), expression at mRNA level was augmented by carrot juice ingestion, while their protein levels showed a significant reduction, which corroborated with decreased monounsaturated fatty acids (MUFA), particularly palmitoleic (C16:1) and oleic (C18:1) acids. Notably, it also improved the long chain n-3 polyunsaturated fatty acid, docosahexaenoic acid (DHA; C22:6) content of the liver in HFr-CJ. In conclusion, carrot juice ingestion decreased the SCD1-mediated production of MUFA and improved DHA levels in liver, under high fructose diet-fed conditions. However, these changes did not significantly lower the hepatic triglyceride levels.

Carrot Juice Administration Decreases Liver Stearoyl-CoA Desaturase 1 and Improves Docosahexaenoic Acid Levels, but Not Steatosis in High Fructose Diet-Fed Weanling Wistar Rats

PubMed Central

Mahesh, Malleswarapu; Bharathi, Munugala; Reddy, Mooli Raja Gopal; Kumar, Manchiryala Sravan; Putcha, Uday Kumar; Vajreswari, Ayyalasomayajula; Jeyakumar, Shanmugam M.

2016-01-01

Non-alcoholic fatty liver disease (NAFLD) is one of the most prevalent liver diseases associated with an altered lifestyle, besides genetic factors. The control and management of NAFLD mostly depend on lifestyle modifications, due to the lack of a specific therapeutic approach. In this context, we assessed the effect of carrot juice on the development of high fructose-induced hepatic steatosis. For this purpose, male weanling Wistar rats were divided into 4 groups, fed either a control (Con) or high fructose (HFr) diet of AIN93G composition, with or without carrot juice (CJ) for 8 weeks. At the end of the experimental period, plasma biochemical markers, such as triglycerides, alanine aminotransferase, and β-hydroxy butyrate levels were comparable among the 4 groups. Although, the liver injury marker, aspartate aminotransferase, levels in plasma showed a reduction, hepatic triglycerides levels were not significantly reduced by carrot juice ingestion in the HFr diet-fed rats (HFr-CJ). On the other hand, the key triglyceride synthesis pathway enzyme, hepatic stearoyl-CoA desaturase 1 (SCD1), expression at mRNA level was augmented by carrot juice ingestion, while their protein levels showed a significant reduction, which corroborated with decreased monounsaturated fatty acids (MUFA), particularly palmitoleic (C16:1) and oleic (C18:1) acids. Notably, it also improved the long chain n-3 polyunsaturated fatty acid, docosahexaenoic acid (DHA; C22:6) content of the liver in HFr-CJ. In conclusion, carrot juice ingestion decreased the SCD1-mediated production of MUFA and improved DHA levels in liver, under high fructose diet-fed conditions. However, these changes did not significantly lower the hepatic triglyceride levels. PMID:27752492

Antigenic relatedness between glycoproteins of human respiratory syncytial virus subgroups A and B: evaluation of the contributions of F and G glycoproteins to immunity.

PubMed Central

Johnson, P R; Olmsted, R A; Prince, G A; Murphy, B R; Alling, D W; Walsh, E E; Collins, P L

1987-01-01

The degree of antigenic relatedness between human respiratory syncytial virus (RSV) subgroups A and B was estimated from antibody responses induced in cotton rats by respiratory tract infection with RSV. Glycoprotein-specific enzyme-linked immunosorbent assays of antibody responses induced by RSV infection demonstrated that the F glycoproteins of subgroups A and B were antigenically closely related (relatedness, R approximately 50%), whereas the G glycoproteins were only distantly related (R approximately 5%). Intermediate levels of antigenic relatedness (R approximately 25%) were seen in neutralizing antibodies from cotton rats infected with RSV of the two subgroups. Immunity against the F glycoprotein of subgroup A, induced by vaccinia-A2-F, conferred a high level of protection which was of comparable magnitude against challenge by RSV of either subgroup. In comparison, immunity against the G glycoprotein of subgroup A, induced by vaccinia-A2-G, conferred less complete, but significant, protection. Importantly, in vaccinia-A2-G-immunized animals, suppression of homologous challenge virus replication was significantly greater (13-fold) than that observed for the heterologous virus. PMID:3305988

Glycoproteins Enrichment and LC-MS/MS Glycoproteomics in Central Nervous System Applications.

PubMed

Zhu, Rui; Song, Ehwang; Hussein, Ahmed; Kobeissy, Firas H; Mechref, Yehia

2017-01-01

Proteins and glycoproteins play important biological roles in central nervous systems (CNS). Qualitative and quantitative evaluation of proteins and glycoproteins expression in CNS is critical to reveal the inherent biomolecular mechanism of CNS diseases. This chapter describes proteomic and glycoproteomic approaches based on liquid chromatography/tandem mass spectrometry (LC-MS or LC-MS/MS) for the qualitative and quantitative assessment of proteins and glycoproteins expressed in CNS. Proteins and glycoproteins, extracted by a mass spectrometry friendly surfactant from CNS samples, were subjected to enzymatic (tryptic) digestion and three down-stream analyses: (1) a nano LC system coupled with a high-resolution MS instrument to achieve qualitative proteomic profile, (2) a nano LC system combined with a triple quadrupole MS to quantify identified proteins, and (3) glycoprotein enrichment prior to LC-MS/MS analysis. Enrichment techniques can be applied to improve coverage of low abundant glycopeptides/glycoproteins. An example described in this chapter is hydrophilic interaction liquid chromatographic (HILIC) enrichment to capture glycopeptides, allowing efficient removal of peptides. The combination of three LC-MS/MS-based approaches is capable of the investigation of large-scale proteins and glycoproteins from CNS with an in-depth coverage, thus offering a full view of proteins and glycoproteins changes in CNS.

Making Home Sweet and Sturdy: Toxoplasma gondii ppGalNAc-Ts Glycosylate in Hierarchical Order and Confer Cyst Wall Rigidity

PubMed Central

Tomita, Tadakimi; Sugi, Tatsuki; Yakubu, Rama; Tu, Vincent; Ma, Yanfen

2017-01-01

ABSTRACT The protozoan intracellular parasite Toxoplasma gondii forms latent cysts in the central nervous system (CNS) and persists for the lifetime of the host. This cyst is cloaked with a glycosylated structure called the cyst wall. Previously, we demonstrated that a mucin-like glycoprotein, CST1, localizes to the cyst wall and confers structural rigidity on brain cysts in a mucin-like domain-dependent manner. The mucin-like domain of CST1 is composed of 20 units of threonine-rich tandem repeats that are O-GalNAc glycosylated. A family of enzymes termed polypeptide N-acetylgalactosaminyltransferases (ppGalNAc-Ts) initiates O-GalNAc glycosylation. To identify which isoforms of ppGalNAc-Ts are responsible for the glycosylation of the CST1 mucin-like domain and to evaluate the function of each ppGalNAc-T in the overall glycosylation of the cyst wall, all five ppGalNAc-T isoforms were deleted individually from the T. gondii genome. The ppGalNAc-T2 and -T3 deletion mutants produced various glycosylation defects on the cyst wall, implying that many cyst wall glycoproteins are glycosylated by T2 and T3. Both T2 and T3 glycosylate the CST1 mucin-like domain, and this glycosylation is necessary for CST1 to confer structural rigidity on the cyst wall. We established that T2 is required for the initial glycosylation of the mucin-like domain and that T3 is responsible for the sequential glycosylation on neighboring acceptor sites, demonstrating hierarchical glycosylation by two distinct initiating and filling-in ppGalNAc-Ts in an intact organism. PMID:28074022

Selenium biofortification of broccoli and carrots grown in soil amended with Se-enriched hyperaccumulator Stanleya pinnata

USDA-ARS?s Scientific Manuscript database

Amending soils with Se-hyperaccumulator plant derived sources of selenium (Se) may be useful for increasing Se content in food crops in Se-deficient regions of the world. In this study, we evaluated total Se and the different chemical species of Se in broccoli and carrots grown in soils amended with...

Selectivity lists of pesticides to beneficial arthropods for IPM programs in carrot--first results.

PubMed

Hautier, L; Jansen, J-P; Mabon, N; Schiffers, B

2005-01-01

In order to improve IPM programs in carrot, 7 fungicides, 12 herbicides and 9 insecticides commonly used in Belgium were tested for their toxicity towards five beneficial arthropods representative of most important natural enemies encountered in carrot: parasitic wasps - Aphidius rhopalosiphi (De Stefani-Perez) (Hym., Aphidiidae), ladybirds - Adalia bipunctata (L.) (Col., Coccinellidae), hoverfly - Episyrphus balteatus (Dipt.. Syrphidae), rove beetle - Aleochara bilineata (Col., Staphylinidae) and carabid beetle - Bembidion lampros (Col., Carabidae). Initialy, all plant protection products were tested on inert substrate glass plates or sand according to the insect. Products with a corrected mortality (CM) or a parasitism reduction (PR) lower than 30% were kept for the constitution of positive list (green list). The other compounds were further tested on plant for A. rhopalosiphi, A. bipunctata, E. balteatus and soil for B. lampros and A. bilineata. With these extended laboratory tests results, products were listed in toxicity class: green category [CM or PR < or = 30%], yellow category [30% < CM or PR < or = 60%] and orange category [60% < CM or PR < or = 80%]. Products with toxicity higher than 80% on plants or that reduce parasitism more than 80% on soil were put in red category and are not recommended to Integrated Pest Management programs in carrot. Results showed that all fungicides tested were harmless to beneficials except Tebuconazole, which was slightly harmful for A. bipunctata. Herbicides were also harmless for soil beneficials, except Chlorpropham. This product was very toxic on sand towards A. bilineata and must be tested on soil. All soil insecticides tested were very toxic for ground beneficials and considered as non-selective. Their use in IPM is subject to questioning in view of negative impacts on beneficials. Among foliar insecticides, Dimethoate and Deltamethrin are not recommended for IPM because their high toxicity for all beneficials. The

Paramyxovirus Glycoproteins and the Membrane Fusion Process.

PubMed

Aguilar, Hector C; Henderson, Bryce A; Zamora, J Lizbeth; Johnston, Gunner P

2016-09-01

The family Paramyxoviridae includes many viruses that significantly affect human and animal health. An essential step in the paramyxovirus life cycle is viral entry into host cells, mediated by virus-cell membrane fusion. Upon viral entry, infection results in expression of the paramyxoviral glycoproteins on the infected cell surface. This can lead to cell-cell fusion (syncytia formation), often linked to pathogenesis. Thus membrane fusion is essential for both viral entry and cell-cell fusion and an attractive target for therapeutic development. While there are important differences between viral-cell and cell-cell membrane fusion, many aspects are conserved. The paramyxoviruses generally utilize two envelope glycoproteins to orchestrate membrane fusion. Here, we discuss the roles of these glycoproteins in distinct steps of the membrane fusion process. These findings can offer insights into evolutionary relationships among Paramyxoviridae genera and offer future targets for prophylactic and therapeutic development.

Paramyxovirus Glycoproteins and the Membrane Fusion Process

PubMed Central

Aguilar, Hector C.; Henderson, Bryce A.; Zamora, J. Lizbeth; Johnston, Gunner P.

2016-01-01

The family Paramyxoviridae includes many viruses that significantly affect human and animal health. An essential step in the paramyxovirus life cycle is viral entry into host cells, mediated by virus-cell membrane fusion. Upon viral entry, infection results in expression of the paramyxoviral glycoproteins on the infected cell surface. This can lead to cell-cell fusion (syncytia formation), often linked to pathogenesis. Thus membrane fusion is essential for both viral entry and cell-cell fusion and an attractive target for therapeutic development. While there are important differences between viral-cell and cell-cell membrane fusion, many aspects are conserved. The paramyxoviruses generally utilize two envelope glycoproteins to orchestrate membrane fusion. Here, we discuss the roles of these glycoproteins in distinct steps of the membrane fusion process. These findings can offer insights into evolutionary relationships among Paramyxoviridae genera and offer future targets for prophylactic and therapeutic development. PMID:28138419

Metabolism of pharmaceutical and personal care products by carrot cell cultures.

PubMed

Wu, Xiaoqin; Fu, Qiuguo; Gan, Jay

2016-04-01

With the increasing use of treated wastewater and biosolids in agriculture, residues of pharmaceutical and personal care products (PPCPs) in these reused resources may contaminate food produce via plant uptake, constituting a route for human exposure. Although various PPCPs have been reported to be taken up by plants in laboratories or under field conditions, at present little information is available on their metabolism in plants. In this study, we applied carrot cell cultures to investigate the plant metabolism of PPCPs. Five phase I metabolites of carbamazepine were identified and the potential metabolism pathways of carbamazepine were proposed. We also used the carrot cell cultures as a rapid screening tool to initially assess the metabolism potentials of 18 PPCPs. Eleven PPCPs, including acetaminophen, caffeine, meprobamate, primidone, atenolol, trimethoprim, DEET, carbamazepine, dilantin, diazepam, and triclocarban, were found to be recalcitrant to metabolism. The other 7 PPCPs, including triclosan, naproxen, diclofenac, ibuprofen, gemfibrozil, sulfamethoxazole, and atorvastatin, displayed rapid metabolism, with 0.4-47.3% remaining in the culture at the end of the experiment. Further investigation using glycosidase hydrolysis showed that 1.3-20.6% of initially spiked naproxen, diclofenac, ibuprofen, and gemfibrozil were transformed into glycoside conjugates. Results from this study showed that plant cell cultures may be a useful tool for initially exploring the potential metabolites of PPCPs in plants as well as for rapidly screening the metabolism potentials of a variety of PPCPs or other emerging contaminants, and therefore may be used for prioritizing compounds for further comprehensive evaluations. Copyright © 2015 Elsevier Ltd. All rights reserved.

Biosynthesis and maturation of cellular membrane glycoproteins.

PubMed

Hunt, L A

1979-01-01

The biosynthesis and the processing of asparagine-linked oligosaccharides of cellular membrane glycoproteins were examined in monolayer cultures of BHK21 cells and human diploid fibroblasts after pulse- and pulse-chase labeling with [2-3H]mannose. After pronase digestion, radiolabeled glycopeptides were characterized by high-resolution gel filtration, with or without additional digestion with various exoglycosidases and endoglycosidases. Pulse-labeled glycoproteins contained a relatively homogenous population of neutral oligosaccharides (major species: Man9GlcNAc2ASN). The vast majority of these asparagine-linked oligosaccharides was smaller than the major fraction of lipid-linked oligosaccharides from the cell and was apparently devoid of terminal glucose. After pulse-chase or long labeling periods, a significant fraction of the large oligomannosyl cores was processed by removal of mannose units and addition of branch sugars (NeuNAc-Gal-GlcNAc), resulting in complex acidic structures containing three and possibly five mannoses. In addition, some of the large oligomannosyl cores were processed by the removal of only several mannoses, resulting in a mixture of neutral structures with 5-9 mannoses. This oligomannosyl core heterogeneity in both neutral and acidic oligosaccharides linked to asparagine in cellular membrane glycoproteins was analogous to the heterogeneity reported for the oligosaccharides of avian RNA tumor virus glycoproteins (Hunt LA, Wright SE, Etchison JR, Summers DF: J Virol 29:336, 1979).

Influence of cooking conditions on organoleptic and health-related properties of artichokes, green beans, broccoli and carrots.

PubMed

Guillén, Sofía; Mir-Bel, Jorge; Oria, Rosa; Salvador, María L

2017-02-15

Colour, pigments, total phenolic content and antioxidant activity were investigated in artichokes, green beans, broccoli and carrots cooked under different conditions. Domestic induction hobs with temperature control were used to evaluate the effect of boiling, sous-vide cooking and water immersion cooking at temperatures below 100°C on the properties of each vegetable. Sous-vide cooking preserved chlorophyll, carotenoids, phenolic content and antioxidant activity to a greater extent than boiling for all of the vegetables tested and retained colour better, as determined by a(∗). A reduction of only 10-15°C in the cooking temperature was enough to improve the properties of the samples cooked by water immersion, except for green beans. Artichokes and carrots suffered pronounced losses of antioxidant activity during boiling (17.0 and 9.2% retention, respectively), but the stability of this parameter significantly increased with sous-vide cooking (84.9 and 55.3% retention, respectively). Copyright © 2016 Elsevier Ltd. All rights reserved.

Carbohydrate moieties of myelin-associated glycoprotein, major glycoprotein of the peripheral nervous system myelin and other myelin glycoproteins potentially involved in cell adhesion.

PubMed

Badache, A; Burger, D; Villarroya, H; Robert, Y; Kuchler, S; Steck, A J; Zanetta, J P

1992-01-01

The myelin-associated glycoprotein (MAG) and the major glycoprotein of the peripheral nervous system myelin (P0) are two members of the family of cell adhesion molecules (CAMs). A role in cell adhesion of the carbohydrate moiety of these molecules has been attributed to the presence of N-glycans bearing the HNK-1 carbohydrate epitope. On the other hand, it has been suggested that these glycoproteins could be ligands of an endogenous mannose-binding lectin present in myelin, the cerebellar soluble lectin (CSL). In order to further document the heterogeneity of the glycans of these two CAMs, we have used several probes: an anti-carbohydrate antibody of the HNK-1 type, called Elec-39, the plant lectin concanavalin A (ConA), and the endogenous lectin CSL involved in myelin compaction. This study shows that CSL binds to a small proportion of the polypeptide chains of MAG found in adult CNS of rats and man and the polypeptide chains of P0 molecules from adult human and rat sciatic nerve. For MAG from adult rat brain, the binding of CSL is restricted to glycans of polypeptide chains which could be separated from the others according to their solubility properties. These MAG molecular entities react also with the Elec-39 antibody and with ConA. These results confirm that P0 and MAG are heterogeneous in their carbohydrate moieties.(ABSTRACT TRUNCATED AT 250 WORDS)

Strategies to overcome or circumvent P-glycoprotein mediated multidrug resistance.

PubMed

Yuan, Hongyu; Li, Xun; Wu, Jifeng; Li, Jinpei; Qu, Xianjun; Xu, Wenfang; Tang, Wei

2008-01-01

Cancer patients who receive chemotherapy often experience intrinsic or acquired resistance to a broad spectrum of chemotherapeutic agents. The phenomenon, termed multidrug resistance (MDR), is often associated with the over-expression of P-glycoprotein, a transmembrane protein pump, which can enhance efflux of a various chemicals structurally unrelated at the expense of ATP depletion, resulting in decrease of the intracellular cytotoxic drug accumulation. The MDR has been a big threaten to the human health and the war fight for it continues. Although several other mechanisms for MDR are elucidated in recent years, considerable efforts attempting to inverse MDR are involved in exploring P-glycoprotein modulators and suppressing P-glycoprotein expression. In this review, we will report on the recent advances in various strategies for overcoming or circumventing MDR mediated by P-glycoprotein.

Square-wave voltammetry assays for glycoproteins on nanoporous gold

PubMed Central

Pandey, Binod; Bhattarai, Jay K.; Pornsuriyasak, Papapida; Fujikawa, Kohki; Catania, Rosa; Demchenko, Alexei V.; Stine, Keith J.

2014-01-01

Electrochemical enzyme-linked lectinsorbent assays (ELLA) were developed using nanoporous gold (NPG) as a solid support for protein immobilization and as an electrode for the electrochemical determination of the product of the reaction between alkaline phosphatase (ALP) and p-aminophenyl phosphate (p-APP), which is p-aminophenol (p-AP). Glycoproteins or concanavalin A (Con A) and ALP conjugates were covalently immobilized onto lipoic acid self-assembled monolayers on NPG. The binding of Con A – ALP (or soybean agglutinin – ALP) conjugate to glycoproteins covalently immobilized on NPG and subsequent incubation with p-APP substrate was found to result in square-wave voltammograms whose peak difference current varied with the identity of the glycoprotein. NPG presenting covalently bound glycoproteins was used as the basis for a competitive electrochemical assay for glycoproteins in solution (transferrin and IgG). A kinetic ELLA based on steric hindrance of the enzyme-substrate reaction and hence reduced enzymatic reaction rate after glycoprotein binding is demonstrated using immobilized Con A–ALP conjugates. Using the immobilized Con A-ALP conjugate, the binding affinity of immunoglobulin G (IgG) was found to be 105 nM, and that for transferrin was found to be 650 nM. Minimal interference was observed in the presence of 5 mg mL−1 BSA as a model serum protein in both the kinetic and competitive ELLA. Inhibition studies were performed with methyl D-mannoside for the binding of TSF and IgG to Con A-ALP; IC50 values were found to be 90 μM and 286 μM, respectively. Surface coverages of proteins were estimated using solution depletion and the BCA protein concentration assay. PMID:24611035

Role of Carbohydrate in Glycoprotein Traffic and Secretion

DTIC Science & Technology

1988-01-01

synthesized in normal amounts but accumu- lated intracellularly, with transport to the cell surface being greatly de - layed. Glycoprotein E2 isolated from...UNcLA ,F E 2 Role of Carbohydrate in Glycoprotein Traffic and Secretion JAMES B. PARENT I. Introduction I!. Evidence for Intracellular Transport Signals...Ill. Oligosaccharide Biosynthesis IV. Role of Carbohydrate in Protein Solubility. Structure, and Stability V. Evidence for Carbohydrate Transport

Influence of thermal processing on hydrolysis and stability of folate poly-gamma-glutamates in broccoli (Brassica oleracea var. italica), carrot (Daucus carota) and tomato (Lycopersicon esculentum).

PubMed

Munyaka, Ann Wambui; Verlinde, Philippe; Mukisa, Ivan Muzira; Oey, Indrawati; Van Loey, Ann; Hendrickx, Marc

2010-04-14

The folate poly-gamma-glutamate profile, their concentrations, and hydrolysis by endogenous gamma-glutamyl hydrolase (GGH) were evaluated in broccoli, carrot and tomato. Further studies on the effect of time and temperature on folate poly-gamma-glutamate hydrolysis and stability were carried out in broccoli since this vegetable showed the highest long-chain and total folate poly-gamma-glutamate concentration. The evolution of l-ascorbic acid, total phenols and Trolox equivalent antioxidant capacity (TEAC) values was evaluated in parallel. Upon thermal inactivation of GGH prior to crushing, it was observed that broccoli, carrot and tomato contained poly-gamma-glutamates with one to seven glutamate residues but differed in the predominant poly-gamma-glutamates. Crushing of raw broccoli, carrot and tomato resulted in significant poly-gamma-glutamate profile changes in broccoli and carrot (indicating GGH-catalyzed hydrolysis) but not in tomato. In this study, the actual crushing of raw broccoli matrix had a greater effect on folate poly-gamma-glutamate hydrolysis than incubation conditions (0-30 min at 25-55 degrees C). During treatments at 25-140 degrees C, folate retention was higher at 80 and 100 degrees C than at the other temperatures. A similar trend in thermal stability was observed for folates, vitamin C, total phenols and TEAC value, an indication that conditions that result in endogenous antioxidants degradation might also result in folate degradation.

Polyamine and ethylene biosynthesis in relation to somatic embryogenesis in carrot (Daucus carota L.) cell cultures

Treesearch

Subhash C. Minocha; Cheryl A. Robie; Akhtar J. Khan; Nancy S. Papa; Andrew I. Samuelsen; Rakesh Minocha

1990-01-01

Carrot cell cultures provide a model experimental system for the analysis of biochemical and molecular events associated with morphogenesis in plants (3, 4, 5, 14). Among the biochemical changes accompanying somatic embryogenesis in this tissue is an increased biosynthesis ofpolyamines (1, 2, 7, 10, 11, 13). A variety of inhibitors of polyamine biosynthetic enzymes...

Is the Cry1Ab protein from Bacillus thuringiensis (Bt) taken up by plants from soils previously planted with Bt corn and by carrot from hydroponic culture?

PubMed

Icoz, I; Andow, D; Zwahlen, C; Stotzky, G

2009-07-01

The uptake of the insecticidal Cry1Ab protein from Bacillus thuringiensis (Bt) by various crops from soils on which Bt corn had previously grown was determined. In 2005, the Cry1Ab protein was detected by Western blot in tissues (leaves plus stems) of basil, carrot, kale, lettuce, okra, parsnip, radish, snap bean, and soybean but not in tissues of beet and spinach and was estimated by enzyme-linked immunosorbent assay (ELISA) to be 0.05 +/- 0.003 ng g(-1) of fresh plant tissue in basil, 0.02 +/- 0.014 ng g(-1) in okra, and 0.34 +/- 0.176 ng g(-1) in snap bean. However, the protein was not detected by ELISA in carrot, kale, lettuce, parsnip, radish, and soybean or in the soils by Western blot. In 2006, the Cry1Ab protein was detected by Western blot in tissues of basil, carrot, kale, radish, snap bean, and soybean from soils on which Bt corn had been grown the previous year and was estimated by ELISA to be 0.02 +/- 0.014 ng g(-1) of fresh plant tissue in basil, 0.19 +/- 0.060 ng g(-1) in carrot, 0.05 +/- 0.018 ng g(-1) in kale, 0.04 +/- 0.022 ng g(-1) in radish, 0.53 +/- 0.170 ng g(-1) in snap bean, and 0.15 +/- 0.071 ng g(-1) in soybean. The Cry1Ab protein was also detected by Western blot in tissues of basil, carrot, kale, radish, and snap bean but not of soybean grown in soil on which Bt corn had not been grown since 2002; the concentration was estimated by ELISA to be 0.03 +/- 0.021 ng g(-1) in basil, 0.02 +/- 0.008 ng g(-1) in carrot, 0.04 +/- 0.017 ng g(-1) in kale, 0.02 +/- 0.012 ng g(-1) in radish, 0.05 +/- 0.004 ng g(-1) in snap bean, and 0.09 +/- 0.015 ng g(-1) in soybean. The protein was detected by Western blot in 2006 in most soils on which Bt corn had or had not been grown since 2002. The Cry1Ab protein was detected by Western blot in leaves plus stems and in roots of carrot after 56 days of growth in sterile hydroponic culture to which purified Cry1Ab protein had been added and was estimated by ELISA to be 0.08 +/- 0.021 and 0.60 +/- 0.148 ng g(-1) of

In vitro bioaccessibility of β-carotene, Ca, Mg and Zn in landrace carrots (Daucus carota, L.).

PubMed

Zaccari, Fernanda; Cabrera, María Cristina; Ramos, Ana; Saadoun, Ali

2015-01-01

Four landrace carrots ("Becaria", "CRS", "González" and "Rodríguez") and two marketable cultivars (Kuroda and Brasilia), raw and steamed, were characterised by the total content of β-carotene Ca, Mg and Zn, in vitro bioaccessibility and by colour and were evaluated to determine the effect of particle size in nutrient bioaccessibility. Steaming increased the content of β-carotene extracted from "CRS" and Brasilia (29% and 75%) and decreased the content of β-carotene extracted from "CRS" by 23% in "Rodríguez." In addition, steaming caused a loss of Ca (21%) but did not change the amount of Mg and Zn. The bioaccessibility of β-carotene in raw and pulped carrots was very low (<0.5%). Furthermore, steaming and a smaller particle size increased the bioaccessibility of β-carotene by 3-16 times. Additionally, cooking increased the in vitro bioaccessibility of Ca and Zn but had no effect on Mg. Moreover, homogenisation increased the bioaccessibility by 20% in Ca, 17% in Mg, and 10% in Zn compared to pulping. Copyright © 2014 Elsevier Ltd. All rights reserved.

Amorphous titania modified with boric acid for selective capture of glycoproteins.

PubMed

Jin, Shanxia; Liu, Liping; Zhou, Ping

2018-05-22

Amorphous titania was modified with boric acid, and the resulting material was characterized by scanning electron microscopy, Fourier transform infrared spectroscopy, X-ray powder diffraction and X-ray photoelectron spectrometry. The new material, in contrast to conventional boronate affinity materials containing boronic acid ligands, bears boric acid groups. It is shown to exhibit high specificity for glycoproteins, and this was applied to design a method for solid phase extraction of glycoproteins as shown for ribonuclease B, horse radish peroxidase and ovalbumin. Glycoproteins were captured under slightly alkaline environment and released in acidic solutions. The glycoproteins extracted were detected by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. The binding capacities for ribonuclease B, horse radish peroxidase and ovalbumin typically are 9.3, 26.0 and 53.0 mg ∙ g -1 , respectively. The method was successfully applied to the selective enrichment of ovalbumin from egg white. Graphical abstract Schematic presentation of the capture of glycoproteins by amorphous titania modified with boric acid.

Only the carrot, not the stick: incorporating trust into the enforcement of regulation.

PubMed

Mendoza, Juan P; Wielhouwer, Jacco L

2015-01-01

New enforcement strategies allow agents to gain the regulator's trust and consequently face a lower audit probability. Prior research suggests that, in order to prevent lower compliance, a reduction in the audit probability (the "carrot") must be compensated with the introduction of a higher penalty for non-compliance (the "stick"). However, such carrot-and-stick strategies reflect neither the concept of trust nor the strategies observed in practice. In response to this, we define trust-based regulation as a strategy that incorporates rules that allow trust to develop, and using a generic (non-cooperative) game of tax compliance, we examine whether trust-based regulation is feasible (i.e., whether, in equilibrium, a reduction in the audit probability, without ever increasing the penalty for non-compliance, does not lead to reduced compliance). The model shows that trust-based regulation is feasible when the agent sufficiently values the future. In line with the concept of trust, this strategy is feasible when the regulator is uncertain about the agent's intentions. Moreover, the model shows that (i) introducing higher penalties makes trust-based regulation less feasible, and (ii) combining trust and forgiveness can lead to a lower audit probability for both trusted and distrusted agents. Policy recommendations often point toward increasing deterrence. This model shows that the opposite can be optimal.

Effect of acidification on carrot (Daucus carota) juice cloud stability.

PubMed

Schultz, Alison K; Barrett, Diane M; Dungan, Stephanie R

2014-11-26

Effects of acidity on cloud stability in pasteurized carrot juice were examined over the pH range of 3.5-6.2. Cloud sedimentation, particle diameter, and ζ potential were measured at each pH condition to quantify juice cloud stability and clarification during 3 days of storage. Acidification below pH 4.9 resulted in a less negative ζ potential, an increased particle size, and an unstable cloud, leading to juice clarification. As the acidity increased, clarification occurred more rapidly and to a greater extent. Only a weak effect of ionic strength was observed when sodium salts were added to the juice, but the addition of calcium salts significantly reduced the cloud stability.

Assessing the likelihood of transmission of ‘Candidatus Liberibacter solanacearum’ to carrot by potato psyllid, Bactericera cockerelli (Hemiptera: Triozidae)

USDA-ARS?s Scientific Manuscript database

‘Candidatus Liberibacter solanacearum’ (Lso) is a phloem-limited bacterium that severely affects important Solanaceae and Apiaceae crops, including potato, tomato, pepper, tobacco, carrot and celery. This bacterium is transmitted to solanaceous species by potato psyllid, Bactericera cockerelli, and ...

Tuning orb spider glycoprotein glue performance to habitat humidity.

PubMed

Opell, Brent D; Jain, Dharamdeep; Dhinojwala, Ali; Blackledge, Todd A

2018-03-26

Orb-weaving spiders use adhesive threads to delay the escape of insects from their webs until the spiders can locate and subdue the insects. These viscous threads are spun as paired flagelliform axial fibers coated by a cylinder of solution derived from the aggregate glands. As low molecular mass compounds (LMMCs) in the aggregate solution attract atmospheric moisture, the enlarging cylinder becomes unstable and divides into droplets. Within each droplet an adhesive glycoprotein core condenses. The plasticity and axial line extensibility of the glycoproteins are maintained by hygroscopic LMMCs. These compounds cause droplet volume to track changes in humidity and glycoprotein viscosity to vary approximately 1000-fold over the course of a day. Natural selection has tuned the performance of glycoprotein cores to the humidity of a species' foraging environment by altering the composition of its LMMCs. Thus, species from low-humidity habits have more hygroscopic threads than those from humid forests. However, at their respective foraging humidities, these species' glycoproteins have remarkably similar viscosities, ensuring optimal droplet adhesion by balancing glycoprotein adhesion and cohesion. Optimal viscosity is also essential for integrating the adhesion force of multiple droplets. As force is transferred to a thread's support line, extending droplets draw it into a parabolic configuration, implementing a suspension bridge mechanism that sums the adhesive force generated over the thread span. Thus, viscous capture threads extend an orb spider's phenotype as a highly integrated complex of large proteins and small molecules that function as a self-assembling, highly tuned, environmentally responsive, adhesive biomaterial. Understanding the synergistic role of chemistry and design in spider adhesives, particularly the ability to stick in wet conditions, provides insight in designing synthetic adhesives for biomedical applications. © 2018. Published by The Company of

[Hydroxyproline: Rich glycoproteins of the plant and cell wall]. Annual technical progress report, 1993

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

Varner, J.E.

1993-06-01

Since xylem tissue includes the main cell types which are lignified, we are interested in gene expression of glycine-rich proteins and proline-rich proteins, and other proteins which are involved in secondary cell wall thickening during xylogenesis. Since the main feature of xylogenesis is the deposition of additional wall components, study of the mechanism of xylogenesis will greatly advance our knowledge of the synthesis and assembly of wall macromolecules. We are using the in vitro xylogenesis system from isolated Zinnia mesophyll cells to isolate genes which are specifically expressed during xylogenesis. We have used subtractive hybridization methods to isolate a numbermore » of cDNA clones for differentially regulated genes from the cells after hormonal induction. So far, we have partially characterized 18 different cDNA clones from 239 positive clones. These differentially regulated genes can be divided into three sets according to the characteristics of gene expression in the induction medium and the control medium. The first set is induced in both the induction medium and the control medium without hormones. The second set is induced mainly in the induction medium and in the control medium with the addition of NAA alone. Two of thesegenes are exclusively induced by auxin. The third set of genes is induced mainly in the induction medium. Since these genes are not induced by either auxin or cytokinin alone, they may be directly involved in the process of xylogenesis. Our experiments on the localization of H{sub 2}O{sub 2} production reinforce the earlier ideas of others that H{sub 2}O{sub 2} is involved in normal lignification.« less

Effect of human alpha 2HS glycoprotein on mouse macrophage function.

PubMed Central

Lewis, J G; André, C M

1980-01-01

alpha 2HS glycoprotein was isolated from normal adult serum. The ability of alpha 2HS glycoprotein to promote the endocytosis of radiolabelled DNA and radiolabelled latex particles by mouse macrophages was investigated. The results using both radiolabelled latex particles and radiolabelled DNA show that alpha 2HS glycoprotein enhances the ability of mouse macrophages to take up these radiolabelled substrates as compared to control cells. Images Figure 1 Figure 2 PMID:7439929

Adipokine zinc-α2-glycoprotein regulated by growth hormone and linked to insulin sensitivity.

PubMed

Balaz, Miroslav; Ukropcova, Barbara; Kurdiova, Timea; Gajdosechova, Lucia; Vlcek, Miroslav; Janakova, Zuzana; Fedeles, Jozef; Pura, Mikulas; Gasperikova, Daniela; Smith, Steven R; Tkacova, Ruzena; Klimes, Iwar; Payer, Juraj; Wolfrum, Christian; Ukropec, Jozef

2015-02-01

Hypertrophic obesity is associated with impaired insulin sensitivity and lipid-mobilizing activity of zinc-α2-glycoprotein. Adipose tissue (AT) of growth hormone (GH) -deficient patients is characterized by extreme adipocyte hypertrophy due to defects in AT lipid metabolism. It was hypothesized that zinc-α2-glycoprotein is regulated by GH and mediates some of its beneficial effects in AT. AT from patients with GH deficiency and individuals with obesity-related GH deficit was obtained before and after 5-year and 24-month GH supplementation therapy. GH action was tested in primary human adipocytes. Relationships of GH and zinc-α2-glycoprotein with adipocyte size and insulin sensitivity were evaluated in nondiabetic patients with noncancerous cachexia and hypertrophic obesity. AT in GH-deficient adults displayed a substantial reduction of zinc-α2-glycoprotein. GH therapy normalized AT zinc-α2-glycoprotein. Obesity-related relative GH deficit was associated with almost 80% reduction of zinc-α2-glycoprotein mRNA in AT. GH increased zinc-α2-glycoprotein mRNA in both AT of obese men and primary human adipocytes. Interdependence of GH and zinc-α2-glycoprotein in regulating AT morphology and metabolic phenotype was evident from their relationship with adipocyte size and AT-specific and whole-body insulin sensitivity. The results demonstrate that GH is involved in regulation of AT zinc-α2-glycoprotein; however, the molecular mechanism linking GH and zinc-α2-glycoprotein in AT is yet unknown. © 2014 The Obesity Society.

Glycoprotein expression by adenomatous polyps of the colon

NASA Astrophysics Data System (ADS)

Roney, Celeste A.; Xie, Jianwu; Xu, Biying; Jabour, Paul; Griffiths, Gary; Summers, Ronald M.

2008-03-01

Colon cancer is the second leading cause of cancer related deaths in the United States. Specificity in diagnostic imaging for detecting colorectal adenomas, which have a propensity towards malignancy, is desired. Adenomatous polyp specimens of the colon were obtained from the mouse model of colorectal cancer called adenomatous polyposis coli-multiple intestinal neoplasia (APC Min). Histological evaluation, by the legume protein Ulex europaeus agglutinin I (UEA-1), determined expression of the glycoprotein α-L-fucose. FITC-labelled UEA-1 confirmed overexpression of the glycoprotein by the polyps on fluorescence microscopy in 17/17 cases, of which 13/17 included paraffin-fixed mouse polyp specimens. In addition, FITC-UEA-1 ex vivo multispectral optical imaging of 4/17 colonic specimens displayed over-expression of the glycoprotein by the polyps, as compared to non-neoplastic mucosa. Here, we report the surface expression of α-L-fucosyl terminal residues by neoplastic mucosal cells of APC specimens of the mouse. Glycoprotein expression was validated by the carbohydrate binding protein UEA-1. Future applications of this method are the development of agents used to diagnose cancers by biomedical imaging modalities, including computed tomographic colonography (CTC). UEA-1 targeting to colonic adenomas may provide a new avenue for the diagnosis of colorectal carcinoma by CT imaging.

Microsatellite isolation and marker development in carrot - genomic distribution, linkage mapping, genetic diversity analysis and marker transferability across Apiaceae

USDA-ARS?s Scientific Manuscript database

The Apiaceae family includes several vegetable and spice crop species among which carrot is the most economically important member, with ~21 million tons produced yearly worldwide. Despite its importance, molecular resources in this species are relatively underdeveloped. The availability of informat...

The plant secretory pathway seen through the lens of the cell wall.

PubMed

van de Meene, A M L; Doblin, M S; Bacic, Antony

2017-01-01

Secretion in plant cells is often studied by looking at well-characterised, evolutionarily conserved membrane proteins associated with particular endomembrane compartments. Studies using live cell microscopy and fluorescent proteins have illuminated the highly dynamic nature of trafficking, and electron microscopy studies have resolved the ultrastructure of many compartments. Biochemical and molecular analyses have further informed about the function of particular proteins and endomembrane compartments. In plants, there are over 40 cell types, each with highly specialised functions, and hence potential variations in cell biological processes and cell wall structure. As the primary function of secretion in plant cells is for the biosynthesis of cell wall polysaccharides and apoplastic transport complexes, it follows that utilising our knowledge of cell wall glycosyltransferases (GTs) and their polysaccharide products will inform us about secretion. Indeed, this knowledge has led to novel insights into the secretory pathway, including previously unseen post-TGN secretory compartments. Conversely, our knowledge of trafficking routes of secretion will inform us about polarised and localised deposition of cell walls and their constituent polysaccharides/glycoproteins. In this review, we look at what is known about cell wall biosynthesis and the secretory pathway and how the different approaches can be used in a complementary manner to study secretion and provide novel insights into these processes.

Characterization of a salt-responsive 24-kilodalton glycoprotein in Mesembryanthemum crystallinum.

PubMed Central

Yen, H E; Edwards, G E; Grimes, H D

1994-01-01

A concanavalin A (Con A)-binding polypeptide with a molecular mass of 24 kD (termed "SRgp24") was associated with the intercellular space of Mesembryanthemum crystallinum L. callus. When callus was grown in medium containing between 0 and 100 mM NaCl, SRgp24 was detected by Con A binding. Increasing the NaCl concentration to 200 mM caused a reduction in the amount of SRgp24 within 3 d, and returning the callus to medium without salt resulted in an accumulation of SRgp24. Immunoblot analysis showed that appreciable amounts of SRgp24 accumulated in the leaves when plants were grown under sodium-limiting conditions. Unlike most of the cell-wall Con A-binding proteins in M. crystallinum callus, the carbohydrate moiety of SRgp24 was resistant to endoglycosidase H digestion. After purification of SRgp24, the N terminus was sequenced and found to share 55 to 60% identity with the N terminus of osmotin, a group 5 pathogenesis-related protein (PR-5) that accumulates in salt-adapted tobacco cell suspension. Immunocytochemical assays, with affinity-purified antibodies to SRgp24, indicated that SRgp24 preferentially accumulated in the cell-wall region. We conclude that SRgp24 is a salt-responsive glycoprotein related to the PR-5 family in M. crystallinum. PMID:7972493

Glycoprotein interactions in paramyxovirus fusion

PubMed Central

Iorio, Ronald M; Melanson, Vanessa R; Mahon, Paul J

2009-01-01

The Paramyxoviridae are enveloped, negative-stranded RNA viruses, some of which recognize sialic acid-containing receptors, while others recognize specific proteinaceous receptors. The major cytopathic effect of paramyxovirus infection is membrane fusion-induced syncytium formation. Paramyxoviruses are unusual in that the receptor-binding and fusion-promoting activities reside on two different spike structures, the attachment and fusion glycoproteins, respectively. For most paramyxoviruses, this distribution of functions requires a mechanism by which the two processes can be linked for the promotion of fusion. This is accomplished by a virus-specific interaction between the two proteins. An increasing body of evidence supports the notion that members of this family of viruses utilize this glycoprotein interaction in different ways in order to mediate the regulation of the fusion protein activation, depending on the type of receptor utilized by the virus. PMID:20161127

Glycoprotein Biochemistry--Some Clinical Aspects of Interest to Biochemistry Students.

ERIC Educational Resources Information Center

Smith, Christopher A.; And Others

1991-01-01

Authors describe some clinical features of glycoprotein biochemistry, including recognition, selected blood glycoproteins, glycated proteins, histochemistry, and cancer. The material presented has largely been taught to medical laboratory students; however, it can be used to teach premedical students and pure biochemistry students. Includes two…

Importance of the short cytoplasmic domain of the feline immunodeficiency virus transmembrane glycoprotein for fusion activity and envelope glycoprotein incorporation into virions

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

Celma, Cristina C.P.; Paladino, Monica G.; Gonzalez, Silvia A.

2007-09-30

The mature form of the envelope (Env) glycoprotein of lentiviruses is a heterodimer composed of the surface (SU) and transmembrane (TM) subunits. Feline immunodeficiency virus (FIV) possesses a TM glycoprotein with a cytoplasmic tail of approximately 53 amino acids which is unusually short compared with that of the other lentiviral glycoproteins (more than 100 residues). To investigate the relevance of the FIV TM cytoplasmic domain to Env-mediated viral functions, we characterized the biological properties of a series of Env glycoproteins progressively shortened from the carboxyl terminus. All the mutant Env proteins were efficiently expressed in feline cells and processed intomore » the SU and TM subunits. Deletion of 5 or 11 amino acids from the TM C-terminus did not significantly affect Env surface expression, fusogenic activity or Env incorporation into virions, whereas removal of 17 or 23 residues impaired Env-mediated cell-to-cell fusion. Further truncation of the FIV TM by 29 residues resulted in an Env glycoprotein that was poorly expressed at the cell surface, exhibited only 20% of the wild-type Env fusogenic capacity and was inefficiently incorporated into virions. Remarkably, deletion of the TM C-terminal 35 or 41 amino acids restored or even enhanced Env biological functions. Indeed, these mutant Env glycoproteins bearing cytoplasmic domains of 18 or 12 amino acids were found to be significantly more fusogenic than the wild-type Env and were efficiently incorporated into virions. Interestingly, truncation of the TM cytoplasmic domain to only 6 amino acids did not affect Env incorporation into virions but abrogated Env fusogenicity. Finally, removal of the entire TM cytoplasmic tail or deletion of as many as 6 amino acids into the membrane-spanning domain led to a complete loss of Env functions. Our results demonstrate that despite its relatively short length, the FIV TM cytoplasmic domain plays an important role in modulating Env-mediated viral

Bypassing P-Glycoprotein Drug Efflux Mechanisms: Possible Applications in Pharmacoresistant Schizophrenia Therapy

PubMed Central

Hoosain, Famida G.; Choonara, Yahya E.; Tomar, Lomas K.; Tyagi, Charu; du Toit, Lisa C.

2015-01-01

The efficient noninvasive treatment of neurodegenerative disorders is often constrained by reduced permeation of therapeutic agents into the central nervous system (CNS). A vast majority of bioactive agents do not readily permeate into the brain tissue due to the existence of the blood-brain barrier (BBB) and the associated P-glycoprotein efflux transporter. The overexpression of the MDR1 P-glycoprotein has been related to the occurrence of multidrug resistance in CNS diseases. Various research outputs have focused on overcoming the P-glycoprotein drug efflux transporter, which mainly involve its inhibition or bypassing mechanisms. Studies into neurodegenerative disorders have shown that the P-glycoprotein efflux transporter plays a vital role in the progression of schizophrenia, with a noted increase in P-glycoprotein function among schizophrenic patients, thereby reducing therapeutic outcomes. In this review, we address the hypothesis that methods employed in overcoming P-glycoprotein in cancer and other disease states at the level of the BBB and intestine may be applied to schizophrenia drug delivery system design to improve clinical efficiency of drug therapies. In addition, the current review explores polymers and drug delivery systems capable of P-gp inhibition and modulation. PMID:26491671

Immunogold detection of glycoprotein antigens in sea urchin embryos.

PubMed

Benson, N C; Benson, S C; Wilt, F

1989-01-01

Four developmental stages of sea urchin embryos were labeled with colloidal gold in an attempt to elucidate the intracellular trafficking patterns within the cells that produce the glycoprotein matrix of the embryonic spicule. The primary mesenchyme cells (PMCs) form a syncytium and secrete an organic matrix on which calcium carbonate is laid down to form an endoskeletal spicule. The organic matrix has been isolated and characterized as glycoprotein consisting of four major bands. Polyclonal antibodies to these glycoproteins were used to label embryos from the mesenchyme blastula, early gastrula, late gastrula, and plutei stages of development. The label is concentrated in the Golgi complex and associated vesicles, in secretory vesicles, and in the organic matrix. The density of the labeling increases as development proceeds.

Quantitative mass spectrometric analysis of glycoproteins combined with enrichment methods.

PubMed

Ahn, Yeong Hee; Kim, Jin Young; Yoo, Jong Shin

2015-01-01

Mass spectrometry (MS) has been a core technology for high sensitive and high-throughput analysis of the enriched glycoproteome in aspects of quantitative assays as well as qualitative profiling of glycoproteins. Because it has been widely recognized that aberrant glycosylation in a glycoprotein may involve in progression of a certain disease, the development of efficient analysis tool for the aberrant glycoproteins is very important for deep understanding about pathological function of the glycoprotein and new biomarker development. This review first describes the protein glycosylation-targeting enrichment technologies mainly employing solid-phase extraction methods such as hydrizide-capturing, lectin-specific capturing, and affinity separation techniques based on porous graphitized carbon, hydrophilic interaction chromatography, or immobilized boronic acid. Second, MS-based quantitative analysis strategies coupled with the protein glycosylation-targeting enrichment technologies, by using a label-free MS, stable isotope-labeling, or targeted multiple reaction monitoring (MRM) MS, are summarized with recent published studies. © 2014 The Authors. Mass Spectrometry Reviews Published by Wiley Periodicals, Inc.

Extracellular Vesicle-Associated Transitory Cell Wall Components and Their Impact on the Interaction of Fungi with Host Cells.

PubMed

Nimrichter, Leonardo; de Souza, Marcio M; Del Poeta, Maurizio; Nosanchuk, Joshua D; Joffe, Luna; Tavares, Patricia de M; Rodrigues, Marcio L

2016-01-01

Classic cell wall components of fungi comprise the polysaccharides glucans and chitin, in association with glycoproteins and pigments. During the last decade, however, system biology approaches clearly demonstrated that the composition of fungal cell walls include atypical molecules historically associated with intracellular or membrane locations. Elucidation of mechanisms by which many fungal molecules are exported to the extracellular space suggested that these atypical components are transitorily located to the cell wall. The presence of extracellular vesicles (EVs) at the fungal cell wall and in culture supernatants of distinct pathogenic species suggested a highly functional mechanism of molecular export in these organisms. Thus, the passage of EVs through fungal cell walls suggests remarkable molecular diversity and, consequently, a potentially variable influence on the host antifungal response. On the basis of information derived from the proteomic characterization of fungal EVs from the yeasts Cryptoccocus neoformans and Candida albicans and the dimorphic fungi Histoplasma capsulatum and Paracoccidioides brasiliensis, our manuscript is focused on the clear view that the fungal cell wall is much more complex than previously thought.

Fast determination of phenoxy acid herbicides in carrots and apples using liquid chromatography coupled triple quadrupole mass spectrometry.

PubMed

Santilio, Angela; Stefanelli, Patrizia; Dommarco, Roberto

2009-08-01

A fast, simple and inexpensive method has been developed for the analysis of phenoxy acid herbicides: 2,4-dichlorophenoxyacetic acid (2,4-D), 4-chloro-2-methylphenoxyacetic acid (MCPA), 2-(4-chloro-o-tolyloxy)propionic acid (MCPP), 2-(4-aryloxyphenoxy)propionic acid (Fluazifop) and 2-(4-aryloxyphenoxy)propionic acid (Haloxyfop) in carrots and apples by liquid chromatography coupled to triple quadrupole mass spectrometry (LC/MS/MS). The compounds were analyzed by QuEChERS (quick, easy, cheap, effective, rugged, safe) methodology without cleanup. The recoveries were performed at two spiked levels (0.05 and 0.5 mg/kg) for both matrices with six replicates for each level. The mean recoveries ranged from 70-92% for both apples and carrots. The precision of the method expressed as relative standard deviation (RSD%) was found to be in the range 3-15%. For all compounds, good linearity (r(2) > 0.99) was obtained over the range of concentration from 0.05 micro g/mL to 0.5 micro g/mL, corresponding to the pesticide concentrations of 0.05 mg/kg and 0.5 mg/kg, respectively. The determination limits (LOQs) ranged from 0.01 ng/mL to 1.3 ng/mL in solvent, whereas, the LOQs calculated in matrix ranged from 0.05 ng/g to 21.0 ng/g for apples and from 0.06 ng/g to 10.2 ng/g for carrots. The developed methodology combines the advantages of both QuEChERS and LC/MS/MS producing a very rapid, sensitive and cheap method useful for the routine analytical laboratories.

Putative storage root specific promoters from cassava and yam: cloning and evaluation in transgenic carrots as a model system.

PubMed

Arango, Jacobo; Salazar, Bertha; Welsch, Ralf; Sarmiento, Felipe; Beyer, Peter; Al-Babili, Salim

2010-06-01

A prerequisite for biotechnological improvements of storage roots is the availability of tissue-specific promoters enabling high expression of transgenes. In this work, we cloned two genomic fragments, pMe1 and pDJ3S, controlling the expression of a gene with unknown function from cassava (Manihot esculenta) and of the storage protein dioscorin 3 small subunit gene from yam (Dioscorea japonica), respectively. Using beta-glucuronidase as a reporter, the activities of pMe1 and pDJ3S were evaluated in independent transgenic carrot lines and compared to the constitutive CaMV35S and the previously described cassava p15 promoters. Activities of pMe1 and pDJ3S in storage roots were assessed using quantitative GUS assays that showed pDJ3S as the most active one. To determine organ specificities, uidA transcript levels in leaves, stems and roots were measured by real-time RT-PCR analyses showing highest storage root specificity for pDJ3S. Root cross sections revealed that pMe1 was highly active in secondary xylem. In contrast, pDJ3S was active in all root tissues except for the central xylem. The expression patterns caused by the cassava p15 promoter in carrot storage roots were consistent with its previously described activities for the original storage organ. Our data demonstrate that the pDJ3S and, to a lesser extent, the pMe1 regulatory sequences represent feasible candidates to drive high and preferential expression of genes in carrot storage roots.

Effects of Chrysosporum (Aphanizomenon) ovalisporum extracts containing cylindrospermopsin on growth, photosynthetic capacity, and mineral content of carrots (Daucus carota).

PubMed

Guzmán-Guillén, Remedios; Campos, Alexandre; Machado, Joana; Freitas, Marisa; Azevedo, Joana; Pinto, Edgar; Almeida, Agostinho; Cameán, Ana M; Vasconcelos, Vitor

2017-01-01

Natural toxins produced by freshwater cyanobacteria, such as cylindrospermopsin, have been regarded as an emergent environmental threat. Despite the risks for food safety, the impact of these water contaminants in agriculture is not yet fully understood. Carrots (Daucus carota) are root vegetables, extensively consumed worldwide with great importance for human nourishment and economy. It is, therefore, important to evaluate the possible effects of using water contaminated with cyanotoxins on carrot cultivation. The aim of this work was to investigate cylindrospermopsin effects on D. carota grown in soil and irrigated for 30 days, with a Chrysosporum ovalisporum extract containing environmentally relevant concentrations of cylindrospermopsin (10 and 50 μg/L). The parameters evaluated were plant growth, photosynthetic capacity, and nutritional value (mineral content) in roots of carrots, as these are the edible parts of this plant crop. The results show that, exposure to cylindrospermopsin did not have a clear negative effect on growth or photosynthesis of D. carota, even leading to an increase of both parameters. However, alterations in mineral contents were detected after exposure to crude extracts of C. ovalisporum containing cylindrospermopsin. A general decline was observed for most minerals (Ca, Mg, Na, Fe, Mn, Zn, Mo, and P), although an increase was shown in the case of K and Cu, pointing to a possible interference of the cyanobacterial extract in mineral uptake. This study is the first to evaluate the effects of C. ovalisporum extracts on a root vegetable, however, more research is necessary to understand the effects of this toxin in environmentally relevant scenarios.

Carrot Juice Fermentations as Man-Made Microbial Ecosystems Dominated by Lactic Acid Bacteria.

PubMed

Wuyts, Sander; Van Beeck, Wannes; Oerlemans, Eline F M; Wittouck, Stijn; Claes, Ingmar J J; De Boeck, Ilke; Weckx, Stefan; Lievens, Bart; De Vuyst, Luc; Lebeer, Sarah

2018-06-15

Spontaneous vegetable fermentations, with their rich flavors and postulated health benefits, are regaining popularity. However, their microbiology is still poorly understood, therefore raising concerns about food safety. In addition, such spontaneous fermentations form interesting cases of man-made microbial ecosystems. Here, samples from 38 carrot juice fermentations were collected through a citizen science initiative, in addition to three laboratory fermentations. Culturing showed that Enterobacteriaceae were outcompeted by lactic acid bacteria (LAB) between 3 and 13 days of fermentation. Metabolite-target analysis showed that lactic acid and mannitol were highly produced, as well as the biogenic amine cadaverine. High-throughput 16S rRNA gene sequencing revealed that mainly species of Leuconostoc and Lactobacillus (as identified by 8 and 20 amplicon sequence variants [ASVs], respectively) mediated the fermentations in subsequent order. The analyses at the DNA level still detected a high number of Enterobacteriaceae , but their relative abundance was low when RNA-based sequencing was performed to detect presumptive metabolically active bacterial cells. In addition, this method greatly reduced host read contamination. Phylogenetic placement indicated a high LAB diversity, with ASVs from nine different phylogenetic groups of the Lactobacillus genus complex. However, fermentation experiments with isolates showed that only strains belonging to the most prevalent phylogenetic groups preserved the fermentation dynamics. The carrot juice fermentation thus forms a robust man-made microbial ecosystem suitable for studies on LAB diversity and niche specificity. IMPORTANCE The usage of fermented food products by professional chefs is steadily growing worldwide. Meanwhile, this interest has also increased at the household level. However, many of these artisanal food products remain understudied. Here, an extensive microbial analysis was performed of spontaneous fermented

Establishment of a fluorescence-based method to evaluate endocytosis of desialylated glycoproteins in vitro.

PubMed

Luo, Cheng; Chen, Song; Xu, Na; Sai, Wen Bo; Zhao, Wei; Li, Ying Chun; Hu, Xiao Jing; Tian, Hong; Gao, Xiang Dong; Yao, Wen Bing

2017-04-01

Insufficient sialylation can result in rapid clearance of therapeutic glycoproteins by intracellular degradation, which is mainly mediated by asialoglycoprotein receptors (ASGPRs) on hepatic cells. In contrast, for glycoproteins, a long half-life is often related to high level of terminal sialic acid. These could be extremely important for insufficient sialylated biomedicines in clinic, and development of therapeutic glycoproteins in laboratory. However, how the desialylated glycoproteins are removed and how to evaluate the ASGPRs mediated endocytosis in vitro needs further investigate. Herein we described an integrative characterization of ASGPRs in vitro to elucidate its endocytosis properties. The endocytosis was determined by a fluorescence-based quantization method. The results showed that the ASGPRs could bind to poorly sialylated glycoproteins including asialofetuin and low sialylated recombinant Factor VIIa with a relatively higher ASGPRs binding affinity, and induce a more rapid endocytosis in vitro. Moreover, the mechanism under the internalization of ASGPRs was also investigated, which was found to depend on clathrin and caveolin. Utilizing the relative fluorescence quantification can be suitable for measurement of insufficient sialylated glycoprotein endocytosis and quality control of therapeutic glycoproteins, which could be useful for the understanding of the development of therapeutic glycoproteins. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

First report of ‘Candidatus Liberibacter solanacearum’ associated with the psyllid Bactericera trigonica Hodkinson on carrots in Northern Africa

USDA-ARS?s Scientific Manuscript database

Carrot plants (Daucus carota L.) exhibiting symptoms of yellowing, purpling, and curling of leaves, proliferation of shoots, formation of hairy secondary roots, and plant decline were observed in March 2014 and February 2015 in commercial fields in the Gharb region of Morocco. The symptoms resembled...

Development of a high-throughput SNP resource to advance genomic, genetic and breeding research in carrot (Daucus carota L.)

USDA-ARS?s Scientific Manuscript database

The rapid advancement in high-throughput SNP genotyping technologies along with next generation sequencing (NGS) platforms has decreased the cost, improved the quality of large-scale genome surveys, and allowed specialty crops with limited genomic resources such as carrot (Daucus carota) to access t...

Thyroid hormone upregulates zinc-α2-glycoprotein production in the liver but not in adipose tissue.

PubMed

Simó, Rafael; Hernández, Cristina; Sáez-López, Cristina; Soldevila, Berta; Puig-Domingo, Manel; Selva, David M

2014-01-01

Overproduction of zinc-α2-glycoprotein by adipose tissue is crucial in accounting for the lipolysis occurring in cancer cachexia of certain malignant tumors. The main aim of this study was to explore whether thyroid hormone could enhance zinc-α2-glycoprotein production in adipose tissue. In addition, the regulation of zinc-α2-glycoprotein by thyroid hormone in the liver was investigated. We performed in vitro (HepG2 cells and primary human adipocytes) and in vivo (C57BL6/mice) experiments addressed to examine the effect of thyroid hormone on zinc-α2-glycoprotein production (mRNA and protein levels) in liver and visceral adipose tissue. We also measured the zinc-α2-glycoprotein serum levels in a cohort of patients before and after controlling their hyperthyroidism. Our results showed that thyroid hormone up-regulates zinc-α2-glycoprotein production in HepG2 cells in a dose-dependent manner. In addition, the zinc-α2-glycoprotein proximal promoter contains functional thyroid hormone receptor binding sites that respond to thyroid hormone treatment in luciferase reporter gene assays in HepG2 cells. Furthermore, zinc-α2-glycoprotein induced lipolysis in HepG2 in a dose-dependent manner. Our in vivo experiments in mice confirmed the up-regulation of zinc-α2-glycoprotein induced by thyroid hormone in the liver, thus leading to a significant increase in zinc-α2-glycoprotein circulating levels. However, thyroid hormone did not regulate zinc-α2-glycoprotein production in either human or mouse adipocytes. Finally, in patients with hyperthyroidism a significant reduction of zinc-α2-glycoprotein serum levels was detected after treatment but was unrelated to body weight changes. We conclude that thyroid hormone up-regulates the production of zinc-α2-glycoprotein in the liver but not in the adipose tissue. The neutral effect of thyroid hormones on zinc-α2-glycoprotein expression in adipose tissue could be the reason why zinc-α2-glycoprotein is not related to weight

Thyroid Hormone Upregulates Zinc-α2-glycoprotein Production in the Liver but Not in Adipose Tissue

PubMed Central

Simó, Rafael; Hernández, Cristina; Sáez-López, Cristina; Soldevila, Berta; Puig-Domingo, Manel; Selva, David M.

2014-01-01

Overproduction of zinc-α2-glycoprotein by adipose tissue is crucial in accounting for the lipolysis occurring in cancer cachexia of certain malignant tumors. The main aim of this study was to explore whether thyroid hormone could enhance zinc-α2-glycoprotein production in adipose tissue. In addition, the regulation of zinc-α2-glycoprotein by thyroid hormone in the liver was investigated. We performed in vitro (HepG2 cells and primary human adipocytes) and in vivo (C57BL6/mice) experiments addressed to examine the effect of thyroid hormone on zinc-α2-glycoprotein production (mRNA and protein levels) in liver and visceral adipose tissue. We also measured the zinc-α2-glycoprotein serum levels in a cohort of patients before and after controlling their hyperthyroidism. Our results showed that thyroid hormone up-regulates zinc-α2-glycoprotein production in HepG2 cells in a dose-dependent manner. In addition, the zinc-α2-glycoprotein proximal promoter contains functional thyroid hormone receptor binding sites that respond to thyroid hormone treatment in luciferase reporter gene assays in HepG2 cells. Furthermore, zinc-α2-glycoprotein induced lipolysis in HepG2 in a dose-dependent manner. Our in vivo experiments in mice confirmed the up-regulation of zinc-α2-glycoprotein induced by thyroid hormone in the liver, thus leading to a significant increase in zinc-α2-glycoprotein circulating levels. However, thyroid hormone did not regulate zinc-α2-glycoprotein production in either human or mouse adipocytes. Finally, in patients with hyperthyroidism a significant reduction of zinc-α2-glycoprotein serum levels was detected after treatment but was unrelated to body weight changes. We conclude that thyroid hormone up-regulates the production of zinc-α2-glycoprotein in the liver but not in the adipose tissue. The neutral effect of thyroid hormones on zinc-α2-glycoprotein expression in adipose tissue could be the reason why zinc-α2-glycoprotein is not related to weight

Leishmania cell wall as a potent target for antiparasitic drugs. A focus on the glycoconjugates.

PubMed

Cabezas, Yari; Legentil, Laurent; Robert-Gangneux, Florence; Daligault, Franck; Belaz, Sorya; Nugier-Chauvin, Caroline; Tranchimand, Sylvain; Tellier, Charles; Gangneux, Jean-Pierre; Ferrières, Vincent

2015-08-21

Although leishmaniasis has been studied for over a century, the fight against cutaneous, mucocutaneous and visceral forms of the disease remains a hot topic. This review refers to the parasitic cell wall and more particularly to the constitutive glycoconjugates. The structures of the main glycolipids and glycoproteins, which are species-dependent, are described. The focus is on the disturbance of the lipid membrane by existing drugs and possible new ones, in order to develop future therapeutic agents.

Flavonoids released by carrot (Daucus carota) seedlings stimulate hyphal development of vesicular-arbuscular mycorrhizal fungi in the presence of optimal CO2 enrichment.

PubMed

Poulin, M J; Bel-Rhlid, R; Piché, Y; Chênevert, R

1993-10-01

Carbon dioxide has been previously identified as a critical volatile factor that stimulates hyphal growth ofGigaspora margarita, a vesiculararbuscular mycorrhizal fungus, and we determined the optimal concentration at 2.0%. The beneficial effect of CO2 on fungal development is also visible in the presence of stimulatory (quercetin, myricetin) or inhibitory (naringenin) flavonoids. Sterile root exudates from carrot seedlings stimulate the hyphal development ofG. margarita in the presence of optimal CO2 enrichment. Three flavonols (quercetin, kaempferol, rutin or quercetin 3-rutinoside) and two flavones (apigenin, luteolin) were identified in carrot root exudates by means of HPLC retention time. Flavonols like quercetin and kaempferol are known to have stimulatory effects on hyphal growth ofG. margarita.

Serodiagnosis of infectious mononucleosis with a bovine erythrocyte glycoprotein.

PubMed

Fletcher, M A; Klimas, N G; Latif, Z A; Caldwell, K E

1983-09-01

A glycoprotein from bovine erythrocyte membrane was evaluated in two immunoassays as a reagent for the serodiagnosis of infectious mononucleosis (IM). We previously reported that a partially purified preparation of this glycoprotein, when attached to latex beads, agglutinated in the presence of IM heterophile antibody. In the present study, we used a highly purified form of the glycoprotein both as an agglutinating reagent, covalently bound to latex, and in a solid-phase sandwich-type radioimmunoassay (RIA) for IM antibody detection in a larger population of patients. We tested serum samples from college students with symptoms suggestive of IM with the latex reagent (143 samples) and with the RIA (245 samples). Correlation of these two tests, both with each other and with the classical differentially absorbed, agglutination tests for Paul-Bunnell antibody in IM sera, using fresh sheep or horse cells, was excellent (greater than 97% agreement). The new tests also corresponded in most cases with a rapid, unabsorbed preserved horse erythrocyte slide test. However, in this study of 245 samples, both apparent false-positives (5 samples) and apparent false-negatives (3 samples) were observed with this slide test. In conclusion, we found that the bovine glycoprotein as a reagent can facilitate the diagnosis of IM, giving results comparable to those with erythrocyte agglutination tests on differentially absorbed sera. The advantages are ease and speed of performance (latex test), potential for automation (RIA test), stability and uniformity of the glycoprotein reagent (latex and RIA tests), and most importantly, the ability to use unabsorbed sera (latex and RIA tests).

Physico-chemical parameters, bioactive compounds and microbial quality of thermo-sonicated carrot juice during storage.

PubMed

Martínez-Flores, Héctor E; Garnica-Romo, Ma Guadalupe; Bermúdez-Aguirre, Daniela; Pokhrel, Prashant Raj; Barbosa-Cánovas, Gustavo V

2015-04-01

Thermosonication has been successfully tested in food for microbial inactivation; however, changes in bioactive compounds and shelf-life of treated products have not been thoroughly investigated. Carrot juice was thermo-sonicated (24 kHz, 120 μm amplitude) at 50 °C, 54 °C and 58 °C for 10 min (acoustic power 2204.40, 2155.72, 2181.68 mW/mL, respectively). Quality parameters and microbial growth were evaluated after processing and during storage at 4 °C. Control and sonicated treatments at 50 °C and 54 °C had 10, 12 and 14 d of shelf-life, respectively. Samples sonicated at 58 °C had the best quality; microbial growth remained low at around 3-log for mesophiles, 4.5-log for yeasts and molds and 2-log for enterobacteria after 20 d of storage. Furthermore, thermo-sonicated juice at 58 °C retained >98% of carotenoids and 100% of ascorbic acid. Phenolic compounds increased in all stored, treated juices. Thermo-sonication is therefore a promising technology for preserving the quality of carrot juice by minimising the physicochemical changes during storage, retarding microbial growth and retaining the bioactive compounds. Copyright © 2014 Elsevier Ltd. All rights reserved.

Tromantadine inhibits HSV-1 induced syncytia formation and viral glycoprotein processing

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

Ickes, D.E.

1989-01-01

Tromantadine inhibits a late event in Herpes Simplex Virus Type 1 (HSV-1) replication, visualized by the inhibition of both the size and number of syncytia. Tromantadine can be added at any time between 1 and 9 h post infection with complete inhibition of syncytia formation. Glycan synthesis of the viral glycoproteins, important for syncytia formation, is incomplete due to tromantadine treatment. Tromantadine does not inhibit the initiation of glycosylation, since viral glycoproteins, gX{sub t}, synthesized in the presence of tromantadine still incorporate {sup 3}H-glucosamine. Tromantadine does not inhibit the transport of t e viral glycoproteins to the cell surface, sincemore » glycoproteins B, C, and D are expressed, as demonstrated by immunofluorescence. Tromantadine inhibition of HSV-1 glycoprotein processing is demonstrated by an increase in mobility of the radioimmunoprecipitated gX{sub t}, on SDS-PAGE. The gX{sub t} of KOS, a non-syncytial strain of HSV-1, had a similar increase in mobility, suggesting that the block in glycoprotein processing is a general effect of tromantadine treatment. Fucose, which is incorporated into oligosaccharides in the medial Golgi, is incorporated into gX{sub t}, indicating that the tromantadine block in glycoprotein processing occurs after this step. Lectin binding studies and SDS-PAGE analysis of gC processed in the presence of tromantadine, gC{sub t}, indicates that it has terminal galactose residues in both N- and O-linked glycans (binds Peanut and Ricin Agglutinins, respectively). The inhibition of sialylation of N-linked glycans by tromantadine was indicated by the extent of the increase in SDS-PAGE mobility of the G protein from Vesicular Stomatitis Virus. O-glycanase digestion and SDS-PAGE analysis of gC{sub t} indicate that the O-linked disaccharide NAcGal-Galactose is present.« less

Prediction of conserved sites and domains in glycoproteins B, C and D of herpes viruses.

PubMed

Rasheed, Muhammad Asif; Ansari, Abdur Rahman; Ihsan, Awais; Navid, Muhammad Tariq; Ur-Rehman, Shahid; Raza, Sohail

2018-03-01

Glycoprotein B (gB), C (gC) and D (gD) of herpes simplex virus are implicated in virus adsorption and penetration. The gB, gC and gD are glycoproteins for different processes of virus binding and attachment to the host cells. Moreover, their expression is necessary and sufficient to induce cell fusion in the absence of other glycoproteins. Egress of herpes simplex virus (HSV) and other herpes viruses from cells involves extensive modification of cellular membranes and sequential envelopment, de-envelopment and re-envelopment steps. Viral glycoproteins are important in these processes, and frequently two or more glycoproteins can largely suffice in any step. Hence, we target the 3 important glycoproteins (B, C and D) of eight different herpes viruses of different species. These species include human (HSV1 and 2), bovine (BHV1), equine (EHV1 and 4), chicken (ILT1 and MDV2) and pig (PRV1). By applying different bioinformatics tools, we highlighted the conserved sites in these glycoproteins which might be most significant regarding attachment and infection of the viruses. Moreover the conserved domains in these glycoproteins are also highlighted. From this study, we will able to analyze the role of different viral glycoproteins of different species during herpes virus adsorption and penetration. Moreover, this study will help to construct the antivirals that target the glycoproteins of different herpes viruses. Copyright © 2018 Elsevier Ltd. All rights reserved.

High-throughput microarray mapping of cell wall polymers in roots and tubers during the viscosity-reducing process.

PubMed

Huang, Yuhong; Willats, William G; Lange, Lene; Jin, Yanling; Fang, Yang; Salmeán, Armando A; Pedersen, Henriette L; Busk, Peter Kamp; Zhao, Hai

2016-01-01

Viscosity reduction has a great impact on the efficiency of ethanol production when using roots and tubers as feedstock. Plant cell wall-degrading enzymes have been successfully applied to overcome the challenges posed by high viscosity. However, the changes in cell wall polymers during the viscosity-reducing process are poorly characterized. Comprehensive microarray polymer profiling, which is a high-throughput microarray, was used for the first time to map changes in the cell wall polymers of sweet potato (Ipomoea batatas), cassava (Manihot esculenta), and Canna edulis Ker. over the entire viscosity-reducing process. The results indicated that the composition of cell wall polymers among these three roots and tubers was markedly different. The gel-like matrix and glycoprotein network in the C. edulis Ker. cell wall caused difficulty in viscosity reduction. The obvious viscosity reduction of the sweet potato and the cassava was attributed to the degradation of homogalacturonan and the released 1,4-β-d-galactan and 1,5-α-l-arabinan. © 2015 International Union of Biochemistry and Molecular Biology, Inc.

CHAPTER 7: Glycoprotein Enrichment Analytical Techniques: Advantages and Disadvantages

PubMed Central

Zhu, Rui; Zacharias, Lauren; Wooding, Kerry M.; Peng, Wenjing; Mechref, Yehia

2017-01-01

Protein glycosylation is one of the most important posttranslational modifications. Numerous biological functions are related to protein glycosylation. However, analytical challenges remain in the glycoprotein analysis. To overcome the challenges associated with glycoprotein analysis, many analytical techniques were developed in recent years. Enrichment methods were used to improve the sensitivity of detection while HPLC and mass spectrometry methods were developed to facilitate the separation of glycopeptides/proteins and enhance detection, respectively. Fragmentation techniques applied in modern mass spectrometers allow the structural interpretation of glycopeptides/proteins while automated software tools started replacing manual processing to improve the reliability and throughout of the analysis. In this chapter, the current methodologies of glycoprotein analysis were discussed. Multiple analytical techniques are compared, and advantages and disadvantages of each technique are highlighted. PMID:28109440

21 CFR 866.5420 - Alpha-1-glycoproteins immunological test system.

Code of Federal Regulations, 2014 CFR

2014-04-01

... electrophoresis) in serum and other body fluids. Measurement of specific alpha-1-glycoproteins may aid in the... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Alpha-1-glycoproteins immunological test system. 866.5420 Section 866.5420 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN...

21 CFR 866.5420 - Alpha-1-glycoproteins immunological test system.

Code of Federal Regulations, 2013 CFR

2013-04-01

... electrophoresis) in serum and other body fluids. Measurement of specific alpha-1-glycoproteins may aid in the... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Alpha-1-glycoproteins immunological test system. 866.5420 Section 866.5420 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN...

21 CFR 866.5420 - Alpha-1-glycoproteins immunological test system.

Code of Federal Regulations, 2012 CFR

2012-04-01

... electrophoresis) in serum and other body fluids. Measurement of specific alpha-1-glycoproteins may aid in the... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Alpha-1-glycoproteins immunological test system. 866.5420 Section 866.5420 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN...

21 CFR 866.5420 - Alpha-1-glycoproteins immunological test system.

Code of Federal Regulations, 2011 CFR

2011-04-01

... electrophoresis) in serum and other body fluids. Measurement of specific alpha-1-glycoproteins may aid in the... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Alpha-1-glycoproteins immunological test system. 866.5420 Section 866.5420 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN...

Effects of polyamine biosynthesis inhibitors on S-adenosylmethionine synthetase and S-adenosylmethionine decarboxylase activities in carrot cell cultures

Treesearch

S.C. Minocha; R. Minocha; A. Komamine

1991-01-01

Changes in the activites of S-adcnosylmethionine (SAM) synthetase (methionine adenosyltransferase, EC 2.5.1.6.) and SAM decarboxylase (EC 4.1.1.50) were studied in carrot (Daucus carota) cell cultures in response to 2,4-dichlorophenoxyacetic acid (2,4-D) and several inhibitors of polyamine biosynthesis. Activity of SAM synthetase increased...

HIV-1 envelope glycoprotein

DOEpatents

Caulfield, Michael; Cupo, Albert; Dean, Hansi; Hoffenberg, Simon; King, C. Richter; Klasse, P. J.; Marozsan, Andre; Moore, John P.; Sanders, Rogier W.; Ward, Andrew; Wilson, Ian; Julien, Jean-Philippe

2017-08-22

The present application relates to novel HIV-1 envelope glycoproteins, which may be utilized as HIV-1 vaccine immunogens, and antigens for crystallization, electron microscopy and other biophysical, biochemical and immunological studies for the identification of broad neutralizing antibodies. The present invention encompasses the preparation and purification of immunogenic compositions, which are formulated into the vaccines of the present invention.

Molecularly imprinted nanomicrospheres as matrix solid-phase dispersant combined with gas chromatography for determination of four phosphorothioate pesticides in carrot and yacon.

PubMed

Zhou, Mengchun; Hu, Nana; Shu, Shaohua; Wang, Mo

2015-01-01

An efficient, rapid, and selective method for sample pretreatment, namely, molecularly imprinted matrix solid-phase dispersion (MI-MSPD) coupled with gas chromatography (GC), was developed for the rapid isolation of four phosphorothioate organophosphorus pesticides (tolclofos-methyl, phoxim, chlorpyrifos, and parathion-methyl) from carrot and yacon samples. New molecularly imprinted polymer nanomicrospheres were synthesized by using typical structural analogue tolclofos-methyl as a dummy template via surface grafting polymerization on nanosilica. Then, these four pesticides in carrot and yacon were extracted and adsorbed using the imprinted nanomicrospheres and further determined by gas chromatography. Under the optimized conditions, a good linearity of four pesticides was obtained in a range of 0.05-17.0 ng·g(-1) with R varying from 0.9971 to 0.9996, and the detection limit of the method was 0.012~0.026 ng·g(-1) in carrot and yacon samples. The recovery rates at two spiked levels were in the range of 85.4-105.6% with RSD ≤9.6%. The presented MI-MSPD method combined the advantages of MSPD for allowing the extraction, dispersion, and homogenization in two steps and the advantages of MIPs for high affinity and selectivity towards four phosphorothioate pesticides, which could be applied to the determination of pesticide residues in complicated vegetal samples.

Molecularly Imprinted Nanomicrospheres as Matrix Solid-Phase Dispersant Combined with Gas Chromatography for Determination of Four Phosphorothioate Pesticides in Carrot and Yacon

PubMed Central

Zhou, Mengchun; Hu, Nana; Shu, Shaohua; Wang, Mo

2015-01-01

An efficient, rapid, and selective method for sample pretreatment, namely, molecularly imprinted matrix solid-phase dispersion (MI-MSPD) coupled with gas chromatography (GC), was developed for the rapid isolation of four phosphorothioate organophosphorus pesticides (tolclofos-methyl, phoxim, chlorpyrifos, and parathion-methyl) from carrot and yacon samples. New molecularly imprinted polymer nanomicrospheres were synthesized by using typical structural analogue tolclofos-methyl as a dummy template via surface grafting polymerization on nanosilica. Then, these four pesticides in carrot and yacon were extracted and adsorbed using the imprinted nanomicrospheres and further determined by gas chromatography. Under the optimized conditions, a good linearity of four pesticides was obtained in a range of 0.05–17.0 ng·g−1 with R varying from 0.9971 to 0.9996, and the detection limit of the method was 0.012~0.026 ng·g−1 in carrot and yacon samples. The recovery rates at two spiked levels were in the range of 85.4–105.6% with RSD ≤9.6%. The presented MI-MSPD method combined the advantages of MSPD for allowing the extraction, dispersion, and homogenization in two steps and the advantages of MIPs for high affinity and selectivity towards four phosphorothioate pesticides, which could be applied to the determination of pesticide residues in complicated vegetal samples. PMID:25954569

QUANTITATIVE MASS SPECTROMETRIC ANALYSIS OF GLYCOPROTEINS COMBINED WITH ENRICHMENT METHODS

PubMed Central

Ahn, Yeong Hee; Kim, Jin Young; Yoo, Jong Shin

2015-01-01

Mass spectrometry (MS) has been a core technology for high sensitive and high-throughput analysis of the enriched glycoproteome in aspects of quantitative assays as well as qualitative profiling of glycoproteins. Because it has been widely recognized that aberrant glycosylation in a glycoprotein may involve in progression of a certain disease, the development of efficient analysis tool for the aberrant glycoproteins is very important for deep understanding about pathological function of the glycoprotein and new biomarker development. This review first describes the protein glycosylation-targeting enrichment technologies mainly employing solid-phase extraction methods such as hydrizide-capturing, lectin-specific capturing, and affinity separation techniques based on porous graphitized carbon, hydrophilic interaction chromatography, or immobilized boronic acid. Second, MS-based quantitative analysis strategies coupled with the protein glycosylation-targeting enrichment technologies, by using a label-free MS, stable isotope-labeling, or targeted multiple reaction monitoring (MRM) MS, are summarized with recent published studies. © 2014 The Authors. Mass Spectrometry Reviews Published by Wiley Periodicals, Inc. Rapid Commun. Mass Spec Rev 34:148–165, 2015. PMID:24889823

β-(1,3)-Glucan Unmasking in Some Candida albicans Mutants Correlates with Increases in Cell Wall Surface Roughness and Decreases in Cell Wall Elasticity

DOE PAGES

Hasim, Sahar; Allison, David P.; Retterer, Scott T.; ...

2016-11-14

Candida albicans is among the most common human fungal pathogens, causing a broad range of infections, including life-threatening systemic infections. The cell wall of C. albicans is the interface between the fungus and the innate immune system. The cell wall is composed of an outer layer enriched in mannosylated glycoproteins (mannan) and an inner layer enriched in β-(1,3)-glucan and chitin. Detection of C. albicans by Dectin-1, a C-type signaling lectin specific for β-(1,3)-glucan, is important for the innate immune system to recognize systemic fungal infections. Increased exposure of β-(1,3)-glucan to the immune system occurs when the mannan layer is alteredmore » or removed in a process called unmasking. Nanoscale changes to the cell wall during unmasking were explored in this paper in live cells with atomic force microscopy (AFM). Two mutants, the cho1Δ/Δ and kre5Δ/Δ mutants, were selected as representatives that exhibit modest and strong unmasking, respectively. Comparisons of the cho1Δ/Δ and kre5Δ/Δ mutants to the wild type reveal morphological changes in their cell walls that correlate with decreases in cell wall elasticity. In addition, AFM tips functionalized with Dectin-1 revealed that the forces of binding of Dectin-1 to all of the strains were similar, but the frequency of binding was highest for the kre5Δ/Δ mutant, decreased for the cho1Δ/Δ mutant, and rare for the wild type. These data show that nanoscale changes in surface topology are correlated with increased Dectin-1 adhesion and decreased cell wall elasticity. Finally, AFM, using tips functionalized with immunologically relevant molecules, can map epitopes of the cell wall and increase our understanding of pathogen recognition by the immune system.« less

β-(1,3)-Glucan Unmasking in Some Candida albicans Mutants Correlates with Increases in Cell Wall Surface Roughness and Decreases in Cell Wall Elasticity

PubMed Central

Hasim, Sahar; Allison, David P.; Retterer, Scott T.; Hopke, Alex; Wheeler, Robert T.; Doktycz, Mitchel J.

2016-01-01

ABSTRACT Candida albicans is among the most common human fungal pathogens, causing a broad range of infections, including life-threatening systemic infections. The cell wall of C. albicans is the interface between the fungus and the innate immune system. The cell wall is composed of an outer layer enriched in mannosylated glycoproteins (mannan) and an inner layer enriched in β-(1,3)-glucan and chitin. Detection of C. albicans by Dectin-1, a C-type signaling lectin specific for β-(1,3)-glucan, is important for the innate immune system to recognize systemic fungal infections. Increased exposure of β-(1,3)-glucan to the immune system occurs when the mannan layer is altered or removed in a process called unmasking. Nanoscale changes to the cell wall during unmasking were explored in live cells with atomic force microscopy (AFM). Two mutants, the cho1Δ/Δ and kre5Δ/Δ mutants, were selected as representatives that exhibit modest and strong unmasking, respectively. Comparisons of the cho1Δ/Δ and kre5Δ/Δ mutants to the wild type reveal morphological changes in their cell walls that correlate with decreases in cell wall elasticity. In addition, AFM tips functionalized with Dectin-1 revealed that the forces of binding of Dectin-1 to all of the strains were similar, but the frequency of binding was highest for the kre5Δ/Δ mutant, decreased for the cho1Δ/Δ mutant, and rare for the wild type. These data show that nanoscale changes in surface topology are correlated with increased Dectin-1 adhesion and decreased cell wall elasticity. AFM, using tips functionalized with immunologically relevant molecules, can map epitopes of the cell wall and increase our understanding of pathogen recognition by the immune system. PMID:27849179

β-(1,3)-Glucan Unmasking in Some Candida albicans Mutants Correlates with Increases in Cell Wall Surface Roughness and Decreases in Cell Wall Elasticity

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

Hasim, Sahar; Allison, David P.; Retterer, Scott T.

Candida albicans is among the most common human fungal pathogens, causing a broad range of infections, including life-threatening systemic infections. The cell wall of C. albicans is the interface between the fungus and the innate immune system. The cell wall is composed of an outer layer enriched in mannosylated glycoproteins (mannan) and an inner layer enriched in β-(1,3)-glucan and chitin. Detection of C. albicans by Dectin-1, a C-type signaling lectin specific for β-(1,3)-glucan, is important for the innate immune system to recognize systemic fungal infections. Increased exposure of β-(1,3)-glucan to the immune system occurs when the mannan layer is alteredmore » or removed in a process called unmasking. Nanoscale changes to the cell wall during unmasking were explored in this paper in live cells with atomic force microscopy (AFM). Two mutants, the cho1Δ/Δ and kre5Δ/Δ mutants, were selected as representatives that exhibit modest and strong unmasking, respectively. Comparisons of the cho1Δ/Δ and kre5Δ/Δ mutants to the wild type reveal morphological changes in their cell walls that correlate with decreases in cell wall elasticity. In addition, AFM tips functionalized with Dectin-1 revealed that the forces of binding of Dectin-1 to all of the strains were similar, but the frequency of binding was highest for the kre5Δ/Δ mutant, decreased for the cho1Δ/Δ mutant, and rare for the wild type. These data show that nanoscale changes in surface topology are correlated with increased Dectin-1 adhesion and decreased cell wall elasticity. Finally, AFM, using tips functionalized with immunologically relevant molecules, can map epitopes of the cell wall and increase our understanding of pathogen recognition by the immune system.« less

Genetics Home Reference: glycoprotein VI deficiency

MedlinePlus

... protein called glycoprotein VI (GPVI). This protein is embedded in the outer membrane of blood cell fragments ... erythematosus (SLE). Autoimmune disorders occur when the immune system malfunctions and attacks the body's own cells and ...

Enantioselective Reduction by Crude Plant Parts: Reduction of Benzofuran-2-yl Methyl Ketone with Carrot ("Daucus carota") Bits

ERIC Educational Resources Information Center

Ravia, Silvana; Gamenara, Daniela; Schapiro, Valeria; Bellomo, Ana; Adum, Jorge; Seoane, Gustavo; Gonzalez, David

2006-01-01

The use of biocatalysis and biotransformations are important tools in green chemistry. The enantioselective reduction of a ketone by crude plant parts, using carrot ("Daucus carota") as the reducing agent is presented. The experiment introduces an example of a green chemistry procedure that can be tailored to fit in a regular laboratory session.…

The glycoproteins of Marburg and Ebola virus and their potential roles in pathogenesis.

PubMed

Feldmann, H; Volchkov, V E; Volchkova, V A; Klenk, H D

1999-01-01

Filoviruses cause systemic infections that can lead to severe hemorrhagic fever in human and non-human primates. The primary target of the virus appears to be the mononuclear phagocytic system. As the virus spreads through the organism, the spectrum of target cells increases to include endothelial cells, fibroblasts, hepatocytes, and many other cells. There is evidence that the filovirus glycoprotein plays an important role in cell tropism, spread of infection, and pathogenicity. Biosynthesis of the glycoprotein forming the spikes on the virion surface involves cleavage by the host cell protease furin into two disulfide linked subunits GP1 and GP2. GP1 is also shed in soluble form from infected cells. Different strains of Ebola virus show variations in the cleavability of the glycoprotein, that may account for differences in pathogenicity, as has been observed with influenza viruses and paramyxoviruses. Expression of the spike glycoprotein of Ebola virus, but not of Marburg virus, requires transcriptional editing. Unedited GP mRNA yields the nonstructural glycoprotein sGP, which is secreted extensively from infected cells. Whether the soluble glycoproteins GP1 and sGP interfere with the humoral immune response and other defense mechanisms remains to be determined.

Metabolic fate and distribution of [{sup 14}C]1,3-dichloropropene in carrot, lettuce, radish, tomato, and wheat

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

Barnkow, D.E.; Byrne, S.L.; Huskin, M.A.

1996-10-01

This study examined the nature of the radioactive residue in carrot, lettuce, radish, tomato, and wheat grown in soil treated with uniformly {sup 14}C-labeled cis- and trans-1,3-dichloropropene (1,3-D). Each crop was grown in soil treated at or above the maximum use rate. Carrot root and top, lettuce, radish root and top, tomato fruit and vine, and wheat forage, straw and grain were harvested, processed, and analyzed. The {sup 14}C residues were subjected to sequential extraction with diethyl ether and aqueous acetonitrile. The residues that remained were subjected to extraction, fractionation, and hydrolysis with buffer, enzymes, acid, alkali, and strong oxidizingmore » agents. Analyses of the solubilized residues demonstrated that no detectable parent, 1,3-D, or the putative metabolites 3-chloroallyl alcohol and 3-chloroacrylic acid were present. The components of the extractable residue included most major plant constituents (i.e., protein, pigments, organic acids, sucrose, cellulose, and lignin). Thus, natural incorporation of the {sup 14}C-label into natural plant biochemicals is demonstrated.« less

Dynamics of glycoprotein charge in the evolutionary history of human influenza.

PubMed

Arinaminpathy, Nimalan; Grenfell, Bryan

2010-12-30

Influenza viruses show a significant capacity to evade host immunity; this is manifest both as large occasional jumps in the antigenic phenotype of viral surface molecules and in gradual antigenic changes leading to annual influenza epidemics in humans. Recent mouse studies show that avidity for host cells can play an important role in polyclonal antibody escape, and further that electrostatic charge of the hemagglutinin glycoprotein can contribute to such avidity. We test the role of glycoprotein charge on sequence data from the three major subtypes of influenza A in humans, using a simple method of calculating net glycoprotein charge. Of all subtypes, H3N2 in humans shows a striking pattern of increasing positive charge since its introduction in 1968. Notably, this trend applies to both hemagglutinin and neuraminidase glycoproteins. In the late 1980s hemagglutinin charge reached a plateau, while neuraminidase charge started to decline. We identify key groups of amino acid sites involved in this charge trend. To our knowledge these are the first indications that, for human H3N2, net glycoprotein charge covaries strongly with antigenic drift on a global scale. Further work is needed to elucidate how such charge interacts with other immune escape mechanisms, such as glycosylation, and we discuss important questions arising for future study.

Genetic and biochemical characterization of the GH72 family of cell wall transglycosylases in Neurospora crassa.

PubMed

Ao, Jie; Free, Stephen J

2017-04-01

The Neurospora crassa genome encodes five GH72 family transglycosylases, and four of these enzymes (GEL-1, GEL-2, GEL-3 and GEL-5) have been found to be present in the cell wall proteome. We carried out an extensive genetic analysis on the role of these four transglycosylases in cell wall biogenesis and demonstrated that the transglycosylases are required for the formation of a normal cell wall. As suggested by the proteomic analysis, we found that multiple transglycosylases were being expressed in N. crassa cells and that different combinations of the enzymes are required in different cell types. The combination of GEL-1, GEL-2 and GEL-5 is required for the growth of vegetative hyphae, while the GEL-1, GEL-2, GEL-3 combination is needed for the production of aerial hyphae and conidia. Our data demonstrates that the enzymes are redundant with partially overlapping enzymatic activities, which provides the fungus with a robust cell wall biosynthetic system. Characterization of the transglycosylase-deficient mutants demonstrated that the incorporation of cell wall proteins was severely compromised. Interestingly, we found that the transglycosylase-deficient mutant cell walls contained more β-1,3-glucan than the wild type cell wall. Our results demonstrate that the GH72 transglycosylases are not needed for the incorporation of β-1,3-glucan into the cell wall, but they are required for the incorporation of cell wall glycoprotein into the cell wall. Copyright © 2017 Elsevier Inc. All rights reserved.

Identification and characterization of terpene synthases potentially involved in the formation of volatile terpenes in carrot (Daucus carota L.) roots

USDA-ARS?s Scientific Manuscript database

Plants produce numerous volatile organic compounds, which are important in determining the quality and nutraceutical properties of fruit and root crops, including the taste and the aroma of carrots (Daucus carota L.). A combined chemical, biochemical and molecular study was conducted to evaluate the...

The effect of variety and maturity on the quality of freeze-dried carrots. The effect of microwave blanching on the nutritional and textural quality of freeze-dried spinach

NASA Technical Reports Server (NTRS)

1979-01-01

Using carrots, the quality of freeze-dried products was studied to determine the optimum varieties and maturation stages for quality attributes such as appearance, flavor, texture, and nutritive value. The quality of freeze-dried carrots is discussed in terms of Gardner color, alcohol insoluble solids, viscosity, and core/cortex ratio. Also, microwave blanching of freeze-dried spinach was studied to determine vitamin interrelationships, anatomical changes, and oxidative deteriorations in terms of preprocessing microwave treatments. Statistical methods were employed in the gathering of data and interpretation of results in both studies.

Heterologous Expression of the Carrot Hsp17.7 gene Increased Growth, Cell Viability, and Protein Solubility in Transformed Yeast (Saccharomyces cerevisiae) under Heat, Cold, Acid, and Osmotic Stress Conditions.

PubMed

Ko, Eunhye; Kim, Minhye; Park, Yunho; Ahn, Yeh-Jin

2017-08-01

In industrial fermentation of yeast (Saccharomyces cerevisiae), culture conditions are often modified from the optimal growth conditions of the cells to maintain large-scale cultures and/or to increase recombinant protein production. However, altered growth conditions can be stressful to yeast cells resulting in reduced cell growth and viability. In this study, a small heat shock protein gene from carrot (Daucus carota L.), Hsp17.7, was inserted into the yeast genome via homologous recombination to increase tolerance to stress conditions that can occur during industrial culture. A DNA construct, Translational elongation factor gene promoter-carrot Hsp17.7 gene-Phosphoribosyl-anthranilate isomerase gene (an auxotrophic marker), was generated by a series of PCRs and introduced into the chromosome IV of the yeast genome. Immunoblot analysis showed that carrot Hsp17.7 accumulated in the transformed yeast cell lines. Growth rates and cell viability of these cell lines were higher than control cell lines under heat, cold, acid, and hyperosmotic stress conditions. Soluble protein levels were higher in the transgenic cell lines than control cell lines under heat and cold conditions, suggesting the molecular chaperone function of the recombinant Hsp17.7. This study showed that a recombinant DNA construct containing a HSP gene from carrot was successfully expressed in yeast by homologous recombination and increased tolerances to abiotic stress conditions.

Understanding the interactions of human follicle stimulating hormone with single-walled carbon nanotubes by molecular dynamics simulation and free energy analysis.

PubMed

Mahmoodi, Yasaman; Mehrnejad, Faramarz; Khalifeh, Khosrow

2018-01-01

Interactions of carbon nanotubes (CNTs) and blood proteins are of interest for nanotoxicology and nanomedicine. It is believed that the interactions of blood proteins and glycoproteins with CNTs may have important biological effects. In spite of many experimental studies of single-walled carbon nanotubes (SWCNT) and glycoproteins with different methods, little is known about the atomistic details of their association process or of structural alterations occurring in adsorbed glycoproteins. In this study, we have applied molecular dynamics simulation to investigate the interaction of follicle stimulating hormone (hFSH) with SWCNT. The aim of this work is to investigate possible mechanisms of nanotoxicity at a molecular level. We present details of the molecular dynamics, structure, and free energy of binding of hFSH on the surface of SWCNT. We find that hFSH in aqueous solution strongly adsorbs onto SWCNT via their concave surface as evidenced by high binding free energies for residues in both protein subunits. It was found that hydrophobic, π-cation, and π-π stacking interactions are the main driving forces for the adsorption of the protein at the nanotube surface.

Gum arabic glycoprotein is a twisted hairy rope. A new model based on O-galactosylhydroxyproline as the polysaccharide attachment site

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

Wu Qi; Fong, C.; Lamport, D.T.A.

1991-07-01

Separation of the wound exudate from Acacia senegal (L.) Willd., gum arabic, on a preparative Superose-6 column gave two major fractions: a high molecular weight gum arabic glyco-protein (GAGP) containing about 90% carbohydrate and a lower molecular weight heterogeneous gum arabic polysaccharide fraction. Hydrogen fluoride-deglycosylation of GAGP gave a large hydroxyproline-rich polypeptide backbone (dGAGP). Alkaline hydrolysis of GAGP showed that most of the carbohydrate was attached to the polypeptide backbone as small hydroxyproline (Hyp)-polysaccharide substituents. The data imply a rodlike molecule with numerous small polysaccharide substituents (attached to 24% of the Hyp residues), regularly arranged along a highly periodic polypeptidemore » backbone based, hypothetically, on a 10 to 12 residue repetitive peptide motif. Thus, a simple statistical model of the gum arabic glycoprotein predicts a repeating polysaccharide substituents will maximize intramolecular hydrogen bonding if aligned along the long axis of the molecule, forming in effect a twisted hairy rope. Electron micrographs of rotary shadowed GAGP molecules support that prediction and may also explain show such apparently large molecules can exit the cell by endwise reptation through the small pores of the primary cell wall.« less

Metabolism of Glycoproteins in Turpentine Granuloma*

PubMed Central

Prodi, G.; Pane, G.; Romeo, G.

1970-01-01

The local synthesis of sialic acid and sialic acid containing glycoproteins in granuloma experimentally produced with turpentine has been investigated by incubating them in vitro with 14C glucosamine. The content and activity of chromatographically isolated sialic acid of water soluble and water insoluble fractions of tissue incubated at different times after injection of turpentine was determined. A local synthesis of sialic acid and its incorporation both in the soluble and insoluble fractions were found, with a time depending slope. Chromatography on DEAE Sephadex of glycoproteins obtained from water soluble fraction showed that radioactivity was present in 2 peaks. After papain digestion of the insoluble fraction, the sialic acid containing material could be separated into 2 groups of radioactive glycopeptides on DEAE Sephadex. The data demonstrates that granuloma can synthestize in vitro a considerable variety of glycoproteic materials. PMID:5491911

Chemical synthesis of β-arabinofuranosyl containing oligosaccharides derived from plant cell wall extensins.

PubMed

Kaeothip, Sophon; Boons, Geert-Jan

2013-08-21

Extensins are plant-derived glycoproteins that are densely modified by oligo-arabinofuranosides linked to hydroxyproline residues. These glycoproteins have been implicated in many aspects of plant growth and development. Here, we describe the chemical synthesis of a tetrameric β(1-2)-linked arabinofuranoside that is capped by an α(1-3)-arabinofuranoside and a similar trisaccharide lacking the capping moiety. The challenging β(1-2)-linked arabinofuranosides were installed by using an arabinofuranosyl donor protected with 3,5-O-(di-tert-butylsilane) and a C-2 2-methylnaphthyl (Nap) ether. It was found that the cyclic silane-protecting group of the glycosyl donor greatly increased β-anomeric selectivity. It was, however, imperative to remove the silane-protecting group of an arabinosyl acceptor to achieve optimal anomeric selectivities. The anomeric linker of the synthetic compounds was modified by a biotin moiety for immobilization of the compounds to microtiter plates coated with streptavidine. The resulting microtiter plates were employed to screen for binding against a panel of antibodies elicited against plant cell wall polysaccharides.

Plasmin-Cleaved β-2-Glycoprotein 1 Is an Inhibitor of Angiogenesis

PubMed Central

Sakai, Taro; Balasubramanian, Krishnakumar; Maiti, Sourindra; Halder, Jyotsna B.; Schroit, Alan J.

2007-01-01

β-2-Glycoprotein 1, an abundant plasma glycoprotein, binds anionic cell surfaces and functions as a regulator of thrombosis. Here, we show that cleavage of the kringle domain at Lys317/Thr318 switches its function to a regulator of angiogenesis. In vitro, the cleaved protein specifically inhibited the proliferation and migration of endothelial cells. The protein was without effect on preformed endothelial cell tubes. In vivo, the cleaved protein inhibited neovascularization into subcutaneously implanted Matrigel and Gelfoam sponge implants and the growth of orthotopically injected tumors. Collectively, these data indicate that plasmin-cleaved β-2-glycoprotein 1 is a potent antiangiogenic and antitumor molecule of potential therapeutic significance. PMID:17872974

A sea lamprey glycoprotein hormone receptor similar with gnathostome thyrotropin hormone receptor.

PubMed

Freamat, Mihael; Sower, Stacia A

2008-10-01

The specificity of the vertebrate hypothalamic-pituitary-gonadal and hypothalamic-pituitary-thyroid axes is explained by the evolutionary refinement of the specificity of expression and selectivity of interaction between the glycoprotein hormones GpH (FSH, LH, and TSH) and their cognate receptors GpH-R (FSH-R, LH-R, and TSH-R). These two finely tuned signaling pathways evolved by gene duplication and functional divergence from an ancestral GpH/GpH-R pair. Comparative analysis of the protochordate and gnathostome endocrine systems suggests that this process took place prior or concomitantly with the emergence of the gnathostome lineage. Here, we report identification and characterization of a novel glycoprotein hormone receptor (lGpH-R II) in the Agnathan sea lamprey. This 781 residue protein was found approximately 43% identical with mammalian TSH-R and FSH-R representative sequences, and similarly with these two classes of mammalian receptors it is assembled from ten exons. A synthetic ligand containing the lamprey glycoprotein hormone beta-chain tethered upstream of a mammalian alpha-chain activated the lGpH-R II expressed in COS-7 cells but in a lesser extent than lGpH-R I. Molecular phylogenetic analysis of vertebrate GpH-R protein sequences suggests a closer relationship between lGpH-R II and gnathostome thyrotropin receptors. Overall, the presence and characteristics of the lamprey glycoprotein hormone receptors suggest existence of a primitive functionally overlapping glycoprotein hormone/glycoprotein hormone receptor system in this animal.

Novel functional hepatitis C virus glycoprotein isolates identified using an optimized viral pseudotype entry assay.

PubMed

Urbanowicz, Richard A; McClure, C Patrick; King, Barnabas; Mason, Christopher P; Ball, Jonathan K; Tarr, Alexander W

2016-09-01

Retrovirus pseudotypes are a highly tractable model used to study the entry pathways of enveloped viruses. This model has been extensively applied to the study of the hepatitis C virus (HCV) entry pathway, preclinical screening of antiviral antibodies and for assessing the phenotype of patient-derived viruses using HCV pseudoparticles (HCVpp) possessing the HCV E1 and E2 glycoproteins. However, not all patient-isolated clones produce particles that are infectious in this model. This study investigated factors that might limit phenotyping of patient-isolated HCV glycoproteins. Genetically related HCV glycoproteins from quasispecies in individual patients were discovered to behave very differently in this entry model. Empirical optimization of the ratio of packaging construct and glycoprotein-encoding plasmid was required for successful HCVpp genesis for different clones. The selection of retroviral packaging construct also influenced the function of HCV pseudoparticles. Some glycoprotein constructs tolerated a wide range of assay parameters, while others were much more sensitive to alterations. Furthermore, glycoproteins previously characterized as unable to mediate entry were found to be functional. These findings were validated using chimeric cell-cultured HCV bearing these glycoproteins. Using the same empirical approach we demonstrated that generation of infectious ebolavirus pseudoviruses (EBOVpv) was also sensitive to the amount and ratio of plasmids used, and that protocols for optimal production of these pseudoviruses are dependent on the exact virus glycoprotein construct. These findings demonstrate that it is crucial for studies utilizing pseudoviruses to conduct empirical optimization of pseudotype production for each specific glycoprotein sequence to achieve optimal titres and facilitate accurate phenotyping.

Identification, isolation, and N-terminal sequencing of style glycoproteins associated with self-incompatibility in Nicotiana alata.

PubMed

Jahnen, W; Batterham, M P; Clarke, A E; Moritz, R L; Simpson, R J

1989-05-01

S-Gene-associated glycoproteins (S-glycoproteins) from styles of Nicotiana alata, identified by non-equilibrium two-dimensional electrophoresis, were purified by cation exchange fast protein liquid chromatography with yields of 0.5 to 8 micrograms of protein per style, depending on the S-genotype of the plant. The method relies on the highly basic nature of the S-glycoproteins. The elution profiles of the different S-glycoproteins from the fast protein liquid chromatography column were characteristic of each S-glycoprotein, and could be used to establish the S-genotype of plants in outbreeding populations. In all cases, the S-genotype predicted from the style protein profile corresponded to that predicted from DNA gel blot analysis using S-allele-specific DNA probes and to that established by conventional breeding tests. Amino-terminal sequences of five purified S-glycoproteins showed a high degree of homology with the previously published sequences of N. alata and Lycopersicon esculentum S-glycoproteins.

Synergistic antimicrobial activity of caprylic acid in combination with citric acid against both Escherichia coli O157:H7 and indigenous microflora in carrot juice.

PubMed

Kim, S A; Rhee, M S

2015-08-01

The identification of novel, effective, and non-thermal decontamination methods is imperative for the preservation of unpasteurized and fresh vegetable juices. The aim of this study was to examine the bactericidal effects of caprylic acid + citric acid against the virulent pathogen Escherichia coli O157:H7 and the endogenous microflora in unpasteurized fresh carrot juice. Carrot juice was treated with either caprylic acid, citric acid, or a combination of caprylic acid + citric acid at mild heating temperature (45 °C or 50 °C). The color of the treated carrot juice as well as microbial survival was examined over time. Combined treatment was more effective than individual treatment in terms of both color and microbial survival. Caprylic acid + citric acid treatment (each at 5.0 mM) at 50 °C for 5 min resulted in 7.46 and 3.07 log CFU/ml reductions in the E. coli O157:H7 and endogenous microflora populations, respectively. By contrast, there was no apparent reduction in either population following individual treatment. A validation assay using a low-density E. coli O157:H7 inoculum (3.31 log CFU/ml) showed that combined treatment with caprylic acid (5.0 mM) + citric acid (2.5 mM) at 50 °C for >5 min or with caprylic acid + citric acid (both at 5.0 mM) at either 45 °C or 50 °C for >5 min completely destroyed the bacteria. Combined treatment also increased the redness of the juice, which is a perceived indication of quality. Taken together, these results indicate that combined treatment with low concentrations of caprylic acid and citric acid, which are of biotic origin, can eliminate microorganisms from unpasteurized carrot juice. Copyright © 2015 Elsevier Ltd. All rights reserved.

Combined Effect of Ultrasound and Mild Temperatures on the Inactivation of E. coli in Fresh Carrot Juice and Changes on its Physicochemical Characteristics.

PubMed

Pokhrel, Prashant Raj; Bermúdez-Aguirre, Daniela; Martínez-Flores, Héctor E; Garnica-Romo, M Guadalupe; Sablani, Shyam; Tang, Juming; Barbosa-Cánovas, Gustavo V

2017-10-01

The combination of ultrasound and mild temperatures to process fruits and vegetables juices is a novel approach that is showing promising results for microbial inactivation and preservation of bioactive compounds and sensory attributes. This study centers on investigating the inactivation of Escherichia coli (ATCC 11755) in carrot juice as a result of the combined effect of ultrasound (24 kHz frequency, 120 μm, and 400 W) with temperature (50, 54, and 58 °C) and processing time (0 to 10 min). In addition, the possible changes in physicochemical properties and the retention of bioactive compounds after processing were analyzed. Microbial inactivation with ultrasound treatment at 50 °C resulted in 3.5 log reduction after 10 min, whereas at 54 °C almost 5 log reduction was attained in the same period of time; meanwhile, for treatment at 58 °C, no viable cells were detected (>5 log reduction) after 2 min. There was no significant difference (P > 0.05) on pH (6.80 to 6.82), °Brix (8.0 to 8.5), titratable acidity (0.29% to 0.30%), total carotenoid (1774 to 1835 μg/100 mL), phenolic compounds (20.19 to 20.63 μg/mL), ascorbic acid (4.8 mg/100 mL), and color parameters between fresh and ultrasound treated samples at the studied temperatures. To predict the inactivation patterns, observed values were tested using 3 different general models: first-order, Weibull distribution, and biphasic. The Weibull and biphasic models show good correlation for inactivation under all processing conditions. Results show ultrasound in combination with mild temperature could be effectively used to process fresh carrot juice providing a safe product without affecting physicochemical characteristics. The combination of ultrasound and mild temperatures is effective in reducing microbial load in carrot juice to safe levels. This combination would be beneficial in the industrial processing of carrot juice without altering the quality attributes or bioactive compounds. © 2017 Institute of

Simulations of flow induced structural transition of the β-switch region of glycoprotein Ibα.

PubMed

Han, Mengzhi; Xu, Ji; Ren, Ying; Li, Jinghai

2016-02-01

Binding of glycoprotein Ibα to von Willebrand factor induces platelet adhesion to injured vessel walls and initiates a multistep hemostatic process. It has been hypothesized that the flow condition could induce a loop to β-sheet conformational change in the β-switch region of glycoprotein Ibα, which regulates it binding to the von Willebrand factor and facilitates the blood clot formation and wound healing. In this work, direct molecular dynamics (MD), flow MD and metadynamics, were employed to investigate the mechanisms of this flow induced conformational transition process. Specifically, the free energy landscape of the whole transition process was drawn by metadynamics with the path collective variable approach. The results reveal that without flow, the free energy landscape has two main basins, including a random loop basin stabilized by large conformational entropy and a partially folded β-sheet basin. The free energy barrier separating these two basins is relatively high and the β-switch could not fold from loop to β-sheet state spontaneously. The fully β-sheet conformations located in high free energy regions, which are also unstable and gradually unfold into partially folded β-sheet state with flow. Relatively weak flow could trigger some folding of the β-switch but could not fold it into fully β-sheet state. Under strong flow conditions, the β-switch could readily overcome the high free energy barrier and fold into fully β-sheet state. Copyright © 2015 Elsevier B.V. All rights reserved.

Biochar reduces the bioaccumulation of PAHs from soil to carrot (Daucus carota L.) in the rhizosphere: A mechanism study.

PubMed

Ni, Ni; Song, Yang; Shi, Renyong; Liu, Zongtang; Bian, Yongrong; Wang, Fang; Yang, Xinglun; Gu, Chenggang; Jiang, Xin

2017-12-01

The aim of this study was to reveal the mechanisms on how biochar reduces bioaccumulation of polycyclic aromatic hydrocarbons (PAHs) in tuberous vegetables. Corn straw-derived biochar pyrolyzed at 300°C (CB300) or bamboo-derived biochar pyrolyzed at 700°C (BB700) was amended into PAH-contaminated soil planted with carrot (Daucus carota L.). After 150days, 2% CB300 or 2% BB700 amendments significantly reduced the bioaccumulation of PAHs in carrot root (p<0.05), especially for high-molecular-weight PAHs. In the non-rhizosphere, either CB300 or BB700 suppressed PAH dissipation and decreased the bioavailability via adsorption processes. Compared to the control, the total concentration of PAHs in the rhizosphere was higher in the 2% BB700 treatment but the bioavailable concentration was lower. This indicates that BB700 decreased the bioavailability of PAHs primarily via immobilization (adsorption processes). By contrast, the total and bioavailable PAH concentrations were both lower in the 2% CB300 treatment than those in the control. The abundance of bacteria such as Arthrobacter and Flavobacterium and the total number of genes playing important roles in microbial PAH degradation processes increased significantly (p<0.05), which were likely responsible for the rapid dissipation of PAHs in the 2% CB300 treatment in the rhizosphere. These results indicate that CB300 decreased the PAH bioavailability primarily via increasing degradation of PAHs by indigenous microorganisms. The two biochars both showed better effectiveness at reducing the bioavailability of high-molecular-weight PAHs than the low-molecular-weight PAHs in the rhizosphere. Therefore, the mechanisms on how biochar reduces the PAH uptake into carrot are dependent on the type of biochar (e.g., pyrolysis temperature and feedstock) and root presence. Copyright © 2017 Elsevier B.V. All rights reserved.

Formation of norisoprenoid flavor compounds in carrot (Daucus carota L.) roots: characterization of a cyclic-specific carotenoid cleavage dioxygenase 1 gene

USDA-ARS?s Scientific Manuscript database

Carotenoids are isoprenoid pigments that upon oxidative cleavage lead to the production of norisoprenoids that have profound effect on flavor and aromas of agricultural produce. The biosynthetic pathway to norisoprenoids in carrots (Daucus carota L.) is still widely unknown. We found that geranial i...

Analysis of the 3H8 antigen of Candida albicans reveals new aspects of the organization of fungal cell wall proteins.

PubMed

Sentandreu, Rafael; Caminero, Antonio; Rentería, Itzel; León-Ramirez, Claudia; González-de-la-Vara, Luis; Valentin-Gomez, Eulogio; Ruiz-Herrera, José

2018-06-01

The walls of both, yeast and mycelial cells of Candida albicans possess a species-specific antigen that is recognized by a monoclonal antibody (MAb 3H8). This antigen can be extracted in the form of a very high Mr complex, close or over 106 Da, by treatment, with β-1,3-glucanase, β mercaptoethanol or dithothreitol, or mild alkali, but not by saturated hydrogen fluoride (HF) in pyridine, suggesting that the complex is bound to wall β-1,3 glucans, and to proteins by disulfide bonds, but not to β-1,6 glucans. Through its sensitivity to trypsin and different deglycosylation procedures, it was concluded that the epitope is associated to a glycoprotein containing N-glycosidic, but not O-glycosidic mannan moieties. By means of electrophoresis in polycrylamide gradient gels, followed by mass spectrometric analysis, the epitope was pinpointed to a very high MW complex containing Agglutinin-Like Sequence (ALS) family proteins, and other cytoplasmic, membrane and secreted proteins. The components of this complex are bound by unknown covalent bonds. The material extracted with β mercaptoethanol or dilute alkali appeared under the electron microscope as large aggregates in the form of spheroidal and mostly web-like structures of large sizes. These, and additional data, suggest that this protein complex may constitute an important part of the basic glycoprotein structure of C. albicans. The possibility that similar complexes exist in the wall of other fungi is an attractive, although yet untested possibility.

21 CFR 866.5430 - Beta-2-glycoprotein I immunological test system.

Code of Federal Regulations, 2012 CFR

2012-04-01

... the beta-2-glycoprotein I (a serum protein) in serum and other body fluids. Measurement of beta-2... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Beta-2-glycoprotein I immunological test system. 866.5430 Section 866.5430 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN...

21 CFR 866.5430 - Beta-2-glycoprotein I immunological test system.

Code of Federal Regulations, 2014 CFR

2014-04-01

... the beta-2-glycoprotein I (a serum protein) in serum and other body fluids. Measurement of beta-2... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Beta-2-glycoprotein I immunological test system. 866.5430 Section 866.5430 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN...

21 CFR 866.5440 - Beta-2-glycoprotein III immunological test system.

Code of Federal Regulations, 2012 CFR

2012-04-01

... the beta-2-glycoprotein III (a serum protein) in serum and other body fluids. Measurement of beta-2... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Beta-2-glycoprotein III immunological test system. 866.5440 Section 866.5440 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN...

21 CFR 866.5430 - Beta-2-glycoprotein I immunological test system.

Code of Federal Regulations, 2011 CFR

2011-04-01

... the beta-2-glycoprotein I (a serum protein) in serum and other body fluids. Measurement of beta-2... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Beta-2-glycoprotein I immunological test system. 866.5430 Section 866.5430 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN...

21 CFR 866.5440 - Beta-2-glycoprotein III immunological test system.

Code of Federal Regulations, 2013 CFR

2013-04-01

... the beta-2-glycoprotein III (a serum protein) in serum and other body fluids. Measurement of beta-2... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Beta-2-glycoprotein III immunological test system. 866.5440 Section 866.5440 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN...

21 CFR 866.5440 - Beta-2-glycoprotein III immunological test system.

Code of Federal Regulations, 2011 CFR

2011-04-01

... the beta-2-glycoprotein III (a serum protein) in serum and other body fluids. Measurement of beta-2... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Beta-2-glycoprotein III immunological test system. 866.5440 Section 866.5440 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN...

21 CFR 866.5430 - Beta-2-glycoprotein I immunological test system.

Code of Federal Regulations, 2013 CFR

2013-04-01

... the beta-2-glycoprotein I (a serum protein) in serum and other body fluids. Measurement of beta-2... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Beta-2-glycoprotein I immunological test system. 866.5430 Section 866.5430 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN...

21 CFR 866.5440 - Beta-2-glycoprotein III immunological test system.

Code of Federal Regulations, 2014 CFR

2014-04-01

... the beta-2-glycoprotein III (a serum protein) in serum and other body fluids. Measurement of beta-2... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Beta-2-glycoprotein III immunological test system. 866.5440 Section 866.5440 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN...

A gene-derived SNP-based high resolution linkage map of carrot including the location of QTL conditioning root and leaf anthocyanin pigmentation

USDA-ARS?s Scientific Manuscript database

Background: Purple carrots accumulate large quantities of anthocyanins in their roots and leaves. These flavonoid pigments possess antioxidant activity and are implicated in providing health benefits. The lack of informative and saturated linkage maps associated with well characterized populations s...

Increased Expression of P-Glycoprotein Is Associated With Chlorpyrifos Resistance in the German Cockroach (Blattodea: Blattellidae).

PubMed

Hou, Weiyuan; Jiang, Chu; Zhou, Xiaojie; Qian, Kun; Wang, Lei; Shen, Yanhui; Zhao, Yan

2016-12-01

A principal method for control of the German cockroach, Blattella germanica (L.), is the broad-spectrum organophosphorus insecticide, chlorpyrifos (O,O-diethyl O-3,5,6-trichloro-2-pyridyl phosphorothioate); however, extensive and repeated application has resulted in the development of resistance to chlorpyrifos in this insect. Evidence suggests that ATP-binding cassette protein transporters, including P-glycoprotein, are involved in insecticide resistance. However, little is known of the role of P-glycoprotein in insecticide resistance in the German cockroach. Here, we developed a chlorpyrifos-resistant strain of German cockroach and investigated the relationship between P-glycoprotein and chlorpyrifos resistance using toxicity assays; inhibition studies with two P-glycoprotein inhibitors, verapamil and quinine; P-glycoprotein-ATPase activity assays; and western blotting analysis. After 23 generations of selection from susceptible strain cockroaches, we obtained animals with high resistance to chlorpyrifos. When P-glycoprotein-ATPase activity was inhibited by verapamil and quinine, we observed enhanced susceptibility to chlorpyrifos in both control and chlorpyrifos-resistant cockroaches. No significant alterations of P-glycoprotein expression or ATPase activity were observed in cockroaches acutely exposed to LD50 doses of chlorpyrifos for 24 h, while P-glycoprotein expression and ATPase activity were clearly elevated in the chlorpyrifos-resistant cockroach strain. Thus, we conclude that P-glycoprotein is associated with chlorpyrifos resistance in the German cockroach and that elevated levels of P-glycoprotein expression and ATPase activity may be an important mechanism of chlorpyrifos resistance in the German cockroach. © The Authors 2016. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Making Home Sweet and Sturdy: Toxoplasma gondii ppGalNAc-Ts Glycosylate in Hierarchical Order and Confer Cyst Wall Rigidity.

PubMed

Tomita, Tadakimi; Sugi, Tatsuki; Yakubu, Rama; Tu, Vincent; Ma, Yanfen; Weiss, Louis M

2017-01-10

The protozoan intracellular parasite Toxoplasma gondii forms latent cysts in the central nervous system (CNS) and persists for the lifetime of the host. This cyst is cloaked with a glycosylated structure called the cyst wall. Previously, we demonstrated that a mucin-like glycoprotein, CST1, localizes to the cyst wall and confers structural rigidity on brain cysts in a mucin-like domain-dependent manner. The mucin-like domain of CST1 is composed of 20 units of threonine-rich tandem repeats that are O-GalNAc glycosylated. A family of enzymes termed polypeptide N-acetylgalactosaminyltransferases (ppGalNAc-Ts) initiates O-GalNAc glycosylation. To identify which isoforms of ppGalNAc-Ts are responsible for the glycosylation of the CST1 mucin-like domain and to evaluate the function of each ppGalNAc-T in the overall glycosylation of the cyst wall, all five ppGalNAc-T isoforms were deleted individually from the T. gondii genome. The ppGalNAc-T2 and -T3 deletion mutants produced various glycosylation defects on the cyst wall, implying that many cyst wall glycoproteins are glycosylated by T2 and T3. Both T2 and T3 glycosylate the CST1 mucin-like domain, and this glycosylation is necessary for CST1 to confer structural rigidity on the cyst wall. We established that T2 is required for the initial glycosylation of the mucin-like domain and that T3 is responsible for the sequential glycosylation on neighboring acceptor sites, demonstrating hierarchical glycosylation by two distinct initiating and filling-in ppGalNAc-Ts in an intact organism. Toxoplasma gondii is an obligate intracellular parasite that infects a third of the world's population. It can cause severe congenital disease and devastating encephalitis in immunocompromised individuals. We identified two glycosyltransferases, ppGalNAc-T2 and -T3, which are responsible for glycosylating cyst wall proteins in a hierarchical fashion. This glycosylation confers structural rigidity on the brain cyst. Our studies provide new

The Super Efficient Refrigerator Program: Case study of a Golden Carrot program

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

Eckert, J B

1995-07-01

The work in this report was conducted by the Analytic Studies Division (ASD) of the National Renewable Energy Laboratory (NREL) for the U.S. Department of Energy Office of Energy Efficiency and Renewable Energy, Office of Building Technologies. This case study describes the development and implementation of the Super Efficient Refrigerator Program (SERP), which awarded $30 million to the refrigerator manufacturer that developed and commercialized a refrigerator that exceeded 1993 federal efficiency standards by at least 25%. The program was funded by 24 public and private utilities. As the first Golden Carrot program to be implemented in the United States, SERPmore » was studied as an example for future `market-pull` efforts.« less

Demonstration that endoplasmic reticulum-associated degradation of glycoproteins can occur downstream of processing by endomannosidase.

PubMed

Kukushkin, Nikolay V; Alonzi, Dominic S; Dwek, Raymond A; Butters, Terry D

2011-08-15

During quality control in the ER (endoplasmic reticulum), nascent glycoproteins are deglucosylated by ER glucosidases I and II. In the post-ER compartments, glycoprotein endo-α-mannosidase provides an alternative route for deglucosylation. Previous evidence suggests that endomannosidase non-selectively deglucosylates glycoproteins that escape quality control in the ER, facilitating secretion of aberrantly folded as well as normal glycoproteins. In the present study, we employed FOS (free oligosaccharides) released from degrading glycoproteins as biomarkers of ERAD (ER-associated degradation), allowing us to gain a global rather than single protein-centred view of ERAD. Glucosidase inhibition was used to discriminate between glucosidase- and endomannosidase-mediated ERAD pathways. Endomannosidase expression was manipulated in CHO (Chinese-hamster ovary)-K1 cells, naturally lacking a functional version of the enzyme, and HEK (human embryonic kidney)-293T cells. Endomannosidase was shown to decrease the levels of total FOS, suggesting decreased rates of ERAD. However, following pharmacological inhibition of ER glucosidases I and II, endomannosidase expression resulted in a partial switch between glucosylated FOS, released from ER-confined glycoproteins, to deglucosylated FOS, released from endomannosidase-processed glycoproteins transported from the Golgi/ERGIC (ER/Golgi intermediate compartment) to the ER. Using this approach, we have identified a previously unknown pathway of glycoprotein flow, undetectable by the commonly employed methods, in which secretory cargo is targeted back to the ER after being processed by endomannosidase. © The Authors Journal compilation © 2011 Biochemical Society

Improved platelet survival after cold storage by prevention of glycoprotein Ibα clustering in lipid rafts

PubMed Central

Gitz, Eelo; Koekman, Cornelis A; van den Heuvel, Dave J.; Deckmyn, Hans; Akkerman, Jan W.; Gerritsen, Hans C.; Urbanus, Rolf T.

2012-01-01

Background Storing platelets for transfusion at room temperature increases the risk of microbial infection and decreases platelet functionality, leading to out-date discard rates of up to 20%. Cold storage may be a better alternative, but this treatment leads to rapid platelet clearance after transfusion, initiated by changes in glycoprotein Ibα, the receptor for von Willebrand factor. Design and Methods: We examined the change in glycoprotein Ibα distribution using Förster resonance energy transfer by time-gated fluorescence lifetime imaging microscopy. Results Cold storage induced deglycosylation of glycoprotein Ibα ectodomain, exposing N-acetyl-Dglucosamine residues, which sequestered with GM1 gangliosides in lipid rafts. Raft-associated glycoprotein Ibα formed clusters upon binding of 14-3-3ζ adaptor proteins to its cytoplasmic tail, a process accompanied by mitochondrial injury and phosphatidyl serine exposure. Cold storage left glycoprotein Ibα surface expression unchanged and although glycoprotein V decreased, the fall did not affect glycoprotein Ibα clustering. Prevention of glycoprotein Ibα clustering by blockade of deglycosylation and 14-3-3ζ translocation increased the survival of cold-stored platelets to above the levels of platelets stored at room temperature without compromising hemostatic functions. Conclusions We conclude that glycoprotein Ibα translocates to lipid rafts upon cold-induced deglycosylation and forms clusters by associating with 14-3-3ζ. Interference with these steps provides a means to enable cold storage of platelet concentrates in the near future. PMID:22733027

UDP-glucose:glycoprotein glucosyltransferase (UGGT1) promotes substrate solubility in the endoplasmic reticulum

PubMed Central

Ferris, Sean P.; Jaber, Nikita S.; Molinari, Maurizio; Arvan, Peter; Kaufman, Randal J.

2013-01-01

Protein folding in the endoplasmic reticulum (ER) is error prone, and ER quality control (ERQC) processes ensure that only correctly folded proteins are exported from the ER. Glycoproteins can be retained in the ER by ERQC, and this retention contributes to multiple human diseases, termed ER storage diseases. UDP-glucose:glycoprotein glucosyltransferase (UGGT1) acts as a central component of glycoprotein ERQC, monoglucosylating deglucosylated N-glycans of incompletely folded glycoproteins and promoting subsequent reassociation with the lectin-like chaperones calreticulin and calnexin. The extent to which UGGT1 influences glycoprotein folding, however, has only been investigated for a few selected substrates. Using mouse embryonic fibroblasts lacking UGGT1 or those with UGGT1 complementation, we investigated the effect of monoglucosylation on the soluble/insoluble distribution of two misfolded α1-antitrypsin (AAT) variants responsible for AAT deficiency disease: null Hong Kong (NHK) and Z allele. Whereas substrate solubility increases directly with the number of N-linked glycosylation sites, our results indicate that additional solubility is conferred by UGGT1 enzymatic activity. Monoglucosylation-dependent solubility decreases both BiP association with NHK and unfolded protein response activation, and the solubility increase is blocked in cells deficient for calreticulin. These results suggest that UGGT1-dependent monoglucosylation of N-linked glycoproteins promotes substrate solubility in the ER. PMID:23864712

Spatial localization of the Ebola virus glycoprotein mucin-like domain determined by cryo-electron tomography.

PubMed

Tran, Erin E H; Simmons, James A; Bartesaghi, Alberto; Shoemaker, Charles J; Nelson, Elizabeth; White, Judith M; Subramaniam, Sriram

2014-09-01

The Ebola virus glycoprotein mucin-like domain (MLD) is implicated in Ebola virus cell entry and immune evasion. Using cryo-electron tomography of Ebola virus-like particles, we determined a three-dimensional structure for the full-length glycoprotein in a near-native state and compared it to that of a glycoprotein lacking the MLD. Our results, which show that the MLD is located at the apex and the sides of each glycoprotein monomer, provide a structural template for analysis of MLD function. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Requirements for cell rounding and surface protein down-regulation by Ebola virus glycoprotein.

PubMed

Francica, Joseph R; Matukonis, Meghan K; Bates, Paul

2009-01-20

Ebola virus causes an acute hemorrhagic fever that is associated with high morbidity and mortality. The viral glycoprotein is thought to contribute to pathogenesis, though precise mechanisms are unknown. Cellular pathogenesis can be modeled in vitro by expression of the Ebola viral glycoprotein (GP) in cells, which causes dramatic morphological changes, including cell rounding and surface protein down-regulation. These effects are known to be dependent on the presence of a highly glycosylated region of the glycoprotein, the mucin domain. Here we show that the mucin domain from the highly pathogenic Zaire subtype of Ebola virus is sufficient to cause characteristic cytopathology when expressed in the context of a foreign glycoprotein. Similarly to full length Ebola GP, expression of the mucin domain causes rounding, detachment from the extracellular matrix, and the down-regulation of cell surface levels of beta1 integrin and major histocompatibility complex class 1. These effects were not seen when the mucin domain was expressed in the context of a glycophosphatidylinositol-anchored isoform of the foreign glycoprotein. In contrast to earlier analysis of full length Ebola glycoproteins, chimeras carrying the mucin domains from the Zaire and Reston strains appear to cause similar levels of down-modulation and cell detachment. Cytopathology associated with Ebola glycoprotein expression does not occur when GP expression is restricted to the endoplasmic reticulum. In contrast to a previously published report, our results demonstrate that GP-induced surface protein down-regulation is not mediated through a dynamin-dependent pathway. Overall, these results support a model in which the mucin domain of Ebola GP acts at the cell surface to induce protein down modulation and cytopathic effects.

Phospholipid flippase activity of the reconstituted P-glycoprotein multidrug transporter.

PubMed

Romsicki, Y; Sharom, F J

2001-06-12

The P-glycoprotein multidrug transporter acts as an ATP-powered efflux pump for a large variety of hydrophobic drugs, natural products, and peptides. The protein is proposed to interact with its substrates within the hydrophobic interior of the membrane. There is indirect evidence to suggest that P-glycoprotein can also transport, or "flip", short chain fluorescent lipids between leaflets of the membrane. In this study, we use a fluorescence quenching technique to directly show that P-glycoprotein reconstituted into proteoliposomes translocates a wide variety of NBD lipids from the outer to the inner leaflet of the bilayer. Flippase activity depended on ATP hydrolysis at the outer surface of the proteoliposome, and was inhibited by vanadate. P-Glycoprotein exhibited a broad specificity for phospholipids, and translocated phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, and sphingomyelin. Lipid derivatives that were flipped included molecules with long, short, unsaturated, and saturated acyl chains and species with the NBD group covalently linked to either acyl chains or the headgroup. The extent of lipid translocation from the outer to the inner leaflet in a 20 min period at 37 degrees C was directly estimated, and fell in the range of 0.36-1.83 nmol/mg of protein. Phospholipid flipping was inhibited in a concentration-dependent, saturable fashion by various substrates and modulators, including vinblastine, verapamil, and cyclosporin A, and the efficiency of inhibition correlated well with the affinity of binding to Pgp. Taken together, these results suggest that P-glycoprotein carries out both lipid translocation and drug transport by the same path. The transporter may be a generic flippase for hydrophobic molecules with the correct steric attributes that are present within the membrane interior.

Pathobiology of Pneumocystis pneumonia: life cycle, cell wall and cell signal transduction.

PubMed

Skalski, Joseph H; Kottom, Theodore J; Limper, Andrew H

2015-09-01

Pneumocystis is a genus of ascomycetous fungi that are highly morbid pathogens in immunosuppressed humans and other mammals. Pneumocystis cannot easily be propagated in culture, which has greatly hindered understanding of its pathobiology. The Pneumocystis life cycle is intimately associated with its mammalian host lung environment, and life cycle progression is dependent on complex interactions with host alveolar epithelial cells and the extracellular matrix. The Pneumocystis cell wall is a varied and dynamic structure containing a dominant major surface glycoprotein, β-glucans and chitins that are important for evasion of host defenses and stimulation of the host immune system. Understanding of Pneumocystis cell signaling pathways is incomplete, but much has been deduced by comparison of the Pneumocystis genome with homologous genes and proteins in related fungi. In this mini-review, the pathobiology of Pneumocystis is reviewed, with particular focus on the life cycle, cell wall components and cell signal transduction. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Interaction of forskolin with the P-glycoprotein multidrug transporter

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

Ming s, D.I.; Seamon, K.B.; Speicher, L.A.

1991-08-27

Forskolin and 1,9-dideoxyforskolin, an analogue that does not activate adenylyl cyclase, were tested for their ability to enhance the cytotoxic effects of adriamycin in human ovarian carcinoma cells, SKOV3, which are sensitive to adriamycin and express low levels of P-glycoprotein, and a variant cell line, SKVLB, which overexpresses the P-glycoprotein and has the multidrug reing ance (MDR) phenotype. Forskolin and 1,9-dideoxyforskolin both increased the cytotoxic effects of adriamycin in SKVLB cells, yet had no effect on SKOV3 cells. Two photoactive derivatives of forskolin have been synthesized, 7-O-((2-(3-(4-azido-3-({sup 125}I)iodophenyl)propionamido)ethyl)carbamyl)forskolin, {sup 125}I-6-AIPP-Fsk, and 6-O-((2-(3-(4-azido-3-({sup 125}I)iodophenyl)propionamido)ethyl)carbamyl)forskolin, {sup 125}I-6-AIPP-Fsk, which exhibit specificity for labelingmore » the glucose transporter and aing lyl cyclase, respectively. Both photolabels identified a 140-kDa protein in membranes from SKVLB cells whose labeling was inhibited by forskolin and 1,9-dideoxyforskolin. The data are consistent with forskolin binding to the P-glycoprotein analogous to that of other chemosensitizing drugs that have been shown to partially reverse MDR. The ability of forskolin photolabels to specifically label the transporter, the adenylyl cyclase, and the P-glycoprotein suggests that these proteins may share a common biing g domain for forskolin analogues.« less

Marine Natural Products with P-Glycoprotein Inhibitor Properties

PubMed Central

Lopez, Dioxelis; Martinez-Luis, Sergio

2014-01-01

P-glycoprotein (P-gp) is a protein belonging to the ATP-binding cassette (ABC) transporters superfamily that has clinical relevance due to its role in drug metabolism and multi-drug resistance (MDR) in several human pathogens and diseases. P-gp is a major cause of drug resistance in cancer, parasitic diseases, epilepsy and other disorders. This review article aims to summarize the research findings on the marine natural products with P-glycoprotein inhibitor properties. Natural compounds that modulate P-gp offer great possibilities for semi-synthetic modification to create new drugs and are valuable research tools to understand the function of complex ABC transporters. PMID:24451193

Glycoproteins functionalized natural and synthetic polymers for prospective biomedical applications: A review.

PubMed

Tabasum, Shazia; Noreen, Aqdas; Kanwal, Arooj; Zuber, Mohammad; Anjum, Muhammad Naveed; Zia, Khalid Mahmood

2017-05-01

Glycoproteins have multidimensional properties such as biodegradability, biocompatibility, non-toxicity, antimicrobial and adsorption properties; therefore, they have wide range of applications. They are blended with different polymers such as chitosan, carboxymethyl cellulose (CMC), polyvinyl pyrrolidone (PVP), polycaprolactone (PCL), heparin, polystyrene fluorescent nanoparticles (PS-NPs) and carboxyl pullulan (PC) to improve their properties like thermal stability, mechanical properties, resistance to pH, chemical stability and toughness. Considering the versatile charateristics of glycoprotein based polymers, this review sheds light on synthesis and characterization of blends and composites of glycoproteins, with natural and synthetic polymers and their potential applications in biomedical field such as drug delivery system, insulin delivery, antimicrobial wound dressing uses, targeting of cancer cells, development of anticancer vaccines, development of new biopolymers, glycoproteome research, food product and detection of dengue glycoproteins. All the technical scientific issues have been addressed; highlighting the recent advancement. Copyright © 2017 Elsevier B.V. All rights reserved.

[C-terminal lysosome targeting domain of CD63 modifies cellular localization of rabies virus glycoprotein].

PubMed

Starodubova, E S; Kuzmenko, Y V; Latanova, A A; Preobrazhenskaya, O V; Karpov, V L

2017-01-01

The glycoprotein of rabies virus is the central antigen elicited the immune response to infection; therefore, the majority of developing anti-rabies vaccines are based on this protein. In order to increase the efficacy of DNA immunogen encoding rabies virus glycoprotein, the construction of chimeric protein with the CD63 domain has been proposed. The CD63 is a transmembrane protein localized on the cell surface and in lysosomes. The lysosome targeting motif GYEVM is located at its C-terminus. We used the domain that bears this motif (c-CD63) to generate chimeric glycoprotein in order to relocalize it into lysosomes. Here, it was shown that, in cells transfected with plasmid that encodes glycoprotein with c-CD63 motif at the C-terminus, the chimeric protein was predominantly observed in lysosomes and at the cell membrane where the unmodified glycoprotein is localized in the endoplasmic reticulum and at the cell surface. We suppose that current modification of the glycoprotein may improve the immunogenicity of anti-rabies DNA vaccines due to more efficient antibody production.

Combined effect of starch/montmorillonite coating and passive MAP in antioxidant activity, total phenolics, organic acids and volatile of fresh-cut carrots.

PubMed

Guimarães, Isabela Costa; dos Reis, Kelen Cristina; Menezes, Evandro Galvão Tavares; Borges, Paulo Rogério Siriano; Rodrigues, Ariel Costa; Leal, Renato; Hernandes, Thais; de Carvalho, Elisângela Helena Nunes; Vilas Boas, Eduardo Valério de Barros

2016-01-01

This work evaluates fresh-cut carrots (FCC) coated with montmorillonite (MMT) subjected to passive modified atmosphere packaging. Carrots were sanitized, cooled, peeled and sliced. Half of the FCC were coated with MMT nanoparticle film and the other half were not. All FCCs were packed in a polypropylene rigid tray, covered with a polypropylene rigid lid or sealed with polyethylene + propylene film, in four treatments (RL, rigid lid; RLC, rigid lid + coating; ST, sealed tray; STC, sealed tray + coating). FCCs were stored at 4 °C and were analyzed weekly for 4 weeks (total antioxidant activity by 2,2-diphenyl-1-picryl hydrazyl method and the β-carotene/linoleic acid, phenolic compounds, organic acids and volatile compounds). The use of coating film with starch nanoparticles and a modified atmosphere leads to the preservation of the total antioxidant activity, the volatile and organic acids of FCC.

Yield, chemical composition and nutritional quality responses of carrot, radish and turnip to elevated atmospheric carbon dioxide.

PubMed

Azam, Andaleeb; Khan, Ikhtiar; Mahmood, Abid; Hameed, Abdul

2013-10-01

Future concentration of carbon dioxide in the atmosphere is very important due to its apparent economic and environmental impact in terms of climate change. However, a compressive assessment of its effect on the nutritional and chemical characteristics of food crops has yet to be established. In the present study the impact of elevated atmospheric CO2 on the yield, chemical composition and nutritional quality of three root vegetables, carrot (Daucus carota L. cv. T-1-111), radish (Raphanus sativus L. cv. Mino) and turnip (Brassica rapa L. cv. Grabe) has been investigated. The yield of carrot, radish and turnip increased by 69, 139 and 72%, respectively, when grown under elevated CO2 conditions. Among the proximate composition, protein, vitamin C and fat contents decreased significantly for all the vegetables while sugar and fibre contents were increased. Response of the vegetables to elevated CO2 , in terms of elemental composition, was different with a significant decrease in many important minerals. Elevated CO2 decreased the amount of majority of the fatty acids and amino acids in these vegetables. It was observed that elevated CO2 increased the yield of root vegetables but many important nutritional parameters including protein, vitamin C, minerals, essential fatty acids and amino acids were decreased. © 2013 Society of Chemical Industry.

Encapsulation of black carrot juice using spray and freeze drying.

PubMed

Murali, S; Kar, Abhijit; Mohapatra, Debabandya; Kalia, Pritam

2015-12-01

Black carrot juice extracted using pectinase enzyme was encapsulated in three different carrier materials (maltodextrin 20DE, gum arabic and tapioca starch) using spray drying at four inlet temperatures (150, 175, 200 and 225 ℃) and freeze drying at a constant temperature of - 53 ℃ and vacuum of 0.22-0.11 mbar with the constant feed mixture. The products were analyzed for total anthocyanin content, antioxidant activity, water solubility index, encapsulation efficiency and total colour change. For both the drying methods followed in this study, maltodextrin 20DE as the carrier material has proven to be better in retaining maximum anthocyanin and antioxidant activity compared to gum arabic and tapioca starch. The best spray dried product, was obtained at 150 ℃. The most acceptable was the freeze dried product with maximum anthocyanin content, antioxidant activity, water solubility index, encapsulation efficiency and colour change. © The Author(s) 2014.

Statins Suppress Ebola Virus Infectivity by Interfering with Glycoprotein Processing.

PubMed

Shrivastava-Ranjan, Punya; Flint, Mike; Bergeron, Éric; McElroy, Anita K; Chatterjee, Payel; Albariño, César G; Nichol, Stuart T; Spiropoulou, Christina F

2018-05-01

Ebola virus (EBOV) infection is a major public health concern due to high fatality rates and limited effective treatments. Statins, widely used cholesterol-lowering drugs, have pleiotropic mechanisms of action and were suggested as potential adjunct therapy for Ebola virus disease (EVD) during the 2013-2016 outbreak in West Africa. Here, we evaluated the antiviral effects of statin (lovastatin) on EBOV infection in vitro Statin treatment decreased infectious EBOV production in primary human monocyte-derived macrophages and in the hepatic cell line Huh7. Statin treatment did not interfere with viral entry, but the viral particles released from treated cells showed reduced infectivity due to inhibition of viral glycoprotein processing, as evidenced by decreased ratios of the mature glycoprotein form to precursor form. Statin-induced inhibition of infectious virus production and glycoprotein processing was reversed by exogenous mevalonate, the rate-limiting product of the cholesterol biosynthesis pathway, but not by low-density lipoprotein. Finally, statin-treated cells produced EBOV particles devoid of the surface glycoproteins required for virus infectivity. Our findings demonstrate that statin treatment inhibits EBOV infection and suggest that the efficacy of statin treatment should be evaluated in appropriate animal models of EVD. IMPORTANCE Treatments targeting Ebola virus disease (EVD) are experimental, expensive, and scarce. Statins are inexpensive generic drugs that have been used for many years for the treatment of hypercholesterolemia and have a favorable safety profile. Here, we show the antiviral effects of statins on infectious Ebola virus (EBOV) production. Our study reveals a novel molecular mechanism in which statin regulates EBOV particle infectivity by preventing glycoprotein processing and incorporation into virus particles. Additionally, statins have anti-inflammatory and immunomodulatory effects. Since inflammation and dysregulation of the immune

P-glycoprotein substrate transport assessed by comparing cellular and vesicular ATPase activity.

PubMed

Nervi, Pierluigi; Li-Blatter, Xiaochun; Aänismaa, Päivi; Seelig, Anna

2010-03-01

We compared the P-glycoprotein ATPase activity in inside-out plasma membrane vesicles and living NIH-MDR1-G185 cells with the aim to detect substrate transport. To this purpose we used six substrates which differ significantly in their passive influx through the plasma membrane. In cells, the cytosolic membrane leaflet harboring the substrate binding site of P-glycoprotein has to be approached by passive diffusion through the lipid membrane, whereas in inside-out plasma membrane vesicles, it is accessible directly from the aqueous phase. Compounds exhibiting fast passive influx compared to active efflux by P-glycoprotein induced similar ATPase activity profiles in cells and inside-out plasma membrane vesicles, because their concentrations in the cytosolic leaflets were similar. Compounds exhibiting similar influx as efflux induced in contrast different ATPase activity profiles in cells and inside-out vesicles. Their concentration was significantly lower in the cytosolic leaflet of cells than in the cytosolic leaflet of inside-out membrane vesicles, indicating that P-glycoprotein could cope with passive influx. P-glycoprotein thus transported all compounds at a rate proportional to ATP hydrolysis (i.e. all compounds were substrates). However, it prevented substrate entry into the cytosol only if passive influx of substrates across the lipid bilayer was in a similar range as active efflux. Copyright 2009 Elsevier B.V. All rights reserved.

Prevalence of Clostridium botulinum and thermophilic heat-resistant spores in raw carrots and green beans used in French canning industry.

PubMed

Sevenier, V; Delannoy, S; André, S; Fach, P; Remize, F

2012-04-16

Two categories of vegetables (carrots and green beans) that are widely used in the manufacture of canned food were surveyed for their spore contamination. Samples were recovered from 10 manufactures spread over all producing areas in France. Two samples over 316 raw vegetables collected were found positive for botulinum neurotoxin producing Clostridia spores as tested by PCR-based GeneDisc assay. Both positive samplestested positive for the type B neurotoxin gene (bont/B). In parallel, heat-resistant spores of thermophilic bacteria that are likely to be associated with canned food spoilage after prolonged incubation at 55 °C were surveyed after specific enrichment. Prevalence varied between 1.6% for Moorella thermoacetica/thermoautotrophica in green bean samples and 8.6% for either Geobacillus stearothermophilus or Thermoanaerobacterium spp. in carrot samples. Vegetable preparation, e.g. washing and edge cutting, considerably reduced spore contamination levels. These data constitute the first wide examination of vegetables specifically cultivated for industrialpurposes for their contamination by spores of thermophilic bacterial species. Copyright © 2012 Elsevier B.V. All rights reserved.

The NS1 Glycoprotein Can Generate Dramatic Antibody-Enhanced Dengue Viral Replication in Normal Out-Bred Mice Resulting in Lethal Multi-Organ Disease

PubMed Central

Falconar, Andrew K. I.; Martinez, Fernando

2011-01-01

Antibody-enhanced replication (AER) of dengue type-2 virus (DENV-2) strains and production of antibody-enhanced disease (AED) was tested in out-bred mice. Polyclonal antibodies (PAbs) generated against the nonstructural-1 (NS1) glycoprotein candidate vaccine of the New Guinea-C (NG-C) or NSx strains reacted strongly and weakly with these antigens, respectively. These PAbs contained the IgG2a subclass, which cross-reacted with the virion-associated envelope (E) glycoprotein of the DENV-2 NSx strain, suggesting that they could generate its AER via all mouse Fcγ-receptor classes. Indeed, when these mice were challenged with a low dose (<0.5 LD50) of the DENV-2 NSx strain, but not the NG-C strain, they all generated dramatic and lethal DENV-2 AER/AED. These AER/AED mice developed life-threatening acute respiratory distress syndrome (ARDS), displayed by diffuse alveolar damage (DAD) resulting from i) dramatic interstitial alveolar septa-thickening with mononuclear cells, ii) some hyperplasia of alveolar type-II pneumocytes, iii) copious intra-alveolar protein secretion, iv) some hyaline membrane-covered alveolar walls, and v) DENV-2 antigen-positive alveolar macrophages. These mice also developed meningo-encephalitis, with greater than 90,000-fold DENV-2 AER titers in microglial cells located throughout their brain parenchyma, some of which formed nodules around dead neurons. Their spleens contained infiltrated megakaryocytes with DENV-2 antigen-positive red-pulp macrophages, while their livers displayed extensive necrosis, apoptosis and macro- and micro-steatosis, with DENV-2 antigen-positive Kuppfer cells and hepatocytes. Their infections were confirmed by DENV-2 isolations from their lungs, spleens and livers. These findings accord with those reported in fatal human “severe dengue” cases. This DENV-2 AER/AED was blocked by high concentrations of only the NG-C NS1 glycoprotein. These results imply a potential hazard of DENV NS1 glycoprotein-based vaccines

Intracellular transport and stability of varicella-zoster virus glycoprotein K

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

Hall, Susan L.; Govero, Jennifer L.; Heineman, Thomas C.

2007-02-20

VZV gK, an essential glycoprotein that is conserved among the alphaherpesviruses, is believed to participate in membrane fusion and cytoplasmic virion morphogenesis based on analogy to its HSV-1 homolog. However, the production of VZV gK-specific antibodies has proven difficult presumably due to its highly hydrophobic nature and, therefore, VZV gK has received limited study. To overcome this obstacle, we inserted a FLAG epitope into gK near its amino terminus and produced VZV recombinants expressing epitope-tagged gK (VZV gK-F). These recombinants grew indistinguishably from native VZV, and FLAG-tagged gK could be readily detected in VZV gK-F-infected cells. FACS analysis established thatmore » gK is transported to the plasma membrane of infected cells, while indirect immunofluorescence demonstrated that gK accumulates predominately in the Golgi. Using VZV gK-F-infected cells we demonstrated that VZV gK, like several other herpesvirus glycoproteins, is efficiently endocytosed from the plasma membrane. However, pulse-labeling experiments revealed that the half-life of gK is considerably shorter than that of other VZV glycoproteins including gB, gE and gH. This finding suggests that gK may be required in lower abundance than other viral glycoproteins during virion morphogenesis or viral entry.« less

Global site-specific analysis of glycoprotein N-glycan processing.

PubMed

Cao, Liwei; Diedrich, Jolene K; Ma, Yuanhui; Wang, Nianshuang; Pauthner, Matthias; Park, Sung-Kyu Robin; Delahunty, Claire M; McLellan, Jason S; Burton, Dennis R; Yates, John R; Paulson, James C

2018-06-01

N-glycans contribute to the folding, stability and functions of the proteins they decorate. They are produced by transfer of the glycan precursor to the sequon Asn-X-Thr/Ser, followed by enzymatic trimming to a high-mannose-type core and sequential addition of monosaccharides to generate complex-type and hybrid glycans. This process, mediated by the concerted action of multiple enzymes, produces a mixture of related glycoforms at each glycosite, making analysis of glycosylation difficult. To address this analytical challenge, we developed a robust semiquantitative mass spectrometry (MS)-based method that determines the degree of glycan occupancy at each glycosite and the proportion of N-glycans processed from high-mannose type to complex type. It is applicable to virtually any glycoprotein, and a complete analysis can be conducted with 30 μg of protein. Here, we provide a detailed description of the method that includes procedures for (i) proteolytic digestion of glycoprotein(s) with specific and nonspecific proteases; (ii) denaturation of proteases by heating; (iii) sequential treatment of the glycopeptide mixture with two endoglycosidases, Endo H and PNGase F, to create unique mass signatures for the three glycosylation states; (iv) LC-MS/MS analysis; and (v) data analysis for identification and quantitation of peptides for the three glycosylation states. Full coverage of site-specific glycosylation of glycoproteins is achieved, with up to thousands of high-confidence spectra hits for each glycosite. The protocol can be performed by an experienced technician or student/postdoc with basic skills for proteomics experiments and takes ∼7 d to complete.

Platelet glycoproteins associated with aspirin-treatment upon platelet activation

PubMed Central

Shah, Punit; Yang, Weiming; Sun, Shisheng; Pasay, Jered; Faraday, Nauder; Zhang, Hui

2017-01-01

Platelet glycoproteins are known to play central roles in hemostasis and vascular integrity and have pathologic roles in vascular occlusive diseases such as myocardial infarction and stroke. Characterizing glycoproteins within and secreted by platelets can provide insight into the mechanisms that underlie vascular pathologies and the therapeutic benefits or failure of anti-platelet agents. To study the impact of aspirin, which is commonly prescribed for primary and secondary cardiovascular prevention, on the platelet glycoproteome, we evaluated washed platelets from ten donors. The platelet glycoproteome, was studied using an iTRAQ in resting and stimulated states and with and without aspirin treatment. Using solid phase extraction of glycosite-containing peptides (SPEG), we were able to identify 799 unique N-linked glycosylation sites (glycosites) in platelets, representing the largest and the most comprehensive analysis to date. We were able to identity a number of glycoproteins impacted by aspirin treatment, which we validated using global proteomics analysis of platelets and their secreted proteins. In our analyses, metallopeptidase inhibitor 1 (TIMP1) was the single most significantly affected glycoprotein by aspirin treatment. ELISA assays confirmed proteomic results and validated our strategy. Functional analysis demonstrated that TIMP1 levels were highly correlated with platelet reactivity in vitro, with a correlation coefficient of −0.5. The release of TIMP1 from platelets, which was previously unknown to be affected by aspirin treatment, may play important roles in hemostasis and/or vascular integrity. If validated, our findings may be useful for developing assays that assess platelet response to aspirin or other anti-platelet therapies. PMID:27452734

Enzymatic cell wall degradation of Chlorella vulgaris and other microalgae for biofuels production.

PubMed

Gerken, Henri G; Donohoe, Bryon; Knoshaug, Eric P

2013-01-01

Cell walls of microalgae consist of a polysaccharide and glycoprotein matrix providing the cells with a formidable defense against its environment. We characterized enzymes that can digest the cell wall and weaken this defense for the purpose of protoplasting or lipid extraction. A growth inhibition screen demonstrated that chitinase, lysozyme, pectinase, sulfatase, β-glucuronidase, and laminarinase had the broadest effect across the various Chlorella strains tested and also inhibited Nannochloropsis and Nannochloris strains. Chlorella is typically most sensitive to chitinases and lysozymes, both enzymes that degrade polymers containing N-acetylglucosamine. Using a fluorescent DNA stain, we developed rapid methodology to quantify changes in permeability in response to enzyme digestion and found that treatment with lysozyme in conjunction with other enzymes has a drastic effect on cell permeability. Transmission electron microscopy of enzymatically treated Chlorella vulgaris indicates that lysozyme degrades the outer surface of the cell wall and removes hair-like fibers protruding from the surface, which differs from the activity of chitinase. This action on the outer surface of the cell causes visible protuberances on the cell surface and presumably leads to the increased settling rate when cells are treated with lysozyme. We demonstrate radical ultrastructural changes to the cell wall in response to treatment with various enzyme combinations which, in some cases, causes a greater than twofold increase in the thickness of the cell wall. The enzymes characterized in this study should prove useful in the engineering and extraction of oils from microalgae.

Sweating the small stuff: Glycoproteins in human sweat and their unexplored potential for microbial adhesion.

PubMed

Peterson, Robyn A; Gueniche, Audrey; Adam de Beaumais, Ségolène; Breton, Lionel; Dalko-Csiba, Maria; Packer, Nicolle H

2016-03-01

There is increasing evidence that secretory fluids such as tears, saliva and milk play an important role in protecting the human body from infection via a washing mechanism involving glycan-mediated adhesion of potential pathogens to secretory glycoproteins. Interaction of sweat with bacteria is well established as the cause of sweat-associated malodor. However, the role of sweat glycoproteins in microbial attachment has received little, if any, research interest in the past. In this review, we demonstrate how recent published studies involving high-throughput proteomic analysis have inadvertently, and fortuitously, exposed an abundance of glycoproteins in sweat, many of which have also been identified in other secretory fluids. We bring together research demonstrating microbial adhesion to these secretory glycoproteins in tears, saliva and milk and suggest a similar role of the sweat glycoproteins in mediating microbial attachment to sweat and/or skin. The contribution of glycan-mediated microbial adhesion to sweat glycoproteins, and the associated impact on sweat derived malodor and pathogenic skin infections are unchartered new research areas that we are beginning to explore. © The Author 2015. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Analytical Pipeline for Discovery and Verification of Glycoproteins from Plasma-Derived Extracellular Vesicles as Breast Cancer Biomarkers.

PubMed

Chen, I-Hsuan; Aguilar, Hillary Andaluz; Paez Paez, J Sebastian; Wu, Xiaofeng; Pan, Li; Wendt, Michael K; Iliuk, Anton B; Zhang, Ying; Tao, W Andy

2018-05-15

Glycoproteins comprise more than half of current FDA-approved protein cancer markers, but the development of new glycoproteins as disease biomarkers has been stagnant. Here we present a pipeline to develop glycoproteins from extracellular vesicles (EVs) through integrating quantitative glycoproteomics with a novel reverse phase glycoprotein array and then apply it to identify novel biomarkers for breast cancer. EV glycoproteomics show promise in circumventing the problems plaguing current serum/plasma glycoproteomics and allowed us to identify hundreds of glycoproteins that have not been identified in blood. We identified 1,453 unique glycopeptides representing 556 glycoproteins in EVs, among which 20 were verified significantly higher in individual breast cancer patients. We further applied a novel glyco-specific reverse phase protein array to quantify a subset of the candidates. Together, this study demonstrates the great potential of this integrated pipeline for biomarker discovery.

Homologous and heterologous antibody responses of mice immunized with purified feline herpesvirus type 1 and canine herpesvirus glycoproteins.

PubMed

Limcumpao, J A; Horimoto, T; Xuan, X N; Tohya, Y; Azetaka, M; Takahashi, E; Mikami, T

1991-06-01

The three glycoproteins each of feline herpesvirus type 1 (FHV-1) and canine herpesvirus (CHV) were purified by affinity chromatography using glycoprotein-specific monoclonal antibodies and used individually or in combination in immunizing mice to determine their relative immunogenicity. All the glycoproteins induced detectable virus neutralizing antibodies to the homologous virus but FHV-1 gp143/108 and its cross-reacting counterpart, CHV gp145/112, elicited the highest titers not only to the homologous virus but to the heterologous virus as well. The production of ELISA antibodies after glycoprotein immunization was variable, while hemagglutination-inhibiting antibodies were produced by only 1 out of 10 FHV-1 gp60-inoculated mice. In general, the antibody titers induced by CHV glycoproteins were lower than those by FHV-1 glycoproteins. These results indicate that these glycoproteins may be useful as subunit vaccines against FHV-1 and CHV infections.

Regulation of Herpes Simplex Virus Glycoprotein-Induced Cascade of Events Governing Cell-Cell Fusion

PubMed Central

Saw, Wan Ting; Eisenberg, Roselyn J.; Cohen, Gary H.

2016-01-01

ABSTRACT Receptor-dependent herpes simplex virus (HSV)-induced cell-cell fusion requires glycoproteins gD, gH/gL, and gB. Our current model posits that during fusion, receptor-activated conformational changes in gD activate gH/gL, which subsequently triggers the transformation of the prefusion form of gB into a fusogenic state. To examine the role of each glycoprotein in receptor-dependent cell-cell fusion, we took advantage of our discovery that fusion by wild-type herpes simplex virus 2 (HSV-2) glycoproteins occurs twice as fast as that achieved by HSV-1 glycoproteins. By sequentially swapping each glycoprotein between the two serotypes, we established that fusion speed was governed by gH/gL, with gH being the main contributor. While the mutant forms of gB fuse at distinct rates that are dictated by their molecular structure, these restrictions can be overcome by gH/gL of HSV-2 (gH2/gL2), thereby enhancing their activity. We also found that deregulated forms of gD of HSV-1 (gD1) and gH2/gL2 can alter the fusogenic potential of gB, promoting cell fusion in the absence of a cellular receptor, and that deregulated forms of gB can drive the fusion machinery to even higher levels. Low pH enhanced fusion by affecting the structure of both gB and gH/gL mutants. Together, our data highlight the complexity of the fusion machinery, the impact of the activation state of each glycoprotein on the fusion process, and the critical role of gH/gL in regulating HSV-induced fusion. IMPORTANCE Cell-cell fusion mediated by HSV glycoproteins requires gD, gH/gL, gB, and a gD receptor. Here, we show that fusion by wild-type HSV-2 glycoproteins occurs twice as fast as that achieved by HSV-1 glycoproteins. By sequentially swapping each glycoprotein between the two serotypes, we found that the fusion process was controlled by gH/gL. Restrictions imposed on the gB structure by mutations could be overcome by gH2/gL2, enhancing the activity of the mutants. Under low-pH conditions or when

Use of lambdagt11 to isolate genes for two pseudorabies virus glycoproteins with homology to herpes simplex virus and varicella-zoster virus glycoproteins

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

Petrovskis, E.A.; Timmins, J.G.; Post, L.E.

1986-10-01

A library of pseudorabies virus (PRV) DNA fragments was constructed in the expression cloning vector lambdagt11. The library was screened with antisera which reacted with mixtures of PRV proteins to isolate recombinant bacteriophages expressing PRV proteins. By the nature of the lambdagt11 vector, the cloned proteins were expressed in Escherichia coli as ..beta..-galactosidase fusion proteins. The fusion proteins from 35 of these phages were purified and injected into mice to raise antisera. The antisera were screened by several different assays, including immunoprecipitation of (/sup 14/C)glucosamine-labeled PRV proteins. This method identified phages expressing three different PRV glycoproteins: the secreted glycoprotein, gX;more » gI; and a glycoprotein that had not been previously identified, which we designate gp63. The gp63 and gI genes map adjacent to each other in the small unique region of the PRV genome. The DNA sequence was determined for the region of the genome encoding gp63 and gI. It was found that gp63 has a region of homology with a herpes simplex virus type 1 (HSV-1) protein, encoded by US7, and also with varicella-zoster virus (VZV) gpIV. The gI protein sequence has a region of homology with HSV-1 gE and VZV gpI. It is concluded that PRV, HSV, and VZV all have a cluster of homologous glycoprotein genes in the small unique components of their genomes and that the organization of these genes is conserved.« less

Bioactivity of proteins isolated from Lactobacillus plantarum L67 treated with Zanthoxylum piperitum DC glycoprotein.

PubMed

Song, S; Oh, S; Lim, K-T

2015-06-01

Lactobacilli in the human gastrointestinal tract have beneficial effects on the health of their host. To enhance these effects, the bioactivity of lactobacilli can be fortified through exogenous dietary or pharmacological agents, such as glycoproteins. To elucidate the inductive effect of Zanthoxylum piperitum DC (ZPDC) glycoprotein on Lactobacillus plantarum L67, we evaluated the radical-scavenging activity, anti-oxidative enzymes (SOD, GPx and CAT), growth rate, ATPase activity and β-galactosidase activity of this strain. When Lact. plantarum L67 was treated with ZPDC glycoprotein at different concentrations, the intensities of a few SDS-PAGE bands were slightly changed. The amount of a 23 kDa protein was increased upon treatment with increasing concentrations of ZPDC glycoprotein. The results of this study indicate that the radical-scavenging activity for O2(-) and OH¯, but not for the DPPH radical, increased in a concentration-dependent manner after treatment with ZPDC glycoprotein. The activation of anti-oxidative enzymes (SOD, GPx and CAT), growth rate and β-galactosidase activity also increased in a concentration-dependent manner in response to ZPDC glycoprotein treatment, whereas ATPase activity was decreased. In summary, ZPDC glycoprotein stimulated an increase in the bioactivity of Lact. plantarum L67. Significance and impact of the study: This study demonstrated that Lactobacillus plantarum L67 possesses anti-oxidative activity. This strain of lactic bacteria has been known to have various probiotic uses, such as yogurt starters and dietary additional supplements. We found, through this experiment, that the protein has a strong anti-oxidative character, and the activity can be enhanced by treatment with Zanthoxylum piperitum DC (ZPDC) glycoprotein. This study may be application of Lact. plantarum L67 treated by ZPDC glycoprotein in yogurt fermentation. It could be one of the avenues of minimizing yogurt postacidification during storage. In addition

Maize Genotype and Food Matrix Affect the Provitamin A Carotenoid Bioefficacy from Staple and Carrot-fortified Feeds in Mongolian Gerbils (Meriones unguiculatus)

PubMed Central

Schmaelzle, Samantha; Gannon, Bryan; Crawford, Serra; Arscott, Sara A.; Goltz, Shellen; Palacios-Rojas, Natalia; Pixley, Kevin V.; Simon, Philipp W.; Tanumihardjo, Sherry A.

2014-01-01

Biofortification to increase provitamin A carotenoids is an agronomic approach to alleviate vitamin A deficiency. Two studies compared biofortified foods using in vitro and in vivo methods. Study 1 screened maize genotypes (n = 44) using in vitro analysis, which demonstrated decreasing micellarization with increasing provitamin A. Thereafter, seven 50% biofortified maize feeds that hypothesized a one-to-one equivalency between β-cryptoxanthin and β-carotene were fed to Mongolian gerbils. Total liver retinol differed among the maize groups (P = 0.0043). Study 2 assessed provitamin A bioefficacy from 0.5% high-carotene carrots added to 60% staple-food feeds, followed by in vitro screening. Liver retinol was highest in the potato and banana groups, maize group retinol did not differ from baseline, and all treatments differed from control (P < 0.0001). In conclusion, β-cryptoxanthin and β-carotene have similar bioefficacy; meal matrix effects influence provitamin A absorption from carrot; and in vitro micellarization does not predict bioefficacy. PMID:24341827

Enrichment of Glycoproteins using Nano-scale Chelating Con A Monolithic Capillary Chromatography

PubMed Central

Feng, Shun; Yang, Na; Pennathur, Subramaniam; Goodison, Steve; Lubman, David M.

2009-01-01

Immobilized lectin chromatography can be employed for glycoprotein enrichment, but commonly used columns have limitations of yield and resolution. In order to improve efficiency and to make the technique applicable to minimal sample material, we have developed a nano-scale chelating Concanavalin A (Con A) monolithic capillary prepared using GMA-EDMA (glycidyl methacrylate–co-ethylene dimethacrylate) as polymeric support. Con A was immobilized on Cu(II)-charged iminodiacetic acid (IDA) regenerable sorbents by forming a IDA:Cu(II):Con A sandwich affinity structure that has high column capacity as well as stability. When compared with conventional Con A lectin chromatography, the monolithic capillary enabled the better reproducible detection of over double the number of unique N-glycoproteins in human urine samples. Utility for analysis of minimal biological samples was confirmed by the successful elucidation of glycoprotein profiles in mouse urine samples at the microliter scale. The improved efficiency of the nano-scale monolithic capillary will impact the analysis of glycoproteins in complex biological samples, especially where only limited material may be available. PMID:19366252

Ubiquitination of exposed glycoproteins by SCFFBXO27 directs damaged lysosomes for autophagy

PubMed Central

Yoshida, Yukiko; Yasuda, Sayaka; Fujita, Toshiharu; Hamasaki, Maho; Murakami, Arisa; Kawawaki, Junko; Iwai, Kazuhiro; Saeki, Yasushi; Yoshimori, Tamotsu; Matsuda, Noriyuki; Tanaka, Keiji

2017-01-01

Ubiquitination functions as a signal to recruit autophagic machinery to damaged organelles and induce their clearance. Here, we report the characterization of FBXO27, a glycoprotein-specific F-box protein that is part of the SCF (SKP1/CUL1/F-box protein) ubiquitin ligase complex, and demonstrate that SCFFBXO27 ubiquitinates glycoproteins in damaged lysosomes to regulate autophagic machinery recruitment. Unlike F-box proteins in other SCF complexes, FBXO27 is subject to N-myristoylation, which localizes it to membranes, allowing it to accumulate rapidly around damaged lysosomes. We also screened for proteins that are ubiquitinated upon lysosomal damage, and identified two SNARE proteins, VAMP3 and VAMP7, and five lysosomal proteins, LAMP1, LAMP2, GNS, PSAP, and TMEM192. Ubiquitination of all glycoproteins identified in this screen increased upon FBXO27 overexpression. We found that the lysosomal protein LAMP2, which is ubiquitinated preferentially on lysosomal damage, enhances autophagic machinery recruitment to damaged lysosomes. Thus, we propose that SCFFBXO27 ubiquitinates glycoproteins exposed upon lysosomal damage to induce lysophagy. PMID:28743755

Characterization of the T-cell response to Dau c 1, the Bet v 1-homolog in carrot.

PubMed

Zulehner, N; Nagl, B; Briza, P; Roulias, A; Ballmer-Weber, B; Zlabinger, G J; Ferreira, F; Bohle, B

2017-02-01

In contrast to other Bet v 1-related food allergens, the major carrot allergen, Dau c 1, has been suggested to induce food allergy independently from Bet v 1. As T cells are crucial in the sensitization process, we sought to characterize the T-cell response to Dau c 1 and its cross-reactivity with Bet v 1. Dau c 1-specific T-cell lines (TCL) and clones (TCC) established from PBMC of birch pollen-allergic patients with carrot allergy were analyzed for reactivity to Bet v 1, epitope specificity, allergen-induced cytokine secretion, and expression of integrins α4β7 and α4β1, critical for gut and lung homing, respectively. mRNA expression of GATA3 and Tbet was analyzed in sorted CD3 + CD4 + CFSE low cells proliferating upon stimulation of PBMC with Dau c 1 or Bet v 1. Dau c 1 was incubated with endolysosomal proteases, and the resulting fragments were identified by mass spectrometry. Among 14 distinct T-cell-activating regions, Dau c 1 139-153 was recognized by 55% of the patients. Only 6 of 15 (40%) Dau c 1-specific TCL and 9 of 21 (43%) TCC reacted with Bet v 1. Bet v 1-nonreactive TCC were mainly Th1-like and showed a higher expression of the integrin β7 and a significantly lower expression of the integrin β1 than Bet v 1-positive TCC. A Th1-like response was also detected in Dau c 1-reactive CD3 + CD4 + CFSE low cells. Full-length Dau c 1 was still detectable after 48 h of endolysosomal degradation. Proteolytic fragments of Dau c 1 matched its T-cell-activating regions. Dau c 1 displays several characteristics of sensitizing allergens, namely a major T-cell-activating region, low susceptibility to endolysosomal degradation, and induction of a Bet v 1-independent T-cell response. These cellular insights confirm that the major carrot allergen has a special status among Bet v 1-related food allergens. © 2016 The Authors. Allergy Published by John Wiley & Sons Ltd.

CHARACTERIZATION OF SULFUR CONTAINING ANALOGS OF MONOMETHYLARSONIC ACID IN AQUEOUS PHASE STANDARDS AND CARROT EXTRACTS BY IC-ICP-MS AND IC-ESI-MS/MS

EPA Science Inventory

Recently, sulfur analogs of well known arsenicals have been identified, generating a need for stable species-specific standards. This presentation will focus on the identification and characterization of a novel species, monomethylthioarsonic acid (MMTA), in carrots. A standard...

The haemagglutination activity of equine herpesvirus type 1 glycoprotein C.

PubMed

Andoh, Kiyohiko; Hattori, Shiho; Mahmoud, Hassan Y A H; Takasugi, Maaya; Shimoda, Hiroshi; Bannai, Hiroshi; Tsujimura, Koji; Matsumura, Tomio; Kondo, Takashi; Kirisawa, Rikio; Mochizuki, Masami; Maeda, Ken

2015-01-02

Equine herpesvirus type 1 (EHV-1) has haemagglutination (HA) activity toward equine red blood cells (RBCs), but the identity of its haemagglutinin is unknown. To identify the haemagglutinin of EHV-1, the major glycoproteins of EHV-1 were expressed in 293T cells, and the cells or cell lysates were mixed with equine RBCs. The results showed that only EHV-1 glycoprotein C (gC)-producing cells adsorbed equine RBCs, and that the lysate of EHV-1 gC-expressing cells agglutinated equine RBCs. EHV-1 lacking gC did not show HA activity. HA activity was inhibited by monoclonal antibodies (MAbs) specific for gC, but not by antibodies directed against other glycoproteins. In addition, HA activity was not inhibited by the addition of heparin. These results indicate that EHV-1 gC can bind equine RBCs irrespective of heparin, in contrast to other herpesvirus gC proteins. Copyright © 2014 Elsevier B.V. All rights reserved.

Stress-Induced Accumulation of DcAOX1 and DcAOX2a Transcripts Coincides with Critical Time Point for Structural Biomass Prediction in Carrot Primary Cultures (Daucus carota L.)

PubMed Central

Campos, M. Doroteia; Nogales, Amaia; Cardoso, Hélia G.; Kumar, Sarma R.; Nobre, Tânia; Sathishkumar, Ramalingam; Arnholdt-Schmitt, Birgit

2016-01-01

Stress-adaptive cell plasticity in target tissues and cells for plant biomass growth is important for yield stability. In vitro systems with reproducible cell plasticity can help to identify relevant metabolic and molecular events during early cell reprogramming. In carrot, regulation of the central root meristem is a critical target for yield-determining secondary growth. Calorespirometry, a tool previously identified as promising for predictive growth phenotyping has been applied to measure the respiration rate in carrot meristem. In a carrot primary culture system (PCS), this tool allowed identifying an early peak related with structural biomass formation during lag phase of growth, around the 4th day of culture. In the present study, we report a dynamic and correlated expression of carrot AOX genes (DcAOX1 and DcAOX2a) during PCS lag phase and during exponential growth. Both genes showed an increase in transcript levels until 36 h after explant inoculation, and a subsequent down-regulation, before the initiation of exponential growth. In PCS growing at two different temperatures (21°C and 28°C), DcAOX1 was also found to be more expressed in the highest temperature. DcAOX genes’ were further explored in a plant pot experiment in response to chilling, which confirmed the early AOX transcript increase prior to the induction of a specific anti-freezing gene. Our findings point to DcAOX1 and DcAOX2a as being reasonable candidates for functional marker development related to early cell reprogramming. While the genomic sequence of DcAOX2a was previously described, we characterize here the complete genomic sequence of DcAOX1. PMID:26858746

The effect of P-glycoprotein on methadone hydrochloride flux in equine intestinal mucosa.

PubMed

Linardi, R L; Stokes, A M; Andrews, F M

2013-02-01

Methadone is an effective analgesic opioid that may have a place for the treatment of pain in horses. However, its absorption seems to be impaired by the presence of a transmembrane protein, P-glycoprotein, present in different tissues including the small intestine in other species. This study aims to determine the effect of the P-glycoprotein on methadone flux in the equine intestinal mucosa, as an indicator of in vivo drug absorption. Jejunum tissues from five horses were placed into the Ussing chambers and exposed to methadone solution in the presence or absence of Rhodamine 123 or verapamil. Electrical measurements demonstrated tissue viability for 120 min, and the flux of methadone across the jejunal membrane (mucosal to submucosal direction) was calculated based on the relative drug concentration measured by ELISA. The flux of methadone was significantly higher only in the presence of verapamil. P-glycoprotein was immunolocalized in the apical membrane of the jejunal epithelial cells (enterocytes), mainly located in the tip of the villi compared to cells of the crypts. P-glycoprotein is present in the equine jejunum and may possibly mediate the intestinal transport of methadone. This study suggests that P-glycoprotein may play a role in the poor intestinal absorption of methadone in vivo. © 2012 Blackwell Publishing Ltd.

HSV-1 Glycoproteins Are Delivered to Virus Assembly Sites Through Dynamin-Dependent Endocytosis.

PubMed

Albecka, Anna; Laine, Romain F; Janssen, Anne F J; Kaminski, Clemens F; Crump, Colin M

2016-01-01

Herpes simplex virus-1 (HSV-1) is a large enveloped DNA virus that belongs to the family of Herpesviridae. It has been recently shown that the cytoplasmic membranes that wrap the newly assembled capsids are endocytic compartments derived from the plasma membrane. Here, we show that dynamin-dependent endocytosis plays a major role in this process. Dominant-negative dynamin and clathrin adaptor AP180 significantly decrease virus production. Moreover, inhibitors targeting dynamin and clathrin lead to a decreased transport of glycoproteins to cytoplasmic capsids, confirming that glycoproteins are delivered to assembly sites via endocytosis. We also show that certain combinations of glycoproteins colocalize with each other and with the components of clathrin-dependent and -independent endocytosis pathways. Importantly, we demonstrate that the uptake of neutralizing antibodies that bind to glycoproteins when they become exposed on the cell surface during virus particle assembly leads to the production of non-infectious HSV-1. Our results demonstrate that transport of viral glycoproteins to the plasma membrane prior to endocytosis is the major route by which these proteins are localized to the cytoplasmic virus assembly compartments. This highlights the importance of endocytosis as a major protein-sorting event during HSV-1 envelopment. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

[Pregnancy-specific beta-glycoprotein in the serum of women with a complicated early pregnancy].

PubMed

Radikov, N

1989-01-01

The author determined pregnancy specific beta 1-glycoprotein in 109 women with threatened early pregnancy as 32 of the women suffered from abortus imminens with several unsuccessful pregnancies in the past as well as 67 women with abortus incipiens with bleeding ex utero. The author established that 87% of women with abortus imminens and preserved pregnancies had values of beta 1-glycoprotein close to those of normal pregnancy for the respective gestational week. 93% of women with abortus incipiens preserved pregnancies till term, but the specific glycoprotein was with in normal ranges. Spontaneous abortion occurred in 7% of women with low values under the 10th percentile. The present study show that examination of pregnancy specific beta 1-glycoprotein in women with threatened early pregnancy is of prognostic significance for the outcome of pregnancy.

Study on the extraction and purification of glycoprotein from the yellow seahorse, Hippocampus kuda Bleeker

PubMed Central

Su, Yuting; Xu, Yongjian

2015-01-01

The optimum parameters of extraction for glycoprotein from seahorse were examined and determined by Box-Behnken combined with ultrasonic extraction technology. Column chromatography of glycoprotein was used for further purification. The optimal extraction conditions of seahorse glycoprotein were extracting time 4.3 h, salt concentration 0.08 mol/L, extracting temperature 73°C, raw material, and water ratio 1:6. At the optimal conditions, the yield of saccharide reached to 1.123%, and the yield of protein reached to 5.898%. For purifying the crude glycoprotein, the stage renounces of DEAE-52 column chromatography were done, respectively, with 0.05, 0.1, 0.5 mol/L NaHCO3 solution, and further purification was done with Sephadex G-100 column chromatography. Finally, two pieces of seahorse glycoprotein were obtained by the column chromatography, that is, HG-11 and HG-21. The saccharide content was 56.7975% and 39.479%, the protein content was 30.5475% and 51.747%, respectively. PMID:26288722

Effect of microgravity environment on cell wall regeneration, cell divisions, growth, and differentiation of plants from protoplasts (7-IML-1)

NASA Technical Reports Server (NTRS)

Rasmussen, Ole

1992-01-01

The primary goal of this project is to investigate if microgravity has any influence on growth and differentiation of protoplasts. Formation of new cell walls on rapeseed protoplasts takes place within the first 24 hours after isolation. Cell division can be observed after 2-4 days and formation of cell aggregates after 5-7 days. Therefore, it is possible during the 7 day IML-1 Mission to investigate if cell wall formation, cell division, and cell differentiation are influenced by microgravity. Protoplasts of rapeseeds and carrot will be prepared shortly before launch and injected into 0.6 ml polyethylene bags. Eight bags are placed in an aluminum block inside the ESA Type 1 container. The containers are placed at 4 C in PTCU's and transferred to orbiter mid-deck. At 4 C all cell processes are slowed down, including cell wall formation. Latest access to the shuttle will be 12 hours before launch. In orbit the containers will be transferred from the PTC box to the 22 C Biorack incubator. The installation of a 1 g centrifuge in Biorack will make it possible to distinguish between effects of near weightlessness and effects caused by cosmic radiation and other space flight factors including vibrations. Parallel control experiments will be carried out on the ground. Other aspects of the experiment are discussed.

Purification and characterization of a soluble glycoprotein from garlic (Allium sativum) and its in vitro bioactivity.

PubMed

Wang, Yan; Zou, Tingting; Xiang, Minghui; Jin, Chenzhong; Zhang, Xuejiao; Chen, Yong; Jiang, Qiuqing; Hu, Yihong

2016-10-02

A soluble glycoprotein was purified to homogeneity from ripe garlic (Allium sativum) bulbs using ammonium sulfate precipitation, Sephadex G-100 gel filtration, and diethylaminoethyl-52 cellulose anion-exchange chromatography. A native mass of 55.7 kDa estimated on gel permeation chromatography and a molecular weight of 13.2 kDa observed on sodium dodecyl sulfate-polyacrylamide gel electrophoresis supported that the glycoprotein is a homotetramer. β-Elimination reaction result suggested that the glycoprotein is an N-linked type. Fourier-transform infrared spectroscopy proved that it contains sugar. Gas chromatography-mass spectrometer analysis showed that its sugar component was galactose. The glycoprotein has 1,1-diphenyl-2-picrylhydrazil free radical scavenging activity and the peroxidation inhibition ability to polyunsaturated fatty acid. These results indicated that the glycoprotein has potential for food additives, functional foods, and even biotechnological and medical applications.

Truncation of the human immunodeficiency virus type 1 transmembrane glycoprotein cytoplasmic domain blocks virus infectivity.

PubMed Central

Dubay, J W; Roberts, S J; Hahn, B H; Hunter, E

1992-01-01

Human immunodeficiency virus type 1 contains a transmembrane glycoprotein with an unusually long cytoplasmic domain. To determine the role of this domain in virus replication, a series of single nucleotide changes that result in the insertion of premature termination codons throughout the cytoplasmic domain has been constructed. These mutations delete from 6 to 192 amino acids from the carboxy terminus of gp41 and do not affect the amino acid sequence of the regulatory proteins encoded by rev and tat. The effects of these mutations on glycoprotein biosynthesis and function as well as on virus infectivity have been examined in the context of a glycoprotein expression vector and the viral genome. All of the mutant glycoproteins were synthesized, processed, and transported to the cell surface in a manner similar to that of the wild-type glycoprotein. With the exception of mutants that remove the membrane anchor domain, all of the mutant glycoproteins retained the ability to cause fusion of CD4-bearing cells. However, deletion of more than 19 amino acids from the C terminus of gp41 blocked the ability of mutant virions to infect cells. This defect in virus infectivity appeared to be due at least in part to a failure of the virus to efficiently incorporate the truncated glycoprotein. Similar data were obtained for mutations in two different env genes and two different target cell lines. These results indicate that the cytoplasmic domain of gp41 plays a critical role during virus assembly and entry in the life cycle of human immunodeficiency virus type 1. Images PMID:1357190

Glycoproteomic analysis of bronchoalveolar lavage (BAL) fluid identifies tumor-associated glycoproteins from lung adenocarcinoma.

PubMed

Li, Qing Kay; Shah, Punit; Li, Yan; Aiyetan, Paul O; Chen, Jing; Yung, Rex; Molena, Daniela; Gabrielson, Edward; Askin, Frederic; Chan, Daniel W; Zhang, Hui

2013-08-02

Cytological examination of cells from bronchoalveolar lavage (BAL) is commonly used for the diagnosis of lung cancer. Proteins released from lung cancer cells into BAL may serve as biomarkers for cancer detection. In this study, N-glycoproteins in eight cases of BAL fluid, as well as eight lung adenocarcinoma tissues and eight tumor-matched normal lung tissues, were analyzed using the solid-phase extraction of N-glycoprotein (SPEG), iTRAQ labeling, and liquid chromatography tandem mass spectrometry (LC-MS/MS). Of 80 glycoproteins found in BAL specimens, 32 were identified in both cancer BAL and cancer tissues, with levels of 25 glycoproteins showing at least a 2-fold difference between cancer and benign BAL. Among them, eight glycoproteins showed greater than 2-fold elevations in cancer BAL, including Neutrophil elastase (NE), Integrin alpha-M, Cullin-4B, Napsin A, lysosome-associated membrane protein 2 (LAMP2), Cathepsin D, BPI fold-containing family B member 2, and Neutrophil gelatinase-associated lipocalin. The levels of Napsin A in cancer BAL were further verified in independently collected 39 BAL specimens using an ELISA assay. Our study demonstrates that potential protein biomarkers in BAL fluid can be detected and quantified.

Presynaptic neurones may contribute a unique glycoprotein to the extracellular matrix at the synapse

NASA Astrophysics Data System (ADS)

Caroni, Pico; Carlson, Steven S.; Schweitzer, Erik; Kelly, Regis B.

1985-04-01

As the extracellular matrix at the original site of a neuromuscular junction seems to play a major part in the specificity of synaptic regeneration1-5, considerable attention has been paid to unique molecules localized to this region6-11. Here we describe an extracellular matrix glycoprotein of the elasmobranch electric organ that is localized near the nerve endings. By immunological criteria, it is synthesized in the cell bodies, transported down the axons and is related to a glycoprotein in the synaptic vesicles of the neurones that innervate the electric organ. It is apparently specific for these neurones, as it cannot be detected elsewhere in the nervous system of the fish. Therefore, neurones seem to contribute unique extracellular matrix glycoproteins to the synaptic region. Synaptic vesicles could be involved in transporting these glycoproteins to or from the nerve terminal surface.

Chemoenzymatic Site-Specific Labeling of Influenza Glycoproteins as a Tool to Observe Virus Budding in Real Time

PubMed Central

Ploegh, Hidde L.

2012-01-01

The influenza virus uses the hemagglutinin (HA) and neuraminidase (NA) glycoproteins to interact with and infect host cells. While biochemical and microscopic methods allow examination of the early steps in flu infection, the genesis of progeny virions has been more difficult to follow, mainly because of difficulties inherent in fluorescent labeling of flu proteins in a manner compatible with live cell imaging. We here apply sortagging as a chemoenzymatic approach to label genetically modified but infectious flu and track the flu glycoproteins during the course of infection. This method cleanly distinguishes influenza glycoproteins from host glycoproteins and so can be used to assess the behavior of HA or NA biochemically and to observe the flu glycoproteins directly by live cell imaging. PMID:22457626

Mannostatin A, a new glycoprotein-processing inhibitor.

PubMed

Tropea, J E; Kaushal, G P; Pastuszak, I; Mitchell, M; Aoyagi, T; Molyneux, R J; Elbein, A D

1990-10-30

Mannostatin A is a metabolite produced by the microorganism Streptoverticillium verticillus and reported to be a potent competitive inhibitor of rat epididymal alpha-mannosidase. When tested against a number of other arylglycosidases, mannostatin A was inactive toward alpha- and beta-glucosidase and galactosidase as well as beta-mannosidase, but it was a potent inhibitor of jack bean, mung bean, and rat liver lysosomal alpha-mannosidases, with estimated IC50's of 70 nM, 450 nM, and 160 nM, respectively. The type of inhibition was competitive in nature. This compound also proved to be an effective competitive inhibitor of the glycoprotein-processing enzyme mannosidase II (IC50 of about 10-15 nM with p-nitrophenyl alpha-D-mannopyranoside as substrate, and about 90 nM with [3H]mannose-labeled GlcNAc-Man5GlcNAc as substrate). However, it was virtually inactive toward mannosidase I. The N-acetylated derivative of mannostatin A had no inhibitory activity. In cell culture studies, mannostatin A also proved to be a potent inhibitor of glycoprotein processing. Thus, in influenza virus infected Madin Darby canine kidney (MDCK) cells, mannostatin A blocked the normal formation of complex types of oligosaccharides on the viral glycoproteins and caused the accumulation of hybrid types of oligosaccharides. This observation is in keeping with other data which indicate that the site of action of mannostatin A is mannosidase II. Thus, mannostatin A represents the first nonalkaloidal processing inhibitor and adds to the growing list of chemical structures that can have important biological activity.

Platelet Glycoprotein Ib-IX and Malignancy

DTIC Science & Technology

2010-09-01

provide a unique microenvironment supporting the accumulation of more platelets and the elaboration of a fibrin - rich network produced by coagulation...process and can initiate the formation of a platelet - rich thrombus by tethering the platelet to a thrombogenic surface. Several ligands binding to GP Ib... Platelet Glycoprotein Ib-IX and Malignancy PRINCIPAL INVESTIGATOR: Jerry Ware, Ph.D

Antibodies to the Glycoprotein GP2 Subunit Cross-React between Old and New World Arenaviruses.

PubMed

Amanat, Fatima; Duehr, James; Oestereich, Lisa; Hastie, Kathryn M; Ollmann Saphire, Erica; Krammer, Florian

2018-01-01

Arenaviruses pose a major public health threat and cause numerous infections in humans each year. Although most viruses belonging to this family do not cause disease in humans, some arenaviruses, such as Lassa virus and Machupo virus, are the etiological agents of lethal hemorrhagic fevers. The absence of a currently licensed vaccine and the highly pathogenic nature of these viruses both make the necessity of developing viable vaccines and therapeutics all the more urgent. Arenaviruses have a single glycoprotein on the surface of virions, the glycoprotein complex (GPC), and this protein can be used as a target for vaccine development. Here, we describe immunization strategies to generate monoclonal antibodies (MAbs) that cross-react between the glycoprotein complexes of both Old World and New World arenaviruses. Several monoclonal antibodies isolated from immunized mice were highly cross-reactive, binding a range of Old World arenavirus glycoproteins, including that of Lassa virus. One such monoclonal antibody, KL-AV-2A1, bound to GPCs of both New World and Old World viruses, including Lassa and Machupo viruses. These cross-reactive antibodies bound to epitopes present on the glycoprotein 2 subunit of the glycoprotein complex, which is relatively conserved among arenaviruses. Monoclonal antibodies binding to these epitopes, however, did not inhibit viral entry as they failed to neutralize a replication-competent vesicular stomatitis virus pseudotyped with the Lassa virus glycoprotein complex in vitro In addition, no protection from virus challenge was observed in in vivo mouse models. Even so, these monoclonal antibodies might still prove to be useful in the development of clinical and diagnostic assays. IMPORTANCE Several viruses in the Arenaviridae family infect humans and cause severe hemorrhagic fevers which lead to high case fatality rates. Due to their pathogenicity and geographic tropisms, these viruses remain very understudied. As a result, an effective

Apple, carrot, and hibiscus edible films containing the plant antimicrobials carvacrol and cinnamaldehyde inactivate Salmonella Newport on organic leafy greens in sealed plastic bags

USDA-ARS?s Scientific Manuscript database

The objective of this study was to investigate the antimicrobial effects of carvacrol and cinnamaldehyde incorporated into apple, carrot and hibiscus based edible films against Salmonella Newport in contaminated organic leafy greens. The leafy greens tested included romaine and iceberg lettuce, and ...

Glycoprotein-Based Enzyme-Linked Immunosorbent Assays for Serodiagnosis of Infectious Laryngotracheitis

PubMed Central

Kanabagatte Basavarajappa, Mallikarjuna; Song, Haichen; Lamichhane, Chinta

2015-01-01

For detection of infectious laryngotracheitis virus (ILTV) antibody, glycoprotein B-, C-, and D-based enzyme-linked immunosorbent assays (B-, C-, and D-ELISAs, respectively) were developed. The B- and D-ELISAs showed enhanced detection of anti-ILTV antibodies in infected chickens compared to that of the commercial ELISA. Furthermore, the D-ELISA was efficient in detecting seroconversion with vectored vaccine, using recombinant Newcastle disease virus (rNDV) expressing glycoprotein D (gD) as the vaccine vector. PMID:25694519

A Multifaceted Study of Scedosporium boydii Cell Wall Changes during Germination and Identification of GPI-Anchored Proteins

PubMed Central

Ghamrawi, Sarah; Gastebois, Amandine; Zykwinska, Agata; Vandeputte, Patrick; Marot, Agnès; Mabilleau, Guillaume; Cuenot, Stéphane; Bouchara, Jean-Philippe

2015-01-01

Scedosporium boydii is a pathogenic filamentous fungus that causes a wide range of human infections, notably respiratory infections in patients with cystic fibrosis. The development of new therapeutic strategies targeting S. boydii necessitates a better understanding of the physiology of this fungus and the identification of new molecular targets. In this work, we studied the conidium-to-germ tube transition using a variety of techniques including scanning and transmission electron microscopy, atomic force microscopy, two-phase partitioning, microelectrophoresis and cationized ferritin labeling, chemical force spectroscopy, lectin labeling, and nanoLC-MS/MS for cell wall GPI-anchored protein analysis. We demonstrated that the cell wall undergoes structural changes with germination accompanied with a lower hydrophobicity, electrostatic charge and binding capacity to cationized ferritin. Changes during germination also included a higher accessibility of some cell wall polysaccharides to lectins and less CH3/CH3 interactions (hydrophobic adhesion forces mainly due to glycoproteins). We also extracted and identified 20 GPI-anchored proteins from the cell wall of S. boydii, among which one was detected only in the conidial wall extract and 12 only in the mycelial wall extract. The identified sequences belonged to protein families involved in virulence in other fungi like Gelp/Gasp, Crhp, Bglp/Bgtp families and a superoxide dismutase. These results highlighted the cell wall remodeling during germination in S. boydii with the identification of a substantial number of cell wall GPI-anchored conidial or hyphal specific proteins, which provides a basis to investigate the role of these molecules in the host-pathogen interaction and fungal virulence. PMID:26038837

A Multifaceted Study of Scedosporium boydii Cell Wall Changes during Germination and Identification of GPI-Anchored Proteins.

PubMed

Ghamrawi, Sarah; Gastebois, Amandine; Zykwinska, Agata; Vandeputte, Patrick; Marot, Agnès; Mabilleau, Guillaume; Cuenot, Stéphane; Bouchara, Jean-Philippe

2015-01-01

Scedosporium boydii is a pathogenic filamentous fungus that causes a wide range of human infections, notably respiratory infections in patients with cystic fibrosis. The development of new therapeutic strategies targeting S. boydii necessitates a better understanding of the physiology of this fungus and the identification of new molecular targets. In this work, we studied the conidium-to-germ tube transition using a variety of techniques including scanning and transmission electron microscopy, atomic force microscopy, two-phase partitioning, microelectrophoresis and cationized ferritin labeling, chemical force spectroscopy, lectin labeling, and nanoLC-MS/MS for cell wall GPI-anchored protein analysis. We demonstrated that the cell wall undergoes structural changes with germination accompanied with a lower hydrophobicity, electrostatic charge and binding capacity to cationized ferritin. Changes during germination also included a higher accessibility of some cell wall polysaccharides to lectins and less CH3/CH3 interactions (hydrophobic adhesion forces mainly due to glycoproteins). We also extracted and identified 20 GPI-anchored proteins from the cell wall of S. boydii, among which one was detected only in the conidial wall extract and 12 only in the mycelial wall extract. The identified sequences belonged to protein families involved in virulence in other fungi like Gelp/Gasp, Crhp, Bglp/Bgtp families and a superoxide dismutase. These results highlighted the cell wall remodeling during germination in S. boydii with the identification of a substantial number of cell wall GPI-anchored conidial or hyphal specific proteins, which provides a basis to investigate the role of these molecules in the host-pathogen interaction and fungal virulence.

Application of visible/near infrared spectroscopy to quality control of fresh fruits and vegetables in large-scale mass distribution channels: a preliminary test on carrots and tomatoes.

PubMed

Beghi, Roberto; Giovenzana, Valentina; Tugnolo, Alessio; Guidetti, Riccardo

2018-05-01

The market for fruits and vegetables is mainly controlled by the mass distribution channel (MDC). MDC buyers do not have useful instruments to rapidly evaluate the quality of the products. Decisions by the buyers are driven primarily by pricing strategies rather than product quality. Simple, rapid and easy-to-use methods for objectively evaluating the quality of postharvest products are needed. The present study aimed to use visible and near-infrared (vis/NIR) spectroscopy to estimate some qualitative parameters of two low-price products (carrots and tomatoes) of various brands, as well as evaluate the applicability of this technique for use in stores. A non-destructive optical system (vis/NIR spectrophotometer with a reflection probe, spectral range 450-1650 nm) was tested. The differences in quality among carrots and tomatoes purchased from 13 stores on various dates were examined. The reference quality parameters (firmness, water content, soluble solids content, pH and colour) were correlated with the spectral readings. The models derived from the optical data gave positive results, in particular for the prediction of the soluble solids content and the colour, with better results for tomatoes than for carrots. The application of optical techniques may help MDC buyers to monitor the quality of postharvest products, leading to an effective optimization of the entire supply chain. © 2017 Society of Chemical Industry. © 2017 Society of Chemical Industry.

Xylose-rich polysaccharides from the primary walls of embryogenic cell line of Pinus caribaea.

PubMed

Mollard, A; Domon, J M; David, H; Joseleau, J P

1997-08-01

Embryogenic cell lines of Pinus caribaea were isolated from somatic embryogenesis from zygotic embryos. Previous studies showed that the proteins and glycoproteins were characteristic of the embryogenic state. In the present work we were seeking typical feature in the polysaccharide from the cell walls of embryogenic calli at nine days of culture. Sequential extraction with water, ammonium oxalate, dimethyl sulfoxide, sodium borohydride and 4.3 M potassium hydroxide revealed that the extracted polysaccharides contained high proportions of arabinose and significant amounts of xylose. Fractionation of the hydrosoluble polymers on DEAE cellulose afforded a xylose-rich fraction (80% xylose, 24% glucose and lower properties of fucose and mannose). Methylation analysis and 13C-NMR spectra showed that the glycan backbone consisted of beta 1 --> 4 linked xylosyl residues Similar study of the fractions extracted respectively with DMSO and 4.3 M KOH showed the presence of polydisperse glycoxylans but excluded the presence of xyloglucan in significant amount. This could be a characteristic feature of embryogenic cells walls of Pinus caribaea or could be typical of cells grown as calluses. In the various fractions obtained from DEAE cellulose chromatography of the alkaline extract the infrequent occurrence of fucoxylans beside an arabinogalactan showed again the unusual nature of the cell wall polymers of this embryogenic lines, which seems to differ greatly from those found in the primary wall of cells from suspension cultures.

Identification of the glycoproteins of lymphocystis disease virus (LDV) of fish.

PubMed

Robin, J; Laperrière, A; Berthiaume, L

1986-01-01

Analysis of highly purified fish Lymphocystis Disease Virus (LDV), strain Leetown NFH, by three different methods, namely periodic Acid Schiff reaction, radiolabelling with tritiated fucose and N-acetyl-D-glucosamine and staining with three lectins, indicated that ten glycoproteins were associated with the virus structure. Six of them were detected by all of the three methods, three by both radiolabelling and lectin staining but only one by the lectin technique. Localization of these glycoproteins at the surface or inside the virion is discussed.

Ineffective hand washing and the contamination of carrots after using a field latrine.

PubMed

Monaghan, J M; Hutchison, M L

2016-04-01

A study was undertaken to simulate the likely effects of a field worker with poor hygienic practices that had returned to work too soon after recovering from an infection by an enteric pathogen. The studies simulated a variety of hand-washing practices from no washing to washing with soap and water followed by an application of alcohol gel after using a field latrine. The numbers of generic Escherichia coli isolated from workers' hands declined with increasing thoroughness of hand-washing treatments with unwashed hands > water > water and soap > water, soap and alcohol gel. Where gloves were worn the counts obtained for the treatments were significantly reduced, but it was observed that unwashed hands contaminated gloves during the process of putting them on. Hand contamination following the use of a field latrine transferred contamination to carrots. These results suggest that if no gloves are worn it would be best practice to wash hands with water and soap and apply alcohol gel after using a field latrine. Wearing gloves reduced the risk of contaminating handled produce but workers should still wash their hands after using a field latrine before applying gloves. This study shows that inadequate hand hygiene in the field following the use of a field latrine can transfer bacterial contamination to hand-harvested carrots. Where fresh produce crops are to be handled by workers, wearing gloves reduces the risk of contaminating produce but workers should still wash their hands after using a field latrine before applying gloves. If no gloves are worn it would be best practice to wash hands with water and soap and apply alcohol gel after using a field latrine. © 2016 The Society for Applied Microbiology.

Prostate Cancer Progression and Serum SIBLING (Small Integrin Binding N-Linked Glycoprotein)Levels

DTIC Science & Technology

2007-10-01

termed SIBLINGs (for small integrin binding ligand N-linked glycoproteins) whose members include bone sialoprotein (BSP), osteopontin (OPN), dentin...enzyme-linked immunosorbent assays (ELISAs) for quantitatively determining the levels of bone sialoprotein (BSP), osteopontin (OPN), dentin...synthesized as a chimeric protein, composed of three parts: dentin sialoprotein (DSP), dentin glycoprotein (DGP) and dentin phosphoprotein (DPP, also

The inhibitory and combinative mechanism of HZ08 with P-glycoprotein expressed on the membrane of Caco-2 cell line

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

Zhang, Yanyan; Hu, Yahui; Feng, Yidong

2014-01-15

Recently, the research and development of agents to reverse the phenomenon of multidrug resistance has been an attractive goal as well as a key approach to elevating the clinical survival of cancer patients. Although three generations of P-glycoprotein modulators have been identified, poor clearance and metabolism render these agents too toxic to be used in clinical application. HZ08, which has been under investigation for several years, shows a dramatic reversal effect with low cytotoxicity. For the first time, we aimed to describe the interaction between HZ08 and P-glycoprotein in Caco-2 cell line in which P-glycoprotein is overexpressed naturally. Cytotoxicity andmore » multidrug resistance reversal assays, together with flow cytometry, fluorescence microscopy and siRNA interference as well as Caco-2 monolayer transport model were employed in this study to evaluate the interaction between HZ08 and P-glycoprotein. This study revealed that HZ08 was capable of reversing adriamycin resistance mediated by P-glycoprotein as a result of intracellular enhancement of adriamycin accumulation, which was found to be superior to verapamil. In addition, we confirmed that HZ08 suppressed the transport of Rhodamine123 in the Caco-2 monolayer model but had little effect on P-glycoprotein expression. The transport of HZ08 was diminished by P-glycoprotein inhibitors (verapamil and LY335979) and its accumulation was increased via siRNA targeting MDR1 in Caco-2 cells. Furthermore, considering the binding site of P-glycoprotein, verapamil performed as a competitive inhibitor with HZ08. In conclusion, as a P-glycoprotein substrate, HZ08 inhibited P-glycoprotein activity and may share the same binding site of verapamil to P-glycoprotein. - Highlights: • The cytotoxicity and reversing effect of HZ08 was measured in Caco-2 cell line. • HZ08 inhibited the transport of Rhodamine123 across Caco-2 cell monolayer. • The efflux ratio of HZ08 was dropped when combined with P-glycoprotein

Urinary Alpha-1-Acid Glycoprotein Is a Sensitive Marker of Glomerular Protein Leakage at Altitude.

PubMed

Talks, Ben J; Bradwell, Susie B; Delamere, John; Rayner, Will; Clarke, Alex; Lewis, Chris T; Thomas, Owen D; Bradwell, Arthur R

2018-06-11

Talks, Ben J., Susie B. Bradwell, John Delamere, Will Rayner, Alex Clarke, Chris T. Lewis, Owen D. Thomas, and Arthur R. Bradwell. Urinary alpha-1-acid glycoprotein is a sensitive marker of glomerular protein leakage at altitude. High Alt Med Biol 00:000-000, 2018. Proteinuria is an established feature of ascent to altitude and may be caused by a loss of negative charges on glomerular capillary walls (GCWs). To test this hypothesis, we measured two similar sized but oppositely charged proteins in urine: negatively charged alpha-1-acid glycoprotein (α1-AGP, 41-43 kDa) and positively charged dimeric lambda free light chains (λ-FLCs, 50 kDa). Twenty-four-hour urinary leakage was compared with albumin, a 66 kDa negatively charged protein. We studied 23 individuals (ages 23-78 years, male = 17) at baseline (140 m) and daily during an expedition to 5035 m. The results showed a significant increase in median urinary leakage of α1-AGP (p < 0.0001; 6.85-fold) and albumin (p = 0.0006; 1.65-fold) with ascent to altitude, but no significant increase in leakage of λ-FLCs (p = 0.39; 1.14-fold). α1-AGP correlated with the daily ascent profile (p = 0.0026) and partial pressure of oxygen (p = 0.01), whereas albumin showed no correlation (p = 0.19). Urinary α1-AGP was a more sensitive marker of altitude proteinuria than urinary albumin and λ-FLCs, and supported the possibility of loss of GCW negative charges at altitude.

Arabidopsis leucine-rich repeat extensin (LRX) proteins modify cell wall composition and influence plant growth.

PubMed

Draeger, Christian; Ndinyanka Fabrice, Tohnyui; Gineau, Emilie; Mouille, Grégory; Kuhn, Benjamin M; Moller, Isabel; Abdou, Marie-Therese; Frey, Beat; Pauly, Markus; Bacic, Antony; Ringli, Christoph

2015-06-24

Leucine-rich repeat extensins (LRXs) are extracellular proteins consisting of an N-terminal leucine-rich repeat (LRR) domain and a C-terminal extensin domain containing the typical features of this class of structural hydroxyproline-rich glycoproteins (HRGPs). The LRR domain is likely to bind an interaction partner, whereas the extensin domain has an anchoring function to insolubilize the protein in the cell wall. Based on the analysis of the root hair-expressed LRX1 and LRX2 of Arabidopsis thaliana, LRX proteins are important for cell wall development. The importance of LRX proteins in non-root hair cells and on the structural changes induced by mutations in LRX genes remains elusive. The LRX gene family of Arabidopsis consists of eleven members, of which LRX3, LRX4, and LRX5 are expressed in aerial organs, such as leaves and stem. The importance of these LRX genes for plant development and particularly cell wall formation was investigated. Synergistic effects of mutations with gradually more severe growth retardation phenotypes in double and triple mutants suggest a similar function of the three genes. Analysis of cell wall composition revealed a number of changes to cell wall polysaccharides in the mutants. LRX3, LRX4, and LRX5, and most likely LRX proteins in general, are important for cell wall development. Due to the complexity of changes in cell wall structures in the lrx mutants, the exact function of LRX proteins remains to be determined. The increasingly strong growth-defect phenotypes in double and triple mutants suggests that the LRX proteins have similar functions and that they are important for proper plant development.

Global analysis of glycoproteins identifies markers of endotoxin tolerant monocytes and GPR84 as a modulator of TNFα expression.

PubMed

Müller, Mario M; Lehmann, Roland; Klassert, Tilman E; Reifenstein, Stella; Conrad, Theresia; Moore, Christoph; Kuhn, Anna; Behnert, Andrea; Guthke, Reinhard; Driesch, Dominik; Slevogt, Hortense

2017-04-12

Exposure of human monocytes to lipopolysaccharide (LPS) induces a temporary insensitivity to subsequent LPS challenges, a cellular state called endotoxin tolerance. In this study, we investigated the LPS-induced global glycoprotein expression changes of tolerant human monocytes and THP-1 cells to identify markers and glycoprotein targets capable to modulate the immunosuppressive state. Using hydrazide chemistry and LC-MS/MS analysis, we analyzed glycoprotein expression changes during a 48 h LPS time course. The cellular snapshots at different time points identified 1491 glycoproteins expressed by monocytes and THP-1 cells. Label-free quantitative analysis revealed transient or long-lasting LPS-induced expression changes of secreted or membrane-anchored glycoproteins derived from intracellular membrane coated organelles or from the plasma membrane. Monocytes and THP-1 cells demonstrated marked differences in glycoproteins differentially expressed in the tolerant state. Among the shared differentially expressed glycoproteins G protein-coupled receptor 84 (GPR84) was identified as being capable of modulating pro-inflammatory TNFα mRNA expression in the tolerant cell state when activated with its ligand Decanoic acid.

Polyphenolic profile and biological activities of black carrot crude extract (Daucus carota L. ssp. sativus var. atrorubens Alef.).

PubMed

Smeriglio, A; Denaro, M; Barreca, D; D'Angelo, V; Germanò, M P; Trombetta, D

2018-01-01

Black carrot (Daucus carota L. ssp. sativus var. atrorubens Alef.) is a valuable source of carbohydrates, minerals and vitamins and contains also high amounts of anthocyanins giving the characteristic deep-purple color. These latter compounds are known as natural dyes used in the food and pharmaceutical industry that have recently attracted much attention for their healthful properties. The aim of this work was to investigate for the first time the polyphenolic profile and biological properties of a black carrot crude extract (BCCE) through an in-depth analysis of the main polyphenolic classes evaluating its antioxidant, cytoprotective and anti-angiogenic properties. Twenty five polyphenols were quantified by LC-DAD-FLD-MS/MS analysis (anthocyanins 78.06%, phenolic acids 17.89% and other flavonoids 4.06%) with polyglycosylated cyanidins as major components. In addition, BCCE showed a strong antioxidant and free radical scavenging activity particularly in the hydrogen transfer-based assays (ORAC and β-carotene bleaching) and a significant increase in the cell viability. Furthermore, BCCE exhibited a strong anti-angiogenic activity at the highest concentration assayed on the chick chorioallantoic membrane (50μg/egg). In conclusion, the obtained results demonstrated the antioxidant, cytoprotective and anti-angiogenic properties of BCCE, which highlight that the higher biological activity of BCCE is probably due to the synergic effects exerted by various polyphenolic classes. Copyright © 2017 Elsevier B.V. All rights reserved.

Glycoprotein-based enzyme-linked immunosorbent assays for serodiagnosis of infectious laryngotracheitis.

PubMed

Kanabagatte Basavarajappa, Mallikarjuna; Song, Haichen; Lamichhane, Chinta; Samal, Siba K

2015-05-01

For detection of infectious laryngotracheitis virus (ILTV) antibody, glycoprotein B-, C-, and D-based enzyme-linked immunosorbent assays (B-, C-, and D-ELISAs, respectively) were developed. The B- and D-ELISAs showed enhanced detection of anti-ILTV antibodies in infected chickens compared to that of the commercial ELISA. Furthermore, the D-ELISA was efficient in detecting seroconversion with vectored vaccine, using recombinant Newcastle disease virus (rNDV) expressing glycoprotein D (gD) as the vaccine vector. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

A multivariate approach using attenuated total reflectance mid-infrared spectroscopy to measure the surface mannoproteins and β-glucans of yeast cell walls during wine fermentations.

PubMed

Moore, John P; Zhang, Song-Lei; Nieuwoudt, Hélène; Divol, Benoit; Trygg, Johan; Bauer, Florian F

2015-11-18

Yeast cells possess a cell wall comprising primarily glycoproteins, mannans, and glucan polymers. Several yeast phenotypes relevant for fermentation, wine processing, and wine quality are correlated with cell wall properties. To investigate the effect of wine fermentation on cell wall composition, a study was performed using mid-infrared (MIR) spectroscopy coupled with multivariate methods (i.e., PCA and OPLS-DA). A total of 40 yeast strains were evaluated, including Saccharomyces strains (laboratory and industrial) and non-Saccharomyces species. Cells were fermented in both synthetic MS300 and Chardonnay grape must to stationery phase, processed, and scanned in the MIR spectrum. PCA of the fingerprint spectral region showed distinct separation of Saccharomyces strains from non-Saccharomyces species; furthermore, industrial wine yeast strains separated from laboratory strains. PCA loading plots and the use of OPLS-DA to the data sets suggested that industrial strains were enriched with cell wall proteins (e.g., mannoproteins), whereas laboratory strains were composed mainly of mannan and glucan polymers.

Possible mechanisms of pollination failure in hybrid carrot seed and implications for industry in a changing climate.

PubMed

Broussard, Melissa Ann; Mas, Flore; Howlett, Brad; Pattemore, David; Tylianakis, Jason M

2017-01-01

Approximately one-third of our food globally comes from insect-pollinated crops. The dependence on pollinators has been linked to yield instability, which could potentially become worse in a changing climate. Insect-pollinated crops produced via hybrid breeding (20% of fruit and vegetable production globally) are especially at risk as they are even more reliant on pollinators than open-pollinated plants. We already observe a wide range of fruit and seed yields between different cultivars of the same crop species, and it is unknown how existing variation will be affected in a changing climate. In this study, we examined how three hybrid carrot varieties with differential performance in the field responded to three temperature regimes (cooler than the historical average, average, and warmer that the historical average). We tested how temperature affected the plants' ability to set seed (seed set, pollen viability) as well as attract pollinators (nectar composition, floral volatiles). We found that there were significant intrinsic differences in nectar phenolics, pollen viability, and seed set between the carrot varieties, and that higher temperatures did not exaggerate those differences. However, elevated temperature did negatively affect several characteristics relating to the attraction and reward of pollinators (lower volatile production and higher nectar sugar concentration) across all varieties, which may decrease the attractiveness of this already pollinator-limited crop. Given existing predictions of lower pollinator populations in a warmer climate, reduced attractiveness would add yet another challenge to future food production.

Possible mechanisms of pollination failure in hybrid carrot seed and implications for industry in a changing climate

PubMed Central

Mas, Flore; Howlett, Brad; Pattemore, David; Tylianakis, Jason M.

2017-01-01

Approximately one-third of our food globally comes from insect-pollinated crops. The dependence on pollinators has been linked to yield instability, which could potentially become worse in a changing climate. Insect-pollinated crops produced via hybrid breeding (20% of fruit and vegetable production globally) are especially at risk as they are even more reliant on pollinators than open-pollinated plants. We already observe a wide range of fruit and seed yields between different cultivars of the same crop species, and it is unknown how existing variation will be affected in a changing climate. In this study, we examined how three hybrid carrot varieties with differential performance in the field responded to three temperature regimes (cooler than the historical average, average, and warmer that the historical average). We tested how temperature affected the plants' ability to set seed (seed set, pollen viability) as well as attract pollinators (nectar composition, floral volatiles). We found that there were significant intrinsic differences in nectar phenolics, pollen viability, and seed set between the carrot varieties, and that higher temperatures did not exaggerate those differences. However, elevated temperature did negatively affect several characteristics relating to the attraction and reward of pollinators (lower volatile production and higher nectar sugar concentration) across all varieties, which may decrease the attractiveness of this already pollinator-limited crop. Given existing predictions of lower pollinator populations in a warmer climate, reduced attractiveness would add yet another challenge to future food production. PMID:28665949

A membrane glycoprotein that accumulates intracellularly: cellular processing of the large glycoprotein of LaCrosse virus.

PubMed

Madoff, D H; Lenard, J

1982-04-01

The intracellular transport and certain posttranslational modifications of the large glycoprotein (G1) of LaCrosse virus (LAC) in BHK cells have been studied. G1 from released LAC virus was characterized by complex oligosaccharides (endo H-resistant) and covalently attached fatty acid. Only a small fraction of total cellular G1 was present on the baby hamster kidney cell surface. Cell-surface G1 contained complex oligosaccharides, while total G1 in infected cells contained largely unprocessed (endo H-sensitive) oligosaccharides. In addition, cell G1 contained significantly less fatty acid than virion-associated G1. Pulse-chase experiments showed that the oligosaccharides of G1 were processed to the complex from much more slowly than the oligosaccharides of the vesicular stomatitis virus (VSV) glycoprotein (G). In addition, transit of LAC G1 to the cell surface and into extracellular virions was two to three fold slower than the transit of VSV G. Thus LAC G1 accumulates intracellularly and is only slowly processed by intracellular processing enzymes. Treatment with monensin caused accumulation in the cell of a form of G1 with partial sensitivity toward endo H, suggesting that monensin may act to inhibit the glycosylation process directly.

A double responsive smart upconversion fluorescence sensing material for glycoprotein.

PubMed

Guo, Ting; Deng, Qiliang; Fang, Guozhen; Yun, Yaguang; Hu, Yongjin; Wang, Shuo

2016-11-15

A novel strategy was developed to prepare double responsive smart upconversion fluorescence material for highly specific enrichment and sensing of glycoprotein. The novel double responsive smart sensing material was synthesized by choosing Horse radish peroxidase (HRP) as modal protein, the grapheme oxide (GO) as support material, upconversion nanoparticles (UCNPs) as fluorescence signal reporter, N-isopropyl acrylamide (NIPAAM) and 4-vinylphenylboronic acid (VPBA) as functional monomers. The structure and component of smart sensing material was investigated by transmission electron microscopy (TEM), Scanning electron microscopy (SEM), X-ray photoelectron spectroscopic (XPS) and Fourier transform infrared (FTIR), respectively. These results illustrated the smart sensing material was prepared successfully. The recognition characterizations of smart sensing material were evaluated, and results showed that the fluorescence intensity of smart sensing material was reduced gradually, as the concentration of protein increased, and the smart sensing material showed selective recognition for HRP among other proteins. Furthermore, the recognition ability of the smart sensing material for glycoprotein was regulated by controlling the pH value and temperature. Therefore, this strategy opens up new way to construct smart material for detection of glycoprotein. Copyright © 2016 Elsevier B.V. All rights reserved.

Leukoreduced red blood cell transfusions do not induce platelet glycoprotein antibodies in patients with sickle cell disease.

PubMed

Nickel, Robert Sheppard; Winkler, Anne M; Horan, John T; Hendrickson, Jeanne E

2016-09-01

Alloimmunization to red blood cell (RBC) antigens after transfusion is well described in patients with sickle cell disease (SCD). We recently demonstrated that leukocyte-reduced RBC transfusions appeared to induce human leukocyte antigen (HLA) antibodies in some children with SCD; now, we hypothesize that residual platelets contained in transfused RBC products may lead to platelet glycoprotein antibody formation. A cross-sectional study was conducted among never pregnant pediatric patients with SCD who either had received many RBC transfusions or had never received any transfusions. Serum was tested for antibodies to platelet-specific glycoproteins using a commercial enzyme immunoassay. Platelet-specific glycoprotein antibodies were found in 12 of 90 patients (13%) in the transfused group versus 5 of 24 patients (21%) in the never transfused group (p = 0.35). The prevalence of antibodies as well as the median standardized optical density for these two groups was not significantly different for any of the studied platelet glycoprotein antigens. There was no association with the presence of platelet-specific glycoprotein antibodies with either RBC or HLA antibodies. Leukocyte-reduced RBC transfusions do not appear to induce platelet-specific glycoprotein antibodies. The positive platelet-specific glycoprotein antibody results from this study may represent platelet autoantibodies, platelet alloantibodies, or false-positive reactions. A better understanding of the immunobiology of patients with SCD at baseline and after blood product exposure may help improve future transfusion and transplantation. © 2016 AABB.

Impact of a human CMP-sialic acid transporter on recombinant glycoprotein sialylation in glycoengineered insect cells.

PubMed

Mabashi-Asazuma, Hideaki; Shi, Xianzong; Geisler, Christoph; Kuo, Chu-Wei; Khoo, Kay-Hooi; Jarvis, Donald L

2013-02-01

Insect cells are widely used for recombinant glycoprotein production, but they cannot provide the glycosylation patterns required for some biotechnological applications. This problem has been addressed by genetically engineering insect cells to express mammalian genes encoding various glycoprotein glycan processing functions. However, for various reasons, the impact of a mammalian cytosine-5'-monophospho (CMP)-sialic acid transporter has not yet been examined. Thus, we transformed Spodoptera frugiperda (Sf9) cells with six mammalian genes to generate a new cell line, SfSWT-4, that can produce sialylated glycoproteins when cultured with the sialic acid precursor, N-acetylmannosamine. We then super-transformed SfSWT-4 with a human CMP-sialic acid transporter (hCSAT) gene to isolate a daughter cell line, SfSWT-6, which expressed the hCSAT gene in addition to the other mammalian glycogenes. SfSWT-6 cells had higher levels of cell surface sialylation and also supported higher levels of recombinant glycoprotein sialylation, particularly when cultured with low concentrations of N-acetylmannosamine. Thus, hCSAT expression has an impact on glycoprotein sialylation, can reduce the cost of recombinant glycoprotein production and therefore should be included in ongoing efforts to glycoengineer the baculovirus-insect cell system. The results of this study also contributed new insights into the endogenous mechanism and potential mechanisms of CMP-sialic acid accumulation in the Golgi apparatus of lepidopteran insect cells.

Role of the Conserved Oligomeric Golgi Complex in the Abnormalities of Glycoprotein Processing in Breast Cancer Cells

DTIC Science & Technology

2005-05-01

AD Award Number: DAMD17-03-1-0243 TITLE: Role of the Conserved Oligomeric Golgi Complex in the Abnormalities of Glycoprotein Processing in Breast...Glycoprotein Processing in Breast Cancer 5b.GRANTNUMBER Cells DAAD17-03-1-0243 5c. PROGRAM ELEMENT NUMBER 6. AUTHOR(S) 5d. PROJECT NUMBER Sergey N... processing of glycoproteins, exocytosis, protein delivery systems, gene expression, western and northern blot analysis, immunotiuorescence, gradient

Accumulation of hydroxyproline-rich glycoprotein mRNAs in response to fungal elicitor and infection.

PubMed

Showalter, A M; Bell, J N; Cramer, C L; Bailey, J A; Varner, J E; Lamb, C J

1985-10-01

Hydroxyproline-rich glycoproteins (HRGPs) are important structural components of plant cell walls and also accumulate in response to infection as an apparent defense mechanism. Accumulation of HRGP mRNA in biologically stressed bean (Phaseolus vulgaris L.) cells was monitored by blot hybridization with (32)P-labeled tomato genomic HRGP sequences. Elicitor treatment of suspension-cultured cells caused a marked increase in hybridizable HRGP mRNA. The response was less rapid but more prolonged than that observed for mRNAs encoding enzymes of phytoalexin biosynthesis. HRGP mRNA also accumulated during race:cultivar-specific interactions between bean hypocotyls and the partially biotrophic fungus Colletotrichum lindemuthianum, the causal agent of anthracnose. In an incompatible interaction (host resistant) there was an early increase in HRGP mRNA correlated with expression of hypersensitive resistance; whereas, in a compatible interaction (host susceptible), marked accumulation of HRGP mRNA occurred as a delayed response at the onset of lesion formation. In both interactions, mRNA accumulation was observed in uninfected cells distant from the site of fungal inoculation, indicating intercellular transmission of an elicitation signal.

High-frequency ultrasound-guided disruption of glycoprotein VI-targeted microbubbles targets atheroprogressison in mice.

PubMed

Metzger, Katja; Vogel, Sebastian; Chatterjee, Madhumita; Borst, Oliver; Seizer, Peter; Schönberger, Tanja; Geisler, Tobias; Lang, Florian; Langer, Harald; Rheinlaender, Johannes; Schäffer, Tilman E; Gawaz, Meinrad

2015-01-01

Targeted contrast-enhanced ultrasound (CEU) using microbubble agents is a promising non-invasive imaging technique to evaluate atherosclerotic lesions. In this study, we decipher the diagnostic and therapeutic potential of targeted-CEU with soluble glycoprotein (GP)-VI in vivo. Microbubbles were conjugated with the recombinant fusion protein GPVI-Fc (MBGPVI) that binds with high affinity to atherosclerotic lesions. MBGPVI or control microbubbles (MBC) were intravenously administered into ApoE(-/-) or wild type mice and binding of the microbubbles to the vessel wall was visualized by high-resolution CEU. CEU molecular imaging signals of MBGPVI were substantially enhanced in the aortic arch and in the truncus brachiocephalicus in ApoE(-/-) as compared to wild type mice. High-frequency ultrasound (HFU)-guided disruption of MBGPVI enhanced accumulation of GPVI in the atherosclerotic lesions, which may interfere with atheroprogression. Thus, we establish targeted-CEU with soluble GPVI as a novel non-invasive molecular imaging method for atherosclerosis. Further, HFU-guided disruption of GPVI-targeted microbubbles is an innovate therapeutic approach that potentially prevents progression of atherosclerotic disease. Copyright © 2014 Elsevier Ltd. All rights reserved.

Pulse-chase Analysis of N-linked Sugar Chains from Glycoproteins in Mammalian Cells

PubMed Central

Avezov, Edward; Ron, Efrat; Izenshtein, Yana; Adan, Yosef; Lederkremer, Gerardo Z.

2010-01-01

Attachment of the Glc3Man9GlcNAc2 precursor oligosaccharide to nascent polypeptides in the ER is a common modification for secretory proteins. Although this modification was implicated in several biological processes, additional aspects of its function are emerging, with recent evidence of its role in the production of signals for glycoprotein quality control and trafficking. Thus, phenomena related to N-linked glycans and their processing are being intensively investigated. Methods that have been recently developed for proteomic analysis have greatly improved the characterization of glycoprotein N-linked glycans. Nevertheless, they do not provide insight into the dynamics of the sugar chain processing involved. For this, labeling and pulse-chase analysis protocols are used that are usually complex and give very low yields. We describe here a simple method for the isolation and analysis of metabolically labeled N-linked oligosaccharides. The protocol is based on labeling of cells with [2-3H] mannose, denaturing lysis and enzymatic release of the oligosaccharides from either a specifically immunoprecipitated protein of interest or from the general glycoprotein pool by sequential treatments with endo H and N-glycosidase F, followed by molecular filtration (Amicon). In this method the isolated oligosaccharides serve as an input for HPLC analysis, which allows discrimination between various glycan structures according to the number of monosaccharide units comprising them, with a resolution of a single monosaccharide. Using this method we were able to study high mannose N-linked oligosaccharide profiles of total cell glycoproteins after pulse-chase in normal conditions and under proteasome inhibition. These profiles were compared to those obtained from an immunoprecipitated ER-associated degradation (ERAD) substrate. Our results suggest that most NIH 3T3 cellular glycoproteins are relatively stable and that most of their oligosaccharides are trimmed to Man9-8GlcNAc2. In

Pulse-chase analysis of N-linked sugar chains from glycoproteins in mammalian cells.

PubMed

Avezov, Edward; Ron, Efrat; Izenshtein, Yana; Adan, Yosef; Lederkremer, Gerardo Z

2010-04-27

Attachment of the Glc3Man9GlcNAc2 precursor oligosaccharide to nascent polypeptides in the ER is a common modification for secretory proteins. Although this modification was implicated in several biological processes, additional aspects of its function are emerging, with recent evidence of its role in the production of signals for glycoprotein quality control and trafficking. Thus, phenomena related to N-linked glycans and their processing are being intensively investigated. Methods that have been recently developed for proteomic analysis have greatly improved the characterization of glycoprotein N-linked glycans. Nevertheless, they do not provide insight into the dynamics of the sugar chain processing involved. For this, labeling and pulse-chase analysis protocols are used that are usually complex and give very low yields. We describe here a simple method for the isolation and analysis of metabolically labeled N-linked oligosaccharides. The protocol is based on labeling of cells with [2-(3)H] mannose, denaturing lysis and enzymatic release of the oligosaccharides from either a specifically immunoprecipitated protein of interest or from the general glycoprotein pool by sequential treatments with endo H and N-glycosidase F, followed by molecular filtration (Amicon). In this method the isolated oligosaccharides serve as an input for HPLC analysis, which allows discrimination between various glycan structures according to the number of monosaccharide units comprising them, with a resolution of a single monosaccharide. Using this method we were able to study high mannose N-linked oligosaccharide profiles of total cell glycoproteins after pulse-chase in normal conditions and under proteasome inhibition. These profiles were compared to those obtained from an immunoprecipitated ER-associated degradation (ERAD) substrate. Our results suggest that most NIH 3T3 cellular glycoproteins are relatively stable and that most of their oligosaccharides are trimmed to Man9-8GlcNAc2

Paramyxovirus glycoprotein incorporation, assembly and budding: a three way dance for infectious particle production.

PubMed

El Najjar, Farah; Schmitt, Anthony P; Dutch, Rebecca Ellis

2014-08-07

Paramyxoviruses are a family of negative sense RNA viruses whose members cause serious diseases in humans, such as measles virus, mumps virus and respiratory syncytial virus; and in animals, such as Newcastle disease virus and rinderpest virus. Paramyxovirus particles form by assembly of the viral matrix protein, the ribonucleoprotein complex and the surface glycoproteins at the plasma membrane of infected cells and subsequent viral budding. Two major glycoproteins expressed on the viral envelope, the attachment protein and the fusion protein, promote attachment of the virus to host cells and subsequent virus-cell membrane fusion. Incorporation of the surface glycoproteins into infectious progeny particles requires coordinated interplay between the three viral structural components, driven primarily by the matrix protein. In this review, we discuss recent progress in understanding the contributions of the matrix protein and glycoproteins in driving paramyxovirus assembly and budding while focusing on the viral protein interactions underlying this process and the intracellular trafficking pathways for targeting viral components to assembly sites. Differences in the mechanisms of particle production among the different family members will be highlighted throughout.

Paramyxovirus Glycoprotein Incorporation, Assembly and Budding: A Three Way Dance for Infectious Particle Production

PubMed Central

El Najjar, Farah; Schmitt, Anthony P.; Dutch, Rebecca Ellis

2014-01-01

Paramyxoviruses are a family of negative sense RNA viruses whose members cause serious diseases in humans, such as measles virus, mumps virus and respiratory syncytial virus; and in animals, such as Newcastle disease virus and rinderpest virus. Paramyxovirus particles form by assembly of the viral matrix protein, the ribonucleoprotein complex and the surface glycoproteins at the plasma membrane of infected cells and subsequent viral budding. Two major glycoproteins expressed on the viral envelope, the attachment protein and the fusion protein, promote attachment of the virus to host cells and subsequent virus-cell membrane fusion. Incorporation of the surface glycoproteins into infectious progeny particles requires coordinated interplay between the three viral structural components, driven primarily by the matrix protein. In this review, we discuss recent progress in understanding the contributions of the matrix protein and glycoproteins in driving paramyxovirus assembly and budding while focusing on the viral protein interactions underlying this process and the intracellular trafficking pathways for targeting viral components to assembly sites. Differences in the mechanisms of particle production among the different family members will be highlighted throughout. PMID:25105277

Most neutralizing human monoclonal antibodies target novel epitopes requiring both Lassa virus glycoprotein subunits

PubMed Central

Robinson, James E.; Hastie, Kathryn M.; Cross, Robert W.; Yenni, Rachael E.; Elliott, Deborah H.; Rouelle, Julie A.; Kannadka, Chandrika B.; Smira, Ashley A.; Garry, Courtney E.; Bradley, Benjamin T.; Yu, Haini; Shaffer, Jeffrey G.; Boisen, Matt L.; Hartnett, Jessica N.; Zandonatti, Michelle A.; Rowland, Megan M.; Heinrich, Megan L.; Martínez-Sobrido, Luis; Cheng, Benson; de la Torre, Juan C.; Andersen, Kristian G.; Goba, Augustine; Momoh, Mambu; Fullah, Mohamed; Gbakie, Michael; Kanneh, Lansana; Koroma, Veronica J.; Fonnie, Richard; Jalloh, Simbirie C.; Kargbo, Brima; Vandi, Mohamed A.; Gbetuwa, Momoh; Ikponmwosa, Odia; Asogun, Danny A.; Okokhere, Peter O.; Follarin, Onikepe A.; Schieffelin, John S.; Pitts, Kelly R.; Geisbert, Joan B.; Kulakoski, Peter C.; Wilson, Russell B.; Happi, Christian T.; Sabeti, Pardis C.; Gevao, Sahr M.; Khan, S. Humarr; Grant, Donald S.; Geisbert, Thomas W.; Saphire, Erica Ollmann; Branco, Luis M.; Garry, Robert F.

2016-01-01

Lassa fever is a severe multisystem disease that often has haemorrhagic manifestations. The epitopes of the Lassa virus (LASV) surface glycoproteins recognized by naturally infected human hosts have not been identified or characterized. Here we have cloned 113 human monoclonal antibodies (mAbs) specific for LASV glycoproteins from memory B cells of Lassa fever survivors from West Africa. One-half bind the GP2 fusion subunit, one-fourth recognize the GP1 receptor-binding subunit and the remaining fourth are specific for the assembled glycoprotein complex, requiring both GP1 and GP2 subunits for recognition. Notably, of the 16 mAbs that neutralize LASV, 13 require the assembled glycoprotein complex for binding, while the remaining 3 require GP1 only. Compared with non-neutralizing mAbs, neutralizing mAbs have higher binding affinities and greater divergence from germline progenitors. Some mAbs potently neutralize all four LASV lineages. These insights from LASV human mAb characterization will guide strategies for immunotherapeutic development and vaccine design. PMID:27161536

Free oligosaccharides to monitor glycoprotein endoplasmic reticulum-associated degradation in Saccharomyces cerevisiae.

PubMed

Hirayama, Hiroto; Seino, Junichi; Kitajima, Toshihiko; Jigami, Yoshifumi; Suzuki, Tadashi

2010-04-16

In eukaryotic cells, N-glycosylation has been recognized as one of the most common and functionally important co- or post-translational modifications of proteins. "Free" forms of N-glycans accumulate in the cytosol of mammalian cells, but the precise mechanism for their formation and degradation remains unknown. Here, we report a method for the isolation of yeast free oligosaccharides (fOSs) using endo-beta-1,6-glucanase digestion. fOSs were undetectable in cells lacking PNG1, coding the cytoplasmic peptide:N-glycanase gene, suggesting that almost all fOSs were formed from misfolded glycoproteins by Png1p. Structural studies revealed that the most abundant fOS was M8B, which is not recognized well by the endoplasmic reticulum-associated degradation (ERAD)-related lectin, Yos9p. In addition, we provide evidence that some of the ERAD substrates reached the Golgi apparatus prior to retrotranslocation to the cytosol. N-Glycan structures on misfolded glycoproteins in cells lacking the cytosol/vacuole alpha-mannosidase, Ams1p, was still quite diverse, indicating that processing of N-glycans on misfolded glycoproteins was more complex than currently envisaged. Under ER stress, an increase in fOSs was observed, whereas levels of M7C, a key glycan structure recognized by Yos9p, were unchanged. Our method can thus provide valuable information on the molecular mechanism of glycoprotein ERAD in Saccharomyces cerevisiae.

Structure of a trimeric variant of the Epstein-Barr virus glycoprotein B

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

Backovic, Marija; Longnecker, Richard; Jardetzky, Theodore S

Epstein-Barr virus (EBV) is a herpesvirus that is associated with development of malignancies of lymphoid tissue. EBV infections are life-long and occur in >90% of the population. Herpesviruses enter host cells in a process that involves fusion of viral and cellular membranes. The fusion apparatus is comprised of envelope glycoprotein B (gB) and a heterodimeric complex made of glycoproteins H and L. Glycoprotein B is the most conserved envelope glycoprotein in human herpesviruses, and the structure of gB from Herpes simplex virus 1 (HSV-1) is available. Here, we report the crystal structure of the secreted EBV gB ectodomain, which formsmore » 16-nm long spike-like trimers, structurally homologous to the postfusion trimers of the fusion protein G of vesicular stomatitis virus (VSV). Comparative structural analyses of EBV gB and VSV G, which has been solved in its pre and postfusion states, shed light on gB residues that may be involved in conformational changes and membrane fusion. Also, the EBV gB structure reveals that, despite the high sequence conservation of gB in herpesviruses, the relative orientations of individual domains, the surface charge distributions, and the structural details of EBV gB differ from the HSV-1 protein, indicating regions and residues that may have important roles in virus-specific entry.« less

Deletion of the α-(1,3)-Glucan Synthase Genes Induces a Restructuring of the Conidial Cell Wall Responsible for the Avirulence of Aspergillus fumigatus

PubMed Central

Beauvais, Anne; Bozza, Silvia; Kniemeyer, Olaf; Formosa, Céline; Balloy, Viviane; Henry, Christine; Roberson, Robert W.; Dague, Etienne; Chignard, Michel; Brakhage, Axel A.; Romani, Luigina; Latgé, Jean-Paul

2013-01-01

α-(1,3)-Glucan is a major component of the cell wall of Aspergillus fumigatus, an opportunistic human fungal pathogen. There are three genes (AGS1, AGS2 and AGS3) controlling the biosynthesis of α-(1,3)-glucan in this fungal species. Deletion of all the three AGS genes resulted in a triple mutant that was devoid of α-(1,3)-glucan in its cell wall; however, its growth and germination was identical to that of the parental strain in vitro. In the experimental murine aspergillosis model, this mutant was less pathogenic than the parental strain. The AGS deletion resulted in an extensive structural modification of the conidial cell wall, especially conidial surface where the rodlet layer was covered by an amorphous glycoprotein matrix. This surface modification was responsible for viability reduction of conidia in vivo, which explains decrease in the virulence of triple agsΔ mutant. PMID:24244155

Changes in intestinal absorption of nutrients and brush border glycoproteins after total parenteral nutrition in rats.

PubMed Central

Miura, S; Tanaka, S; Yoshioka, M; Serizawa, H; Tashiro, H; Shiozaki, H; Imaeda, H; Tsuchiya, M

1992-01-01

The effect of total parenteral nutrition on nutrients absorption and glycoprotein changes of brush border membrane was examined in rat small intestine. In total parenteral nutrition rats, a marked decrease in activity of brush border enzymes was observed mainly in the proximal and middle segments of the intestine. Galactose perfusion of jejunal segment showed that hexose absorption was significantly inhibited, while intestinal absorption of glycine or dipeptide, glycylglycine was not significantly affected by total parenteral nutrition treatment. When brush border membrane glycoprotein profile was examined by [3H]-glucosamine or [3H]-fucose incorporation into jejunal loops, significant changes were observed in the glycoprotein pattern of brush border membrane especially in the high molecular weight range over 120 kDa after total parenteral nutrition treatment, suggesting strong dependency of glycoprotein synthesis on luminal substances. Molecular weight of sucrase isomaltase in brush border membrane detected by specific antibody showed no significant difference, however, in total parenteral nutrition and control rats. Also, molecular weight of specific sodium glucose cotransporter of intestinal brush border membrane detected by selective photoaffinity labelling was not altered in total parenteral nutrition rats. It may be that prolonged absence of oral food intake may produce significant biochemical changes in brush border membrane glycoprotein and absorptive capacity of small intestine, but these changes were not observed in all brush border membrane glycoproteins. Images Figure 1 Figure 2 Figure 3 Figure 4 PMID:1582592

Emerging structural insights into glycoprotein quality control coupled with N-glycan processing in the endoplasmic reticulum.

PubMed

Satoh, Tadashi; Yamaguchi, Takumi; Kato, Koichi

2015-01-30

In the endoplasmic reticulum (ER), the sugar chain is initially introduced onto newly synthesized proteins as a triantennary tetradecasaccharide (Glc3Man9GlcNAc2). The attached oligosaccharide chain is subjected to stepwise trimming by the actions of specific glucosidases and mannosidases. In these processes, the transiently expressed N-glycans, as processing intermediates, function as signals for the determination of glycoprotein fates, i.e., folding, transport, or degradation through interactions of a series of intracellular lectins. The monoglucosylated glycoforms are hallmarks of incompletely folded states of glycoproteins in this system, whereas the outer mannose trimming leads to ER-associated glycoprotein degradation. This review outlines the recently emerging evidence regarding the molecular and structural basis of this glycoprotein quality control system, which is regulated through dynamic interplay among intracellular lectins, glycosidases, and glycosyltransferase. Structural snapshots of carbohydrate-lectin interactions have been provided at the atomic level using X-ray crystallographic analyses. Conformational ensembles of uncomplexed triantennary high-mannose-type oligosaccharides have been characterized in a quantitative manner using molecular dynamics simulation in conjunction with nuclear magnetic resonance spectroscopy. These complementary views provide new insights into glycoprotein recognition in quality control coupled with N-glycan processing.

Glycoprotein changes in non-insulin dependent diabetic rats: effect of N-benzoyl-D-phenylalanine and metformin.

PubMed

Pari, Leelavinothan; Ashokkumar, Natarajan

2006-01-01

The effect of N-benzoyl-D-phenylalanine (NBDP) and metformin on neonatal streptozotocin (nSTZ) induced diabetes has been studied on plasma and tissue glycoproteins. In some pathological conditions, such as cancer, rheumatoid arthritis and diabetes, there is an abnormal glycosylation of acute phase serum proteins. As most serum proteins are produced in the liver, we have examined glycoprotein metabolism in diabetic condition. To induce non-insulin-dependent diabetes mellitus (NIDDM) a single dose of streptozotocin (100 mg/kg body weight) was injected into two day old rats. After 10-12 weeks, rats weighing above 150 g were selected for NIDDM model. In these rat, blood glucose and plasma glycoproteins were significantly increased whereas plasma insulin was significantly decreased. There was a significant decrease in the level of sialic acid and elevated levels of hexose, hexosamine and fucose in tissues. Oral administration of NBDP and metformin to diabetic rats decreased blood glucose and plasma glycoproteins. Plasma insulin and tissue sialic acid were increased whereas tissue concentrations of hexose, hexosamine and fucose were near normal. Our study suggests that NBDP and metformin possess a significant beneficial effect on glycoproteins in addition to their antidiabetic effect.

Carcinoma-specific Ulex europaeus agglutinin-I binding glycoproteins of human colorectal carcinoma and its relation to carcinoembryonic antigen.

PubMed

Matsushita, Y; Yonezawa, S; Nakamura, T; Shimizu, S; Ozawa, M; Muramatsu, T; Sato, E

1985-08-01

Glycoproteins binding to Ulex europaeus agglutinin-I (UEA-I) lectin, which recognizes the terminal alpha-L-fucose residue, were analyzed in 18 cases of human colorectal carcinoma by sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by the Western blotting method. In the distal large bowel (descending and sigmoid colon and rectum), high-molecular-weight glycoproteins binding to UEA-I existed in carcinoma tissue but not in normal mucosa. In the proximal large bowel (ascending and transverse colon), high-molecular-weight glycoproteins binding to UEA-I were found both in normal mucosa and in carcinoma tissue, whereas those from the carcinoma tissue had an apparently lower molecular weight as compared to the weight of those from the normal mucosa. Thus there is a biochemical difference in UEA-I binding glycoproteins between the normal mucosa and the carcinoma tissue, although in our previous histochemical study no difference was observed in UEA-I binding glycoproteins of the proximal large bowel between the carcinoma tissue and the normal mucosa. Furthermore, carcinoembryonic antigen from the carcinoma tissue was found to have the same electrophoretical mobility as the UEA-I binding glycoproteins.

Contribution of tumor endothelial cells to drug resistance: anti-angiogenic tyrosine kinase inhibitors act as p-glycoprotein antagonists.

PubMed

Bani, MariaRosa; Decio, Alessandra; Giavazzi, Raffaella; Ghilardi, Carmen

2017-05-01

Tumor endothelial cells (TEC) differ from the normal counterpart, in both gene expression and functionality. TEC may acquire drug resistance, a characteristic that is maintained in vitro. There is evidence that TEC are more resistant to chemotherapeutic drugs, substrates of ATP-binding cassette (ABC) transporters. TEC express p-glycoprotein (encoded by ABCB1), while no difference in other ABC transporters was revealed compared to normal endothelia. A class of tyrosine kinase inhibitors (TKI), used as angiostatic compounds, interferes with the ATPase activity of p-glycoprotein, thus impairing its functionality. The exposure of ovarian adenocarcinoma TEC to the TKIs sunitinib or sorafenib was found to abrogate resistance (proliferation and motility) to doxorubicin and paclitaxel in vitro, increasing intracellular drug accumulation. A similar effect has been reported by the p-glycoprotein inhibitor verapamil. No beneficial effect was observed in combination with cytotoxic drugs that are not p-glycoprotein substrates. The current paper reviews the mechanisms of TEC chemoresistance and shows the role of p-glycoprotein in mediating such resistance. Inhibition of p-glycoprotein by anti-angiogenic TKI might contribute to the beneficial effect of these small molecules, when combined with chemotherapy, in counteracting acquired drug resistance.

Expression of the rabies virus glycoprotein in transgenic tomatoes.

PubMed

McGarvey, P B; Hammond, J; Dienelt, M M; Hooper, D C; Fu, Z F; Dietzschold, B; Koprowski, H; Michaels, F H

1995-12-01

We have engineered tomato plants (Lycopersicon esculentum Mill var. UC82b) to express a gene for the glycoprotein (G-protein), which coats the outer surface of the rabies virus. The recombinant constructs contained the G-protein gene from the ERA strain of rabies virus, including the signal peptide, under the control of the 35S promoter of cauliflower mosaic virus. Plants were transformed by Agrobacterium tumefaciens-mediated transformation of cotyledons and tissue culture on selective media. PCR confirmed the presence of the G-protein gene in plants surviving selection. Northern blot analysis indicated that RNA of the appropriate molecular weight was produced in both leaves and fruit of the transgenic plants. The recombinant G-protein was immunoprecipitated and detected by Western blot from leaves and fruit using different antisera. The G-protein expressed in tomato appeared as two distinct bands with apparent molecular mass of 62 and 60 kDa as compared to the 66 kDa observed for G-protein from virus grown in BHK cells. Electron microscopy of leaf tissue using immunogold-labeling and antisera specific for rabies G-protein showed localization of the G-protein to the Golgi bodies, vesicles, plasmalemma and cell walls of vascular parenchyma cells. In light of our previous demonstration that orally administered rabies G-protein from the same ERA strain elicits protective immunity in animals, these transgenic plants should provide a valuable tool for the development of edible oral vaccines.

Celastraceae sesquiterpenes as a new class of modulators that bind specifically to human P-glycoprotein and reverse cellular multidrug resistance.

PubMed

Muñoz-Martínez, Francisco; Lu, Peihua; Cortés-Selva, Fernando; Pérez-Victoria, José María; Jiménez, Ignacio A; Ravelo, Angel G; Sharom, Frances J; Gamarro, Francisco; Castanys, Santiago

2004-10-01

Overexpression of ABCB1 (MDR1) P-glycoprotein, a multidrug efflux pump, is one mechanism by which tumor cells may develop multidrug resistance (MDR), preventing the successful chemotherapeutic treatment of cancer. Sesquiterpenes from Celastraceae family are natural compounds shown previously to reverse MDR in several human cancer cell lines and Leishmania strains. However, their molecular mechanism of reversion has not been characterized. In the present work, we have studied the ability of 28 dihydro-beta-agarofuran sesquiterpenes to reverse the P-glycoprotein-dependent MDR phenotype and elucidated their molecular mechanism of action. Cytotoxicity assays using human MDR1-transfected NIH-3T3 cells allowed us to select the most potent sesquiterpenes reversing the in vitro resistance to daunomycin and vinblastine. Flow cytometry experiments showed that the above active compounds specifically inhibited drug transport activity of P-glycoprotein in a saturable, concentration-dependent manner (K(i) down to 0.24 +/- 0.01 micromol/L) but not that of ABCC1 (multidrug resistance protein 1; MRP1), ABCC2 (MRP2), and ABCG2 (breast cancer resistance protein; BCRP) transporters. Moreover, sesquiterpenes inhibited at submicromolar concentrations the P-glycoprotein-mediated transport of [(3)H]colchicine and tetramethylrosamine in plasma membrane from CH(R)B30 cells and P-glycoprotein-enriched proteoliposomes, supporting that P-glycoprotein is their molecular target. Photoaffinity labeling in plasma membrane and fluorescence spectroscopy experiments with purified protein suggested that sesquiterpenes interact with transmembrane domains of P-glycoprotein. Finally, sesquiterpenes modulated P-glycoprotein ATPase-activity in a biphasic, concentration-dependent manner: they stimulated at very low concentrations but inhibited ATPase activity as noncompetitive inhibitors at higher concentrations. Sesquiterpenes from Celastraceae are promising P-glycoprotein modulators with potential

Plant Cell Wall Dynamics in Compatible and Incompatible Potato Response to Infection Caused by Potato Virus Y (PVYNTN)

PubMed Central

Lockhart, Benham E. L.

2018-01-01

The cell wall provides the structure of the plant, and also acts as a barier against biotic stress. The vein necrosis strain of Potato virus Y (PVYNTN) induces necrotic disease symptoms that affect both plant growth and yield. Virus infection triggers a number of inducible basal defense responses, including defense proteins, especially those involved in cell wall metabolism. This study investigates the comparison of cell wall host dynamics induced in a compatible (potato cv. Irys) and incompatible (potato cv. Sárpo Mira with hypersensitive reaction gene Ny-Smira) PVYNTN–host–plant interaction. Ultrastructural analyses revealed numerous cell wall changes induced by virus infection. Furthermore, the localization of essential defensive wall-associated proteins in susceptible and resistant potato host to PVYNTN infection were investigated. The data revealed a higher level of detection of pathogenesis-related protein 2 (PR-2) in a compatible compared to an incompatible (HR) interaction. Immunofluorescence analyses indicated that hydroxyproline-rich glycoproteins (HRGP) (extensin) synthesis was induced, whereas that of cellulose synthase catalytic subunits (CesA4) decreased as a result of PVYNTN infection. The highest level of extensin localization was found in HR potato plants. Proteins involved in cell wall metabolism play a crucial role in the interaction because they affect the spread of the virus. Analysis of CesA4, PR-2 and HRGP deposition within the apoplast and symplast confirmed the active trafficking of these proteins as a step-in potato cell wall remodeling in response to PVYNTN infection. Therefore, cell wall reorganization may be regarded as an element of “signWALLing”—involving apoplast and symplast activation as a specific response to viruses. PMID:29543714

Platelet Glycoprotein lb-1X and Malignancy

DTIC Science & Technology

2010-09-01

supporting the accumulation of more platelets and the elaboration of a fibrin - rich network produced by coagulation factors. This paradigm has been...a platelet - rich thrombus by tethering the platelet to a thrombogenic surface. Several ligands binding to GP Ib-IX have been identified, including...08-1-0576 TITLE: Platelet Glycoprotein lb-1X and Malignancy PRINCIPAL INVESTIGATOR: Dr. Jerry Ware

A plasma membrane-type Ca[sup 2+]-ATPase of 120 kilodaltons on the endoplasmic reticulum from carrot (Daucus carota) cells

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

Chen, F.H.; Ratterman, D.M.; Sze, H.

1993-06-01

Cytosolic Ca[sup 2+] levels are regulated in part by Ca[sup 2+]-pumping ATPases that export Ca[sup 2+] from the cytoplasm; The types and properties of Ca[sup 2+] pumps in plants are not well understood. The kinetic properties of a 120-kD phosphoenzyme (PE) intermediate formed during the reaction cycle of a Ca[sup 2+]-ATPase from suspension-cultured carrot (Daucus carota) cells are characterized. Only one Ca[sup 2+]-dependent phosphoprotein was formed when carrot membrane vesicles were incubated with [[gamma]-[sup 32]P]ATP. Formation of this 120-kD phosphoprotein was inhibited by vanadate, enhanced by La[sup 3+], and decreased by hydroxylamine, confirming its identification as an intermediate of amore » phosphorylated-type Ca[sup 2+]-translocating ATPase. The 120-kD Ca[sup 2+]-ATPase was most abundant in endoplasmic reticulum-enriched fractions, in which the Ca[sup 2+]-ATPase was estimated to be 0.1% of membrane protein. Direct quantitation of Ca[sup 2+]-dependent phosphoprotein was used to examine the kinetics of PE formation. PE formation exhibited a K[sub m] for Ca[sup 2+] of 1 to 2 [mu]m and a K[sub m] for ATP of 67 nm. Relative affinities of substrates, determined by competition experiments, were 0.075 [mu]m for ATP, 1 [mu]m for ADP, 100 [mu]m for ITP, and 250 [mu]m for GTP. Thapsigargin and cyclopiazonic acid, specific inhibitors of animal sarcoplasmic/endoplasmic reticulum Ca[sup 2+]-ATPase, had no effect on PE formation; erythrosin B inhibited with 50% inhibition at <0.1 [mu]m. Calmodulin (1 [mu]m) stimulated PE formation by 25%. The results indicate that the carrot 120-kD Ca[sup 2+]-ATPase is similar but not identical to animal plasma membrane-type Ca[sup 2+]-ATPase and yet is located on endomembranes, such as the endoplasmic reticulum. This type of Ca[sup 2+] pump may reside on the cortical endoplasmic reticulum, thought to play a major role in anchoring the cytoskeleton and in facilitating secretion. 34 refs., 9 figs., 3 tabs.« less

The adhesion of Pseudomonas aeruginosa to high molecular weight human tear film species corresponds to glycoproteins reactive with Sambucus nigra lectin.

PubMed

Aristoteli, Lina Panayiota; Willcox, Mark D P

2006-11-01

Pseudomonas aeruginosa is a pathogen gaining prevalence in contact lens-related corneal ulcers. Tear outflow protects the ocular surface, where high molecular weight tear glycoproteins bind bacteria for removal from the eye. The purpose of the present study was to identify glycoproteins in human tears involved in the adhesion of ocular P. aeruginosa isolates. Basal human tears were applied to a bacterial adhesion assay involving electrophoretic separation of tear components, transfer to nitrocellulose and incubation with biotin-labelled bacteria. Glycoproteins were further characterised using lectin profiling. The results showed large-dimension agarose gels were imperative for the detection of at least four glycoproteins with a migration >200 kDa, including species not previously identified. P. aeruginosa 6294 preferentially bound to a well-defined glycoprotein near the origin of the gel that, unlike other glycoproteins >200 kDa, reacted with Sambucus nigra lectin (sialic acid alpha2-6) but not WGA lectin (N-acetylglucosamine, sialic acid alpha2-3). Adhesion did not involve free biotin label or hydrophobic interactions. Also, the pre-incubation of separated tear glycoproteins with S. nigra lectin increased subsequent adhesion of 6294 to this tear glycoprotein. The less virulent Paer1 strain showed diffuse adhesion in the S. nigra-reactive region at the gel origin. In conclusion, an overlay adhesion assay was developed that identified slow-migrating sialylated glycoprotein species in human tears preferentially bound by P. aeruginosa ocular strains, and S. nigra lectin seemed to enhance the interaction. The study provides a basis for direct investigation of bacterial adhesion to glycoproteins with an apparent migration >200 kDa in tear fluid.

Free Oligosaccharides to Monitor Glycoprotein Endoplasmic Reticulum-associated Degradation in Saccharomyces cerevisiae*

PubMed Central

Hirayama, Hiroto; Seino, Junichi; Kitajima, Toshihiko; Jigami, Yoshifumi; Suzuki, Tadashi

2010-01-01

In eukaryotic cells, N-glycosylation has been recognized as one of the most common and functionally important co- or post-translational modifications of proteins. “Free” forms of N-glycans accumulate in the cytosol of mammalian cells, but the precise mechanism for their formation and degradation remains unknown. Here, we report a method for the isolation of yeast free oligosaccharides (fOSs) using endo-β-1,6-glucanase digestion. fOSs were undetectable in cells lacking PNG1, coding the cytoplasmic peptide:N-glycanase gene, suggesting that almost all fOSs were formed from misfolded glycoproteins by Png1p. Structural studies revealed that the most abundant fOS was M8B, which is not recognized well by the endoplasmic reticulum-associated degradation (ERAD)-related lectin, Yos9p. In addition, we provide evidence that some of the ERAD substrates reached the Golgi apparatus prior to retrotranslocation to the cytosol. N-Glycan structures on misfolded glycoproteins in cells lacking the cytosol/vacuole α-mannosidase, Ams1p, was still quite diverse, indicating that processing of N-glycans on misfolded glycoproteins was more complex than currently envisaged. Under ER stress, an increase in fOSs was observed, whereas levels of M7C, a key glycan structure recognized by Yos9p, were unchanged. Our method can thus provide valuable information on the molecular mechanism of glycoprotein ERAD in Saccharomyces cerevisiae. PMID:20150426

Thiolated polymers: evidence for the formation of disulphide bonds with mucus glycoproteins.

PubMed

Leitner, Verena M; Walker, Greg F; Bernkop-Schnürch, Andreas

2003-09-01

Disulphide bonds between thiolated polymers (thiomers) and cysteine-rich subdomains of mucus glycoproteins are supposed to be responsible for the enhanced mucoadhesive properties of thiomers. This study set out to provide evidence for these covalent interactions using poly(acrylic acid)-cysteine conjugates of 2 and 450 kDa (PAA2-Cys, PAA450-Cys) displaying 402.5-776.0 micromol thiol groups per gram polymer. The effect of the disulphide bond breaker cysteine on thiomer-mucin disulphide bonds was monitored by (1) mucoadhesion studies and (2) rheological studies. Furthermore, (3) diffusion studies and (4) gel filtration studies were performed with thiomer-mucus mixtures. The addition of cysteine significantly (P<0.01) reduced the adhesion of thiomer tablets to porcine mucosa and G'/G" values of thiomer-mucin mixtures, whereas unthiolated controls were not influenced. These results indicate the cleavage of disulphide bonds between thiomer and mucus glycoproteins. Diffusion studies demonstrated that a 12.8-fold higher concentration of the thiomer (PAA2-Cys) remains in the mucin gel than the corresponding unmodified polymer. Gel filtration studies showed that PAA2-Cys was able to form disulphide bonds with mucin glycoproteins resulting in an altered elution profile of the mucin/PAA2-Cys mixture in comparison to mucin alone or mucin/PAA2 mixture. According to these results, the study provides evidence for the formation of covalent bonds between thiomer and mucus glycoproteins.

Methylation of Notch3 modulates chemoresistance via P-glycoprotein.

PubMed

Gu, Xiaoting; Lu, Yangfan; He, Dongxu; Lu, Chunxiao; Jin, Jian; Lu, Xiaojie; Ma, Xin

2016-12-05

The global gene expression and DNA methylation of genes in adriamycin-resistant human breast cancer cells (MCF-7/ADM cells) are similar to those in paclitaxel-resistant MCF-7 cells (MCF-7/PTX) and are significantly different from those in wild-type MCF-7 cells. DNA methylation is associated with chemoresistance in breast cancer and changes the characteristics of chemoresistant and chemosensitive cells. Here, we showed that the tumor-suppressor gene Notch3 was inactivated due to epigenetic silencing DNA hypermethylation in MCF-7/ADM cells. In addition, the drug efflux pump P-glycoprotein was negatively regulated by Notch3 and highly expressed in MCF-7/ADM cells. Taken together, our findings demonstrated that hypermethylation of Notch3 causes activation of P-glycoprotein in adriamycin-resistant cells. Copyright © 2016 Elsevier B.V. All rights reserved.

Immunolocalization and Changes of Hydroxyproline-Rich Glycoproteins During Symbiotic Germination of Dendrobium officinale.

PubMed

Li, Yuan-Yuan; Chen, Xiao-Mei; Zhang, Ying; Cho, Yu-Hsiu; Wang, Ai-Rong; Yeung, Edward C; Zeng, Xu; Guo, Shun-Xing; Lee, Yung-I

2018-01-01

Hydroxyproline-rich glycoproteins (HRGPs) are abundant cell wall components involved in mycorrhizal symbiosis, but little is known about their function in orchid mycorrhizal association. To gain further insight into the role of HRGPs in orchid symbiosis, the location and function of HRGPs were investigated during symbiotic germination of Dendrobium officinale . The presence of JIM11 epitope in developing protocorms was determined using immunodot blots and immunohistochemical staining procedures. Real-time PCR was also employed to verify the expression patterns of genes coding for extensin-like genes selected from the transcriptomic database. The importance of HRGPs in symbiotic germination was further investigated using 3,4-dehydro-L-proline (3,4-DHP), an inhibitor of HRGP biosynthesis. In symbiotic cultures, immunodot blots of JIM11 signals were moderate in mature seeds, and the signals became stronger in swollen embryos. After germination, signal intensities decreased in developing protocorms. In contrast, in asymbiotic cultures, JIM11 signals were much lower as compared with those stages in symbiotic cultures. Immunofluorescence staining enabled the visualization of JIM11 epitope in mature embryo and protocorm cells. Positive signals were initially localized in the larger cells near the basal (suspensor) end of uninfected embryos, marking the future colonization site of fungal hyphae. After 1 week of inoculation, the basal end of embryos had been colonized, and a strong signal was detected mostly at the mid- and basal regions of the enlarging protocorm. As protocorm development progressed, the signal was concentrated in the colonized cells at the basal end. In colonized cells, signals were present in the walls and intracellularly associated with hyphae and the pelotons. The precise localization of JIM11 epitope is further examined by immunogold labeling. In the colonized cells, gold particles were found mainly in the cell wall and the interfacial matrix near the

Immunolocalization and Changes of Hydroxyproline-Rich Glycoproteins During Symbiotic Germination of Dendrobium officinale

PubMed Central

Li, Yuan-Yuan; Chen, Xiao-Mei; Zhang, Ying; Cho, Yu-Hsiu; Wang, Ai-Rong; Yeung, Edward C.; Zeng, Xu; Guo, Shun-Xing; Lee, Yung-I

2018-01-01

Hydroxyproline-rich glycoproteins (HRGPs) are abundant cell wall components involved in mycorrhizal symbiosis, but little is known about their function in orchid mycorrhizal association. To gain further insight into the role of HRGPs in orchid symbiosis, the location and function of HRGPs were investigated during symbiotic germination of Dendrobium officinale. The presence of JIM11 epitope in developing protocorms was determined using immunodot blots and immunohistochemical staining procedures. Real-time PCR was also employed to verify the expression patterns of genes coding for extensin-like genes selected from the transcriptomic database. The importance of HRGPs in symbiotic germination was further investigated using 3,4-dehydro-L-proline (3,4-DHP), an inhibitor of HRGP biosynthesis. In symbiotic cultures, immunodot blots of JIM11 signals were moderate in mature seeds, and the signals became stronger in swollen embryos. After germination, signal intensities decreased in developing protocorms. In contrast, in asymbiotic cultures, JIM11 signals were much lower as compared with those stages in symbiotic cultures. Immunofluorescence staining enabled the visualization of JIM11 epitope in mature embryo and protocorm cells. Positive signals were initially localized in the larger cells near the basal (suspensor) end of uninfected embryos, marking the future colonization site of fungal hyphae. After 1 week of inoculation, the basal end of embryos had been colonized, and a strong signal was detected mostly at the mid- and basal regions of the enlarging protocorm. As protocorm development progressed, the signal was concentrated in the colonized cells at the basal end. In colonized cells, signals were present in the walls and intracellularly associated with hyphae and the pelotons. The precise localization of JIM11 epitope is further examined by immunogold labeling. In the colonized cells, gold particles were found mainly in the cell wall and the interfacial matrix near the

Structure of Respiratory Syncytial Virus Fusion Glycoprotein in the Postfusion Conformation Reveals Preservation of Neutralizing Epitopes

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

McLellan, Jason S.; Yang, Yongping; Graham, Barney S.

2011-09-16

Respiratory syncytial virus (RSV) invades host cells via a type I fusion (F) glycoprotein that undergoes dramatic structural rearrangements during the fusion process. Neutralizing monoclonal antibodies, such as 101F, palivizumab, and motavizumab, target two major antigenic sites on the RSV F glycoprotein. The structures of these sites as peptide complexes with motavizumab and 101F have been previously determined, but a structure for the trimeric RSV F glycoprotein ectodomain has remained elusive. To address this issue, we undertook structural and biophysical studies on stable ectodomain constructs. Here, we present the 2.8-{angstrom} crystal structure of the trimeric RSV F ectodomain in itsmore » postfusion conformation. The structure revealed that the 101F and motavizumab epitopes are present in the postfusion state and that their conformations are similar to those observed in the antibody-bound peptide structures. Both antibodies bound the postfusion F glycoprotein with high affinity in surface plasmon resonance experiments. Modeling of the antibodies bound to the F glycoprotein predicts that the 101F epitope is larger than the linear peptide and restricted to a single protomer in the trimer, whereas motavizumab likely contacts residues on two protomers, indicating a quaternary epitope. Mechanistically, these results suggest that 101F and motavizumab can bind to multiple conformations of the fusion glycoprotein and can neutralize late in the entry process. The structural preservation of neutralizing epitopes in the postfusion state suggests that this conformation can elicit neutralizing antibodies and serve as a useful vaccine antigen.« less

Myo-Inositol trisphosphate mobilizes calcium from fusogenic carrot (Daucus carota L. ) protoplasts

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

Rincon, M.; Boss, W.F.

1987-02-01

To determine whether or not inositol trisphosphate (IP/sub 3/) mobilizes calcium in higher plant cells; they investigated the effect of IP/sub 3/ on Ca/sup 2 +/ fluxes in fusogenic carrot (Daucus carota L.) protoplasts. The protoplasts were incubated in /sup 45/Ca/sup 2 +/-containing medium and the /sup 45/Ca/sup 2 +/ associated with the protoplasts was monitored with time. Addition of IP/sub 3/ (20 micromolar) caused a 17% net loss of the accumulated /sup 45/Ca/sup 2 +/ within 4 minutes. There was a reuptake of /sup 45/Ca/sup 2 +/ and the protoplasts recovered to their initial value by 10 minutes. Phyticmore » acid (IP/sub 6/), also stimulated /sup 45/Ca/sup 2 +/ efflux from the protoplasts. Both the IP/sub 3/- and the IP/sub 6/-induced /sup 45/Ca/sup 2 +/ efflux were inhibited by the calmodulin antagonist, trifluoperazine.« less

Antibodies Targeting Novel Neutralizing Epitopes of Hepatitis C Virus Glycoprotein Preclude Genotype 2 Virus Infection

PubMed Central

Rao, Huiying; Jiang, Dong; Wang, Jianghua; Xie, Xingwang; Wei, Lai

2015-01-01

Currently, there is no effective vaccine to prevent hepatitis C virus (HCV) infection, partly due to our insufficient understanding of the virus glycoprotein immunology. Most neutralizing antibodies (nAbs) were identified using glycoprotein immunogens, such as recombinant E1E2, HCV pseudoparticles or cell culture derived HCV. However, the fact that in the HCV acute infection phase, only a small proportion of patients are self-resolved accompanied with the emergence of nAbs, indicates the limited immunogenicity of glycoprotein itself to induce effective antibodies against a highly evolved virus. Secondly, in previous reports, the immunogen sequence was mostly the genotype of the 1a H77 strain. Rarely, other genotypes/subtypes have been studied, although theoretically one genotype/subtype immunogen is able to induce cross-genotype neutralizing antibodies. To overcome these drawbacks and find potential novel neutralizing epitopes, 57 overlapping peptides encompassing the full-length glycoprotein E1E2 of subtype 1b were synthesized to immunize BALB/c mice, and the neutralizing reactive of the induced antisera against HCVpp genotypes 1–6 was determined. We defined a domain comprising amino acids (aa) 192–221, 232–251, 262–281 and 292–331 of E1, and 421–543, 564–583, 594–618 and 634–673 of E2, as the neutralizing regions of HCV glycoprotein. Peptides PUHI26 (aa 444–463) and PUHI45 (aa 604–618)-induced antisera displayed the most potent broad neutralizing reactive. Two monoclonal antibodies recognizing the PUHI26 and PUHI45 epitopes efficiently precluded genotype 2 viral (HCVcc JFH and J6 strains) infection, but they did not neutralize other genotypes. Our study mapped a neutralizing epitope region of HCV glycoprotein using a novel immunization strategy, and identified two monoclonal antibodies effective in preventing genotype 2 virus infection. PMID:26406225

Use of an informed search space maximizes confidence of site-specific assignment of glycoprotein glycosylation.

PubMed

Khatri, Kshitij; Klein, Joshua A; Zaia, Joseph

2017-01-01

In order to interpret glycopeptide tandem mass spectra, it is necessary to estimate the theoretical glycan compositions and peptide sequences, known as the search space. The simplest way to do this is to build a naïve search space from sets of glycan compositions from public databases and to assume that the target glycoprotein is pure. Often, however, purified glycoproteins contain co-purified glycoprotein contaminants that have the potential to confound assignment of tandem mass spectra based on naïve assumptions. In addition, there is increasing need to characterize glycopeptides from complex biological mixtures. Fortunately, liquid chromatography-mass spectrometry (LC-MS) methods for glycomics and proteomics are now mature and accessible. We demonstrate the value of using an informed search space built from measured glycomes and proteomes to define the search space for interpretation of glycoproteomics data. We show this using α-1-acid glycoprotein (AGP) mixed into a set of increasingly complex matrices. As the mixture complexity increases, the naïve search space balloons and the ability to assign glycopeptides with acceptable confidence diminishes. In addition, it is not possible to identify glycopeptides not foreseen as part of the naïve search space. A search space built from released glycan glycomics and proteomics data is smaller than its naïve counterpart while including the full range of proteins detected in the mixture. This maximizes the ability to assign glycopeptide tandem mass spectra with confidence. As the mixture complexity increases, the number of tandem mass spectra per glycopeptide precursor ion decreases, resulting in lower overall scores and reduced depth of coverage for the target glycoprotein. We suggest use of α-1-acid glycoprotein as a standard to gauge effectiveness of analytical methods and bioinformatics search parameters for glycoproteomics studies. Graphical Abstract Assignment of site specific glycosylation from LC

Calmodulin stimulation of calcium transport in carrot microsomal vesicles. [Daucus carota

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

Pierce, W.S.; Sze, H.

1987-04-01

ATP-dependent /sup 45/Ca/sup 2 +/ uptake into microsomal vesicles isolated from cultured carrot cells (Daucus carota Danvers) was stimulated 2-3 fold by 5 ug/ml calmodulin (CaM). Microsomal vesicles separated with a linear sucrose gradient showed two peaks with CaM-stimulated Ca/sup 2 +/ uptake activities. One peak (at 1.12 g/cc) comigrated with the activity of the antimycin A-insensitive NADH-dependent cytochrome c reductase. This transport activity was enhanced 10-20 fold by 10 mM oxalate and appeared to be associates with vesicles derived primarily from the ER. The other peak of CaM-stimulated Ca/sup 2 +/ uptake (at 1.17 g/cc) was not affected bymore » oxalate. These vesicles are probably derived from the plasma membrane. Preliminary experiments with the low-density vesicles (ER) vesicles, indicate that inositol-1,4,5-trisphosphate caused a transient reduction in intravesicular Ca/sup 2 +/. These results are consistent with the ER being an important site of intracellular Ca/sup 2 +/ regulation.« less

The macromolecular properties of blood-group-specific glycoproteins. Characterization of a series of fractions obtained by solvent fractionation

PubMed Central

Creeth, J. Michael; Bhaskar, K. Ramakrishnan; Donald, Alastair S. R.; Morgan, Walter T. J.

1974-01-01

1. The glycoprotein components of a human ovarian-cyst fluid were isolated by a solvent [95% (w/w) phenol]-extraction procedure; the phenol-insoluble water-soluble glycoprotein was further fractionated by (NH4)2SO4 and by ethanol to yield eight fractions. 2. The fractions were analysed in terms of amino acids, fucose, galactose, N-acetylglucosamine, N-acetylgalactosamine and sialic acid. Variations occurred, particularly in the proportion of peptide; these were partly correlated with varying extent of serological activity. 3. The fractions were characterized physicochemically in terms of buoyant density and degree of spreading in a density gradient, sedimentation velocity and molecular weight; their partial specific volumes and specific refraction increments were also determined. 4. The fractions showed wide variations in their sedimentation-velocity and density-gradient patterns, and gave evidence of pauci-dispersity in density. The fraction regarded as the most typical blood-group-specific glycoprotein sedimented as a single rapidly spreading peak and was of high molecular weight. 5. Significant correlations were observed between the physical properties of the glycoprotein fractions and the amount of their peptide component. The buoyant densities and sedimentation coefficients varied in a manner that suggested the existence of two families of glycoproteins. 6. It is suggested that variability in the extent of glycosylation, or in the degree of cross-linking, might account for the two families of glycoproteins, and that the extent of cross-linkage might also be a factor determining the solubility of these glycoproteins in hot saturated (NH4)2SO4. ImagesFig. 1.PLATE 1 PMID:4219280

Chimeric bovine respiratory syncytial virus with attachment and fusion glycoproteins replaced by bovine parainfluenza virus type 3 hemagglutinin-neuraminidase and fusion proteins.

PubMed

Stope, M B; Karger, A; Schmidt, U; Buchholz, U J

2001-10-01

Chimeric bovine respiratory syncytial viruses (BRSV) expressing glycoproteins of bovine parainfluenza virus type 3 (BPIV-3) instead of BRSV glycoproteins were generated from cDNA. In the BRSV antigenome cDNA, the open reading frames of the major BRSV glycoproteins, attachment protein G and fusion protein F, were replaced individually or together by those of the BPIV-3 hemagglutinin-neuraminidase (HN) and/or fusion (F) glycoproteins. Recombinant virus could not be recovered from cDNA when the BRSV F open reading frame was replaced by the BPIV-3 F open reading frame. However, cDNA recovery of the chimeric virus rBRSV-HNF, with both glycoproteins replaced simultaneously, and of the chimeric virus rBRSV-HN, with the BRSV G protein replaced by BPIV-3 HN, was successful. The replication rates of both chimeras were similar to that of standard rBRSV. Moreover, rBRSV-HNF was neutralized by antibodies specific for BPIV-3, but not by antibodies specific to BRSV, demonstrating that the BRSV glycoproteins can be functionally replaced by BPIV-3 glycoproteins. In contrast, rBRSV-HN was neutralized by BRSV-specific antisera, but not by BPIV-3 specific sera, showing that infection of rBRSV-HN is mediated by BRSV F. Hemadsorption of cells infected with rBRSV-HNF and rBRSV-HN proved that BPIV-3 HN protein expressed by rBRSV is functional. Colocalization of the BPIV-3 glycoproteins with BRSV M protein was demonstrated by confocal laser scan microscopy. Moreover, protein analysis revealed that the BPIV-3 glycoproteins were present in chimeric virions. Taken together, these data indicate that the heterologous glycoproteins were not only expressed but were incorporated into the envelope of recombinant BRSV. Thus, the envelope glycoproteins derived from a member of the Respirovirus genus can together functionally replace their homologs in a Pneumovirus background.

21 CFR 866.5425 - Alpha-2-glycoproteins immunological test system.

Code of Federal Regulations, 2010 CFR

2010-04-01

... the alpha-2-glycoproteins (a group of plasma proteins found in the alpha-2 group when subjected to... some cancers and genetically inherited deficiencies of these plasma proteins. (b) Classification. Class...

21 CFR 866.5425 - Alpha-2-glycoproteins immunological test system.

Code of Federal Regulations, 2012 CFR

2012-04-01

... the alpha-2-glycoproteins (a group of plasma proteins found in the alpha-2 group when subjected to... some cancers and genetically inherited deficiencies of these plasma proteins. (b) Classification. Class...

21 CFR 866.5425 - Alpha-2-glycoproteins immunological test system.

Code of Federal Regulations, 2011 CFR

2011-04-01

... the alpha-2-glycoproteins (a group of plasma proteins found in the alpha-2 group when subjected to... some cancers and genetically inherited deficiencies of these plasma proteins. (b) Classification. Class...

21 CFR 866.5425 - Alpha-2-glycoproteins immunological test system.

Code of Federal Regulations, 2013 CFR

2013-04-01

... the alpha-2-glycoproteins (a group of plasma proteins found in the alpha-2 group when subjected to... some cancers and genetically inherited deficiencies of these plasma proteins. (b) Classification. Class...

21 CFR 866.5425 - Alpha-2-glycoproteins immunological test system.

Code of Federal Regulations, 2014 CFR

2014-04-01

... the alpha-2-glycoproteins (a group of plasma proteins found in the alpha-2 group when subjected to... some cancers and genetically inherited deficiencies of these plasma proteins. (b) Classification. Class...

Induction by carrot allelochemicals of insecticide-metabolising enzymes in the southern armyworm (Spodoptera eridania).

PubMed

Brattsten, L B; Evans, C K; Bonetti, S; Zalkow, L H

1984-01-01

Carrot foliage monoterpenes induce cytochrome P-450 up to 2.9-fold, NADPH cytochrome c (P-450) reductase up to 1.6-fold, NADPH-oxidation up to 3.8-fold, aldrin epoxidation up to 1.5-fold in southern armyworm larval midgut tissues when incorporated in their diet at 0.2% for 3 days. Stigmasterol and ergosterol did not substantially induce microsomal oxidase activities and significantly inhibited GSH S-aryltransferase activity and sulfotransferase activity. Coumarin did not substantially affect microsomal oxidase and sulfotransferase activity but is the most potent inducer of GSH S-aryltransferase activity, increasing this activity 7-fold. None of the chemicals is acutely toxic to the sixth instar larvae or affect the larval weight gain except coumarin which significantly depressed the maximal body weight attained.

Immunoinformatic Analysis of Crimean Congo Hemorrhagic Fever Virus Glycoproteins and Epitope Prediction for Synthetic Peptide Vaccine.

PubMed

Tipu, Hamid Nawaz

2016-02-01

To determine the Crimean Congo Hemorrhagic Fever (CCHF) virus M segement glycoprotein's immunoinformatic parameters, and identify Human Leukocyte Antigen (HLA) class I binders as candidates for synthetic peptide vaccines. Cross-sectional study. Combined Military Hospital, Khuzdar Cantt, in May 2015. Data acquisition, antigenicity prediction, secondary and tertiary structure prediction, residue analysis were done using immunoinformatics tools. HLAclass I binders in glycoprotein's sequence were identified at nanomer length using NetMHC 3.4 and mapped onto tertiary structure. Docking was done for strongest binder against its corresponding allele with CABS-dock. HLAA*0101, 0201, 0301, 2402, 2601 and B*0702, 0801, 2705, 3901, 4001, 5801, 1501 were analyzed against two glycoprotein components of the virus. Atotal of 35 nanomers from GP1, and 3 from GP2 were identified. HLAB*0702 bound maximum number of peptides (6), while HLAB*4001 showed strongest binding affinity. HLAspecific glycoproteins epitope prediction can help identify synthetic peptide vaccine candidates.

Application of Physiologically Based Pharmacokinetic Modeling in Understanding Bosutinib Drug-Drug Interactions: Importance of Intestinal P-Glycoprotein.

PubMed

Yamazaki, Shinji; Loi, Cho-Ming; Kimoto, Emi; Costales, Chester; Varma, Manthena V

2018-05-08

Bosutinib is an orally available Src/Abl tyrosine kinase inhibitor indicated for the treatment of patients with Ph+ chronic myelogenous leukemia at a clinically recommended dose of 500 mg once daily. Clinical results indicated that increases in bosutinib oral exposures were supra-proportional at the lower doses (50 to 200 mg) and approximately dose-proportional at the higher doses (200 to 600 mg). Bosutinib is a substrate of CYP3A4 and P-glycoprotein and exhibits pH-dependent solubility with moderate intestinal permeability. These findings led us to investigate the factors influencing the underlying pharmacokinetic mechanisms of bosutinib with physiologically-based pharmacokinetic (PBPK) models. Our primary objectives were to: 1) refine the previously developed bosutinib PBPK model based on the latest oral bioavailability data and 2) verify the refined PBPK model with P-glycoprotein kinetics based on the bosutinib drug-drug interaction (DDI) results with ketoconazole and rifampin. Additionally, the verified PBPK model was applied to predict bosutinib DDIs with dual CYP3A/P-glycoprotein inhibitors. The results indicated that 1) the refined PBPK model adequately described the observed plasma concentration-time profiles of bosutinib and 2) the verified PBPK model reasonably predicted the effects of ketoconazole and rifampin on bosutinib exposures by accounting for intestinal P-gp inhibition/induction. These results suggested that bosutinib DDI mechanism could involve not only CYP3A4-mediated metabolism but also P-glycoprotein-mediated efflux on absorption. In summary, P-glycoprotein kinetics could constitute a critical element in the PBPK models to understand the pharmacokinetic mechanism of dual CYP3A/P-glycoprotein substrates such as bosutinib exhibiting nonlinear pharmacokinetics due largely to a saturation of intestinal P-glycoprotein-mediated efflux. The American Society for Pharmacology and Experimental Therapeutics.

A hybrid monolithic column based on boronate-functionalized graphene oxide nanosheets for online specific enrichment of glycoproteins.

PubMed

Zhou, Chanyuan; Chen, Xiaoman; Du, Zhuo; Li, Gongke; Xiao, Xiaohua; Cai, Zongwei

2017-05-19

A hybrid monolithic column based on aminophenylboronic acid (APBA)-functionalized graphene oxide (GO) has been developed and used for selective enrichment of glycoproteins. The APBA/GO composites were homogeneously incorporated into a polymer monolithic column with the help of oligomer matrix and followed by in situ polymerization. The effect of dispersion of APBA/GO composites in the polymerization mixture on the performance of the monolithic column was explored in detail. The presence of graphene oxide not only enlarged the BET surface area from 6.3m 2 /g to 169.4m 2 /g, but also provided abundant boronic acid moieties for glycoprotein extraction, which improved the enrichment selectivity and efficiency for glycoproteins. The APBA/GO hybrid monolithic column was incorporated into a sequential injection system, which facilitated online extraction of proteins. Combining the superior properties of extraordinary surface area of GO and the affinity interaction of APBA to glycoproteins, the APBA/GO hybrid monolithic column showed higher enrichment factors for glycoproteins than other proteins without cis-diol-containing groups. Also, under comparable or even shorter processing time and without the addition of any organic solvent, it showed higher binding capacity toward glycoproteins compared with the conventional boronate affinity monolithic column. The practical applicability of this system was demonstrated by processing of egg white samples for extraction of ovalbumin and ovotransferrin, and satisfactory results were obtained by assay with SDS-PAGE. Copyright © 2017 Elsevier B.V. All rights reserved.

[Enrichment of glycoproteins in human serum using concanavalin A-functionalized magnetic nanoparticles and identification by mass spectrometry].

PubMed

Li, Feng; Kang, Jingwu

2014-04-01

Biomedical sciences, and in particular biomarker research, demand efficient glycoprotein enrichment platforms. Herein novel magnetic nanoparticles with an average size around 135 nm in diameter were prepared for the enrichment of glycoproteins in human serum. The prepared magnetic nanoparticles possessed uniform core/shell/shell structure which was composed of 8 nm magnetite internal core and double layers consisting of silica and poly glycidyl methacrylate (GMA). The latter was constructed by seed polymerization. Modified by a polyethylene hydrophilic linker, it made the surfaces of the magnetic nanoparticles highly hydrophilic so as to reduce the nonspecific adsorption of proteins. We examined affinity purification of glycoprotein in diluted human serum using our prepared magnetic nanoparticles with immobilization of concanavalin A (MNP @ ConA). The enriched proteins were reduced, alkylated and digested with trypsin. These peptides then were separated by offline two-dimensional chromatography. Protein identification was realized with nano-high performance liquid chromatography-orbitrap mass spectrometry. A total of 80 proteins were identified, among them 76 proteins were found to be glycoproteins by use of bioinformatic tools. /3-2-Glycoprotein 1 present in serum at low mass concentration around 0.000 01 g/L was also identified. This demonstrates the capability of magnetic nanoparticle for recovering minute amounts of glycoproteins from a fluid exhibiting a dynamic concentration range more than 12 orders of magnitude. Overall, MNP @ ConA has been proven to be an efficient alternative to currently available immobilization supports.

Crystal Structure of the Pre-fusion Nipah Virus Fusion Glycoprotein Reveals a Novel Hexamer-of-Trimers Assembly.

PubMed

Xu, Kai; Chan, Yee-Peng; Bradel-Tretheway, Birgit; Akyol-Ataman, Zeynep; Zhu, Yongqun; Dutta, Somnath; Yan, Lianying; Feng, YanRu; Wang, Lin-Fa; Skiniotis, Georgios; Lee, Benhur; Zhou, Z Hong; Broder, Christopher C; Aguilar, Hector C; Nikolov, Dimitar B

2015-12-01

Nipah virus (NiV) is a paramyxovirus that infects host cells through the coordinated efforts of two envelope glycoproteins. The G glycoprotein attaches to cell receptors, triggering the fusion (F) glycoprotein to execute membrane fusion. Here we report the first crystal structure of the pre-fusion form of the NiV-F glycoprotein ectodomain. Interestingly this structure also revealed a hexamer-of-trimers encircling a central axis. Electron tomography of Nipah virus-like particles supported the hexameric pre-fusion model, and biochemical analyses supported the hexamer-of-trimers F assembly in solution. Importantly, structure-assisted site-directed mutagenesis of the interfaces between F trimers highlighted the functional relevance of the hexameric assembly. Shown here, in both cell-cell fusion and virus-cell fusion systems, our results suggested that this hexamer-of-trimers assembly was important during fusion pore formation. We propose that this assembly would stabilize the pre-fusion F conformation prior to cell attachment and facilitate the coordinated transition to a post-fusion conformation of all six F trimers upon triggering of a single trimer. Together, our data reveal a novel and functional pre-fusion architecture of a paramyxoviral fusion glycoprotein.

Transport characteristics of mammalian Rh and Rh glycoproteins expressed in heterologous systems.

PubMed

Westhoff, C M; Wylie, D E

2006-01-01

The development and use of heterologous expression systems is critical for deciphering the function of mammalian Rh and Rh-glycoproteins. The studies here use Xenopus oocytes, well known for their ability to readily traffic and express difficult membrane proteins, and S. cerevisiae wild-type strains and mutants that are defective in ammonium transport. Data obtained in both of these expression systems revealed that mammalian Rh-glycoprotein-mediated transport (RhAG, RhBG, and RhCG) is an electroneutral process that is driven by the NH4+ concentration and the transmembrane H+ gradient, effectively exchanging NH4+ for H+ in a process that results in transport of net NH3. Homology modeling and functional studies suggest that the more recently evolved erythrocyte blood group proteins, RhCE and RhD, may not function directly in ammonia transport and may be evolving a new function in the RBC membrane. The relationship of Rh and Rh-glycoproteins to the Amt/Mep ammonium transporters is substantiated with functional transport data and structural modeling.

Cytoplasmic tail domain of glycoprotein B is essential for HHV-6 infection

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

Mahmoud, Nora F.; Faculty of Pharmacy, Suez Canal University, Ismailia; Jasirwan, Chyntia

2016-03-15

Human herpesvirus 6 (HHV-6) glycoprotein B (gB) is an abundantly expressed viral glycoprotein required for viral entry and cell fusion, and is highly conserved among herpesviruses. The present study examined the function of HHV-6 gB cytoplasmic tail domain (CTD). A gB CTD deletion mutant was constructed which, in contrast to its revertant, could not be reconstituted. Moreover, deletion of gB cytoplasmic tail impaired the intracellular transport of gB protein to the trans-Golgi network (TGN). Taken together, these results suggest that gB CTD is critical for HHV-6 propagation and important for intracellular transportation. - Highlights: • Glycoprotein B (gB) is highlymore » conserved among herpesviruses. • HHV-6 gB is also abundantly expressed in virions. • In the present study, we showed the function of HHV-6 gB cytoplasmic tail domain (CTD). • We found that deletion of gB CTD impairs the intracellular transport of gB protein to the trans-Golgi network (TGN), and CTD of gB is critical for HHV-6 propagation.« less

Glycoprotein hormone receptors: determinants in leucine-rich repeats responsible for ligand specificity

PubMed Central

Smits, Guillaume; Campillo, Mercedes; Govaerts, Cédric; Janssens, Véronique; Richter, Christine; Vassart, Gilbert; Pardo, Leonardo; Costagliola, Sabine

2003-01-01

Glycoprotein hormone receptors [thyrotropin (TSHr), luteinizing hormone/chorionic gonadotropin (LH/CGr), follicle stimulating hormone (FSHr)] are rhodopsin-like G protein-coupled receptors with a large extracellular N-terminal portion responsible for hormone recognition and binding. In structural models, this ectodomain is composed of two cysteine clusters flanking nine leucine-rich repeats (LRRs). The LRRs form a succession of β-strands and α-helices organized into a horseshoe-shaped structure. It has been proposed that glycoprotein hormones interact with residues of the β-strands making the concave surface of the horseshoe. Gain-of-function homology scanning of the β-strands of glycoprotein hormone receptors allowed identification of the critical residues responsible for the specificity towards human chorionic gonadotropin (hCG). Substitution of eight or two residues of the LH/CGr into the TSHr or FSHr, respectively, resulted in constructs displaying almost the same affinity and sensitivity for hCG as wild-type LH/CGr. Molecular dynamics simulations and additional site-directed mutagenesis provided a structural rationale for the evolution of binding specificity in this duplicated gene family. PMID:12773385

Consumer behaviour towards vegetables: a study on domestic processing of broccoli and carrots by Dutch households.

PubMed

Bongoni, R; Verkerk, R; Dekker, M; Steenbekkers, L P A

2015-06-01

Preferences for sensory properties (e.g. taste and texture) are assumed to control cooking behaviour with respect to vegetables. Conditions such as the cooking method, amount of water used and the time-temperature profile determine the nutritional quality (e.g. vitamins and phytochemicals) of cooked vegetables. Information on domestic processing and any underlying motives can be used to inform consumers about cooking vegetables that are equally liked and are nutrient-rich. Two online self-reporting questionnaires were used to identify domestic processing conditions of broccoli and carrots by Dutch households. Questions on various aspects of domestic processing and consumer motives were included. Descriptive data analysis and hierarchical cluster analysis were performed for both vegetables, separately, to group consumers with similar motives and behaviour towards vegetables. Approximately 70% of consumers boiled vegetables, 8-9% steamed vegetables, 10-15% stir fried raw vegetables and 8-10% stir fried boiled vegetables. Mainly texture was used as a way to decide the 'doneness' of the vegetables. For both vegetables, three clusters of consumers were identified: texture-orientated, health-orientated, or taste-orientated. The texture-orientated consumers are identified as the most prevalent (56-59%) group in the present study. Statistically significant associations are found between domestic processing conditions and clusters, whereas no such association are found between demographic details and clusters. A wide variation in domestic processing of broccoli and carrots is found in the present study. Mainly sensory properties (i.e. texture and taste) determined the domestic processing conditions. The findings of the present study can be used to optimise cooking to yield vegetables that meet consumer's specific sensory preference and are higher in nutrients, and as well as to communicate with target consumer groups. © 2014 The British Dietetic Association Ltd.

Glycosylation and intracellular transport of membrane glycoproteins encoded by murine leukemia viruses. Inhibition by amino acid analogues and by tunicamycin.

PubMed

Polonoff, E; Machida, C A; Kabat, D

1982-12-10

Addition of asparagine-linked oligosaccharides to nascent murine leukemia virus (MuLV)-encoded membrane glycoproteins was inhibited either completely by tunicamycin or specifically at Asn-X-Thr glycosylation sites by incorporation of the threonine analogue beta-hydroxynorvaline. In conditions of partial analogue substitution, a series of subglycosylated components is formed which are related by a constant apparent Mr difference when assayed by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. The total number of asparagine-linked oligosaccharides is then estimated by dividing the measured apparent Mr of one oligosaccharide into the total apparent Mr difference between the complete glycoprotein and the polypeptide chain that is synthesized in cells incubated with tunicamycin. Correct results were obtained using glycoproteins with known numbers of oligosaccharides. Our analyses indicate that the gp70 membrane envelope glycoproteins of certain ecotropic MuLVs contain seven oligosaccharides, whereas the GIX+ antigen-containing variant gp70 contains one fewer Asn-X-Thr-linked oligosaccharide. The membrane glycoprotein encoded by the gag gene of Friend MuLV contains only one asparagine-linked oligosaccharide. Similarly, the gp55 membrane glycoprotein encoded by Friend erythroleukemia virus contains four asparagine-linked oligosaccharides. Pulse-chase and cell surface iodination analyses indicate that MuLV membrane envelope glycoprotein processing by partial proteolysis and transport to the cell surface can be efficiently blocked by structural perturbations caused by incorporation of different amino acid analogues or by loss of oligosaccharides. Our data also suggest that loss of oligosaccharides may expose new antigenic sites in viral membrane glycoproteins and increase their susceptibility to intracellular proteolysis.

Enhancement of Ebola Virus Infection via Ficolin-1 Interaction with the Mucin Domain of GP Glycoprotein.

PubMed

Favier, Anne-Laure; Gout, Evelyne; Reynard, Olivier; Ferraris, Olivier; Kleman, Jean-Philippe; Volchkov, Viktor; Peyrefitte, Christophe; Thielens, Nicole M

2016-06-01

Ebola virus infection requires the surface viral glycoprotein to initiate entry into the target cells. The trimeric glycoprotein is a highly glycosylated viral protein which has been shown to interact with host C-type lectin receptors and the soluble complement recognition protein mannose-binding lectin, thereby enhancing viral infection. Similarly to mannose-binding lectin, ficolins are soluble effectors of the innate immune system that recognize particular glycans at the pathogen surface. In this study, we demonstrate that ficolin-1 interacts with the Zaire Ebola virus (EBOV) glycoprotein, and we characterized this interaction by surface plasmon resonance spectroscopy. Ficolin-1 was shown to bind to the viral glycoprotein with a high affinity. This interaction was mediated by the fibrinogen-like recognition domain of ficolin-1 and the mucin-like domain of the viral glycoprotein. Using a ficolin-1 control mutant devoid of sialic acid-binding capacity, we identified sialylated moieties of the mucin domain to be potential ligands on the glycoprotein. In cell culture, using both pseudotyped viruses and EBOV, ficolin-1 was shown to enhance EBOV infection independently of the serum complement. We also observed that ficolin-1 enhanced EBOV infection on human monocyte-derived macrophages, described to be major viral target cells,. Competition experiments suggested that although ficolin-1 and mannose-binding lectin recognized different carbohydrate moieties on the EBOV glycoprotein, the observed enhancement of the infection likely depended on a common cellular receptor/partner. In conclusion, ficolin-1 could provide an alternative receptor-mediated mechanism for enhancing EBOV infection, thereby contributing to viral subversion of the host innate immune system. A specific interaction involving ficolin-1 (M-ficolin), a soluble effector of the innate immune response, and the glycoprotein (GP) of EBOV was identified. Ficolin-1 enhanced virus infection instead of tipping the

Association between MDR1 gene of gastrointestinal tumors, the expression of P-glycoprotein and resistance to chemotherapeutic drugs.

PubMed

Su, Jian-Li; Wang, Cheng-Hong; Kang, Hong-Gang; Zhang, Jing; Wang, Bao-Zhong; Liu, Mei-Rong; Zhao, Jun; Liu, Lin

2017-09-01

The aim of the present study was to examine and discuss the association between multidrug resistance 1 gene ( MDR1 ) of gastrointestinal tumors, the expression of P-glycoprotein and resistance to chemotherapeutic drugs. In this study, 126 cases of patients with gastrointestinal tumors admitted to hospital from February 2013 to February 2015 were selected. The expression levels of MDR1 gene were obsreved in the control population and patients before and after treatment by fluoresecent quantitative PCR. The protein expression level of P-glycoprotein was determined using western blotting and enzyme-linked immunosorbent assay. In addition, drug resistance was assessed by ATP-TCA chemosensitivity experiments. The results showed that before treatment, the expression of mRNA in MDR1 of tissues of gastrointestinal tract of the 126 cases was 108-fold larger than that of the gastrointestinal tract of the controls (p<0.05), P-glycoprotein was 87-fold larger than the expression level of the controls (p<0.05). The sensitivity of 126 tumor tissues to different chemotherapeutic drugs was determined, and the results showed that most of the tumor tissues were sensitive to chemotherapeutic drugs, and the sensitivity rate reached 96.4%. Following chemotherapy, the expression of mRNA in MDR1 of tumor tissues and the expression of P-glycoprotein decreased (p<0.05). In conclusion, the MDR1 gene and P-glycoprotein have a positive correlation with the occurrence of gastrointestinal tumors, and a negative correlation between the MDR1 gene and P-glycoprotein with resistance of chemotherapeutic drugs. Therefore, the MDR1 gene and P-glycoprotein can be used as references in the identification and diagnosis of gastrointestinal tumors.

The Lyssavirus glycoprotein: A key to cross-immunity.

PubMed

Buthelezi, Sindisiwe G; Dirr, Heini W; Chakauya, Ereck; Chikwamba, Rachel; Martens, Lennart; Tsekoa, Tsepo L; Stoychev, Stoyan H; Vandermarliere, Elien

2016-11-01

Rabies is an acute viral encephalomyelitis in warm-blooded vertebrates, caused by viruses belonging to Rhabdovirus family and genus Lyssavirus. Although rabies is categorised as a neglected disease, the rabies virus (RABV) is the most studied amongst Lyssaviruses which show nearly identical infection patterns. In efforts to improving post-exposure prophylaxis, several anti-rabies monoclonal antibodies (mAbs) targeting the glycoprotein (G protein) sites I, II, III and G5 have been characterized. To explore cross-neutralization capacity of available mAbs and discover new possible B-cell epitopes, we have analyzed all available glycoprotein sequences from Lyssaviruses with a focus on sequence variation and conservation. This information was mapped on the structure of a representative G protein. We proposed several possible cross-neutralizing B-cell epitopes (GUVTTTF, WLRTV, REECLD and EHLVVEEL) in complement to the already well-characterized antigenic sites. The research could facilitate development of novel cross-reactive mAbs against RABV and even more broad, against possibly all Lyssavirus members. Copyright © 2016 Elsevier Inc. All rights reserved.

Isolement et caractérisation de deux subunites constitutives des glycoproteines de structure du tissu sous cutané de lapin.

PubMed

Randoux, A; CornilletStoupy, J; Desanti, M; Borel, J P

1976-09-28

Structural glycoproteins have been extracted by 8 M ureau from the insoluble residue remaining after collagenase digestion of rabbit dermis and purified by Sepharose 4 B chromatography. After reduction and alkylation, Dowex 1 x 2 chromatography allowed separation of two structural glycoproteins (D1 and D2) in an homogenous state as shown by chromatographic and electrophoretic behaviour as well as N terminal amino acid determination. These two glycoproteins have a molecular weight of about 16 000. Their amino acid compositions (very similar), are characterized by a high level of dicarboxylic amino acids and the absence of hydroxyproline and hydroxylysine. The less acidic glycoprotein (D1) has glycine for N terminal amino acid and contains 10.4 percent of bound carbohydrates. The glycoprotein D2 contains 5.1 percent of bound carbohydrates and its N terminal amino acid is glutamic acid.

Identifying the Viral Genes Encoding Envelope Glycoproteins for Differentiation of Cyprinid herpesvirus 3 Isolates

PubMed Central

Han, Jee Eun; Kim, Ji Hyung; Renault, Tristan; Choresca, Casiano; Shin, Sang Phil; Jun, Jin Woo; Park, Se Chang

2013-01-01

Cyprinid herpes virus 3 (CyHV-3) diseases have been reported around the world and are associated with high mortalities of koi (Cyprinus carpio). Although little work has been conducted on the molecular analysis of this virus, glycoprotein genes identified in the present study seem to be valuable targets for genetic comparison of this virus. Three envelope glycoprotein genes (ORF25, 65 and 116) of the CyHV-3 isolates from the USA, Israel, Japan and Korea were compared, and interestingly, sequence insertions or deletions were observed in these target regions. In addition, polymorphisms were presented in microsatellite zones from two glycoprotein genes (ORF65 and 116). In phylogenetic tree analysis, the Korean isolate was remarkably distinguished from USA, Israel, Japan isolates. These findings may be suitable for many applications including isolates differentiation and phylogeny studies. PMID:23435236

Identifying the viral genes encoding envelope glycoproteins for differentiation of Cyprinid herpesvirus 3 isolates.

PubMed

Han, Jee Eun; Kim, Ji Hyung; Renault, Tristan; Choresca, Casiano; Shin, Sang Phil; Jun, Jin Woo; Park, Se Chang

2013-01-31

Cyprinid herpes virus 3 (CyHV-3) diseases have been reported around the world and are associated with high mortalities of koi (Cyprinus carpio). Although little work has been conducted on the molecular analysis of this virus, glycoprotein genes identified in the present study seem to be valuable targets for genetic comparison of this virus. Three envelope glycoprotein genes (ORF25, 65 and 116) of the CyHV-3 isolates from the USA, Israel, Japan and Korea were compared, and interestingly, sequence insertions or deletions were observed in these target regions. In addition, polymorphisms were presented in microsatellite zones from two glycoprotein genes (ORF65 and 116). In phylogenetic tree analysis, the Korean isolate was remarkably distinguished from USA, Israel, Japan isolates. These findings may be suitable for many applications including isolates differentiation and phylogeny studies.

Crucial steps in the structure determination of a coronavirus spike glycoprotein using cryo‐electron microscopy

PubMed Central

Walls, Alexandra; Tortorici, M. Alejandra; Bosch, Berend‐Jan; Frenz, Brandon; Rottier, Peter J. M.; DiMaio, Frank; Rey, Felix A.

2016-01-01

Abstract The tremendous pandemic potential of coronaviruses was demonstrated twice in the last 15 years by two global outbreaks of deadly pneumonia. Entry of coronaviruses into cells is mediated by the transmembrane spike glycoprotein S, which forms a trimer carrying receptor‐binding and membrane fusion functions. Despite their biomedical importance, coronavirus S glycoproteins have proven difficult targets for structural characterization, precluding high‐resolution studies of the biologically relevant trimer. Recent technological developments in single particle cryo‐electron microscopy allowed us to determine the first structure of a coronavirus S glycoprotein trimer which provided a framework to understand the mechanisms of viral entry and suggested potential inhibition strategies for this family of viruses. Here, we describe the key factors that enabled this breakthrough. PMID:27667334

Sensitivity of Aspergillus nidulans to the Cellulose Synthase Inhibitor Dichlobenil: Insights from Wall-Related Genes’ Expression and Ultrastructural Hyphal Morphologies

PubMed Central

Obersriebnig, Michael; Salerno, Marco; Pum, Dietmar; Strauss, Joseph

2013-01-01

The fungal cell wall constitutes an important target for the development of antifungal drugs, because of its central role in morphogenesis, development and determination of fungal-specific molecular features. Fungal walls are characterized by a network of interconnected glycoproteins and polysaccharides, namely α-, β-glucans and chitin. Cell walls promptly and dynamically respond to environmental stimuli by a signaling mechanism, which triggers, among other responses, modulations in wall biosynthetic genes’ expression. Despite the absence of cellulose in the wall of the model filamentous fungus Aspergillus nidulans, we found in this study that fungal growth, spore germination and morphology are affected by the addition of the cellulose synthase inhibitor dichlobenil. Expression analysis of selected genes putatively involved in cell wall biosynthesis, carried out at different time points of drug exposure (i.e. 0, 1, 3, 6 and 24 h), revealed increased expression for the putative mixed linkage β-1,3;1,4 glucan synthase celA together with the β-1,3-glucan synthase fksA and the Rho-related GTPase rhoA. We also compared these data with the response to Congo Red, a known plant/fungal drug affecting both chitin and cellulose biosynthesis. The two drugs exerted different effects at the cell wall level, as shown by gene expression analysis and the ultrastructural features observed through atomic force microscopy and scanning electron microscopy. Although the concentration of dichlobenil required to affect growth of A. nidulans is approximately 10-fold higher than that required to inhibit plant cellulose biosynthesis, our work for the first time demonstrates that a cellulose biosynthesis inhibitor affects fungal growth, changes fungal morphology and expression of genes connected to fungal cell wall biosynthesis. PMID:24312197

Carryover of perfluorooctanoic acid (PFOA) and perfluorooctane sulfonate (PFOS) from soil to plant and distribution to the different plant compartments studied in cultures of carrots (Daucus carota ssp. Sativus), potatoes (Solanum tuberosum), and cucumbers (Cucumis Sativus).

PubMed

Lechner, Mareike; Knapp, Holger

2011-10-26

A vegetation study was carried out to investigate the carryover of Perfluorooctanoic Acid (PFOA) and Perfluorooctane Sulfonate (PFOS) from soil mixed with contaminated sewage sludge to potato, carrot, and cucumber plants. Analysis was done by liquid-extraction using acetonitrile with dispersive SPE cleanup and subsequent HPLC-MS/MS. In order to assess the transfer potential from soil, transfer factors (TF) were calculated for the different plant compartments: TF = [PFC](plant (wet substance))/[PFC](soil (dry weight)). The highest TF were found for the vegetative plant compartments with average values for PFOS below those for PFOA: cucumber, 0.17 (PFOS), 0.88 (PFOA); potato, 0.36 (PFOS), 0.40 (PFOA); carrot, 0.38 (PFOS), 0.53 (PFOA). Transfer of PFOA and PFOS into potato peelings (average values of TF: PFOA 0.03, PFOS 0.04) exceeded the carryover to the peeled tubers (PFOA 0.01, PFOS < 0.01). In carrots, this difference did not occur (average values of TF: PFOA 0.04, PFOS 0.04). Transfer of PFOS into the unpeeled cucumbers was low and comparable to that of peeled potatoes (TF < 0.01). For PFOA, it was higher (TF: 0.03).

LC–MS/MS Quantitation of Esophagus Disease Blood Serum Glycoproteins by Enrichment with Hydrazide Chemistry and Lectin Affinity Chromatography

PubMed Central

2015-01-01

Changes in glycosylation have been shown to have a profound correlation with development/malignancy in many cancer types. Currently, two major enrichment techniques have been widely applied in glycoproteomics, namely, lectin affinity chromatography (LAC)-based and hydrazide chemistry (HC)-based enrichments. Here we report the LC–MS/MS quantitative analyses of human blood serum glycoproteins and glycopeptides associated with esophageal diseases by LAC- and HC-based enrichment. The separate and complementary qualitative and quantitative data analyses of protein glycosylation were performed using both enrichment techniques. Chemometric and statistical evaluations, PCA plots, or ANOVA test, respectively, were employed to determine and confirm candidate cancer-associated glycoprotein/glycopeptide biomarkers. Out of 139, 59 common glycoproteins (42% overlap) were observed in both enrichment techniques. This overlap is very similar to previously published studies. The quantitation and evaluation of significantly changed glycoproteins/glycopeptides are complementary between LAC and HC enrichments. LC–ESI–MS/MS analyses indicated that 7 glycoproteins enriched by LAC and 11 glycoproteins enriched by HC showed significantly different abundances between disease-free and disease cohorts. Multiple reaction monitoring quantitation resulted in 13 glycopeptides by LAC enrichment and 10 glycosylation sites by HC enrichment to be statistically different among disease cohorts. PMID:25134008

Efficacy of soluble glycoprotein fraction from Allium sativum purified by size exclusion chromatography on murine Schistosomiasis mansoni.

PubMed

Aly, Ibrahim; Taher, Eman E; El-Sayed, Hoda; Mohammed, Faten A; ELnain, Gehan; Hamad, Rabab S; Bayoumy, Elsayed M

2017-06-01

In this work, the efficiency of crude MeOH extracts and soluble glycoprotein fraction of Allium sativum purified by size-exclusion chromatography (SEC) on parasitological, histopathological and some biochemical parameters in Schistosoma mansoni infected mice were investigated. Animals were infected by tail immersion with 100 cercariae/each mouse and divided into five groups in addition to the normal control. The results revealed a significant decrease in mean worm burden in all treated mice especially in the group treated with soluble glycoprotein fraction of A. sativum as compared to infected non-treated control with the disappearance of female worms. Administration of the studied extracts revealed remarkable amelioration in the levels of all the measured parameters in S. mansoni infected mice. In addition, treatment of mice with crude A. sativum MeOH extract and soluble glycoprotein fraction of A. sativum decreased significantly the activities of studied enzymes as compared to the infected untreated group. The highest degrees of enhancement in pathological changes was observed in the treated one with soluble glycoprotein fraction of A. sativum compared to the infected group represented by small sized, late fibro-cellular granuloma, the decrease in cellular constituents and degenerative changes in eggs. In conclusion, A. sativum treatment had effective schistosomicidal activities, through reduction of worm burden and tissue eggs, especially when it was given in purified glycoprotein fraction. Moreover, the soluble glycoprotein fraction of A. sativum largely modulates both the size and the number of granulomas. Copyright © 2017 Elsevier Ltd. All rights reserved.

Marked differences in the antigenic structure of human respiratory syncytial virus F and G glycoproteins.

PubMed Central

García-Barreno, B; Palomo, C; Peñas, C; Delgado, T; Perez-Breña, P; Melero, J A

1989-01-01

Monoclonal antibodies directed against the glycoproteins of human respiratory syncytial virus were used in competitive enzyme-linked immunosorbent assays for topological mapping of epitopes. Whereas epitopes of the F glycoprotein could be ascribed to five nonoverlapping antigenic sites, anti-G antibodies recognized unique epitopes, many of whose competition profiles overlapped extensively. Variant viruses selected with a neutralizing (47F) anti-F antibody lost the binding for only 47F and 49F antibodies, which mapped in the same antigenic area. In contrast, viruses selected with an anti-G antibody lost the capacity to bind most of the anti-G antibodies, and their G protein was not recognized by an anti-virus antiserum, indicating major changes in the antigenic structure of the G molecule. Finally, we found great antigenic variation of the G protein among viral isolates. This occurred even within viruses of the same subtype with only limited divergence of amino acid sequence between strains. All of these data indicate marked differences in the antigenic organization of the G and F glycoproteins of respiratory syncytial virus; we discuss these differences in terms of the chemical structure of the glycoproteins. Images PMID:2463385

Decreased levels of alpha-1-acid glycoprotein are related to the mortality of septic patients in the emergency department.

PubMed

Barroso-Sousa, Romualdo; Lobo, Romulo R; Mendonça, Patricia R; Memória, Renan R; Spiller, Fernando; Cunha, Fernando Q; Pazin-Filho, Antonio

2013-01-01

To determine the validity of alpha-1-acid glycoprotein as a novel biomarker for mortality in patients with severe sepsis. We prospectively included patients with severe sepsis or septic shock at the emergency department at a single tertiary referral teaching hospital. All of the patients were enrolled within the first 24 hours of emergency department admission, and clinical data and blood samples were obtained. As the primary outcome, we investigated the association of serum levels of alpha-1-acid glycoprotein and 96-hour mortality with logistic regression analysis and generalized estimating equations adjusted for age, sex, shock status and Acute Physiology and Chronic Health Evaluation II score. Patients with septic shock had lower alpha-1-acid glycoprotein levels at the time of emergency department admission compared to patients without shock (respectively, 149.1 ±42.7 vs. 189.8 ±68.6; p = 0.005). Similarly, non-survivors in the first 96 hours were also characterized by lower levels of alpha-1-acid glycoprotein at the time of emergency department admission compared to survivors (respectively, 132.18 ±50.2 vs. 179.8 ±61.4; p = 0.01). In an adjusted analysis, alpha-1-acid glycoprotein levels ≤120 mg/dL were significantly associated with 96-hour mortality (odds ratio = 14.37; 95% confidence interval = 1.58 to 130.21). Septic shock patients exhibited lower circulating alpha-1-acid glycoprotein levels than patients without shock. Alpha-1-acid glycoprotein levels were independently associated with 96-hour mortality in individuals with severe sepsis.

Curcumin Regulates Colon Cancer by Inhibiting P-Glycoprotein in In-situ Cancerous Colon Perfusion Rat Model.

PubMed

Neerati, Prasad; Sudhakar, Yakkanti A; Kanwar, Jagat R

2013-07-08

Studies on p-glycoprotein was carried out world vide with cell lines like Caco2, MDR1-LLC-PK1 and MDR1-MDCK in-vitro , but most of the results were failed to produce similar results in-vivo. In the present study curcumin inhibitory action on p-glycoprotein increased permeability of irinotecan, so in the colon cancer it would be beneficial if curcumin used as add on therapy. Intra-rectal administered of N-Nitroso N-methyl urea (2 mg/Kg) induced colon cancer. Single pass whole length of colon in-situ perfusion was carried out in rats with irinotecan to study the influence of p-glycoprotein modulators like verapamil and curcumin. The rats were divided in to 5 groups (n=6), Group I served as control perfused with 30 μg/ml of irinotecan, propronolol and phenol red. Group II was cancerous group, induced by N-methyl N-nitroso urea. Group III was perfused with irinotican in cancerous rats. Group IV, perfused with irinotican in presence of verapamil and group V was pre-treated with curcumin and then perfused with irinotican and was estimated by HPLC-UV to effective permeability coefficient. Our qRT-PCR and Western blot results confirmed that about 15-fold decreases in the expression of p-glycoprotein (P-gp) in curcumin treated colon cancer cells. Irinotecan was increased to 0.00066 cm/s and about 11-fold increase in verapamil-coperfused group, where curcumin pre-treated group irinotecan was increases 0.00006 cm/s to 0.00042 cm/s that is about 7-fold increase p-glycoprotein inhibitory activity by verapamil and curcumin found to be significantly enhanced the cancerous colon permeability of irinotecan. Any safe suitable p-glycoprotein inhibitors along with irinotecan will enhance the therapeutic benefit in the treatment of the colon cancer.

Modeling of Oligosaccharides within Glycoproteins from Free-Energy Landscapes

PubMed Central

2017-01-01

In spite of the abundance of glycoproteins in biological processes, relatively little three-dimensional structural data is available for glycan structures. Here, we study the structure and flexibility of the vast majority of mammalian oligosaccharides appearing in N- and O-glycosylated proteins using a bottom up approach. We report the conformational free-energy landscapes of all relevant glycosidic linkages as obtained from local elevation simulations and subsequent umbrella sampling. To the best of our knowledge, this represents the first complete conformational library for the construction of N- and O-glycan structures. Next, we systematically study the effect of neighboring residues, by extensively simulating all relevant trisaccharides and one tetrasaccharide. This allows for an unprecedented comparison of disaccharide linkages in large oligosaccharides. With a small number of exceptions, the conformational preferences in the larger structures are very similar as in the disaccharides. This, finally, allows us to suggest several efficient approaches to construct complete N- and O-glycans on glycoproteins, as exemplified on two relevant examples. PMID:28816453

Radiosensitivity of different tissues from carrot root at different phases of growth in culture

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

Degani, N.; Pickholtz, D.

1980-09-01

The present work compares the effect of ..gamma..-radiation dose and time in culture on the growth of cambium and phloem carrot (Daucus carota) root explants. It was found that the phloem is more radiosensitive than the cambium and that both tissues were more radiosensitive when irradiated on excision at the G/sub 1/ phase rather than at the end of the lag phase on the ninth day of growth in culture when cells were predominantly at the G/sub 2/ phase. The nuclear volumes of cells from both tissues were similar but were larger at the end of the more radioresistant lagmore » phase than those of the G/sub 1/ phase on excision. However, nuclear volume could not account for the differences in radiosensitivity between either the tissues or irradiation times in culture.« less

Mannose-binding lectin binds to Ebola and Marburg envelope glycoproteins, resulting in blocking of virus interaction with DC-SIGN and complement-mediated virus neutralization.

PubMed

Ji, Xin; Olinger, Gene G; Aris, Sheena; Chen, Ying; Gewurz, Henry; Spear, Gregory T

2005-09-01

Mannose-binding lectin (MBL), a serum lectin that mediates innate immune functions including activation of the lectin complement pathway, binds to carbohydrates expressed on some viral glycoproteins. In this study, the ability of MBL to bind to virus particles pseudotyped with Ebola and Marburg envelope glycoproteins was evaluated. Virus particles bearing either Ebola (Zaire strain) or Marburg (Musoke strain) envelope glycoproteins bound at significantly higher levels to immobilized MBL compared with virus particles pseudotyped with vesicular stomatitis virus glycoprotein or with no virus glycoprotein. As observed in previous studies, Ebola-pseudotyped virus bound to cells expressing the lectin DC-SIGN (dendritic cell-specific intercellular adhesion molecule 3-grabbing non-integrin). However, pre-incubation of virus with MBL blocked DC-SIGN-mediated binding to cells, suggesting that the two lectins bind at the same or overlapping sites on the Ebola glycoprotein. Neutralization experiments showed that virus pseudotyped with Ebola or Marburg (Musoke) glycoprotein was neutralized by complement, while the Marburg (Ravn strain) glycoprotein-pseudotyped virus was less sensitive to neutralization. Neutralization was partially mediated through the lectin complement pathway, since a complement source deficient in MBL was significantly less effective at neutralizing viruses pseudotyped with filovirus glycoproteins and addition of purified MBL to the MBL-deficient complement increased neutralization. These experiments demonstrated that MBL binds to filovirus envelope glycoproteins resulting in important biological effects and suggest that MBL can interact with filoviruses during infection in humans.

Evidence for glycoprotein transport into complex plastids.

PubMed

Peschke, Madeleine; Moog, Daniel; Klingl, Andreas; Maier, Uwe G; Hempel, Franziska

2013-06-25

Diatoms are microalgae that possess so-called "complex plastids," which evolved by secondary endosymbiosis and are surrounded by four membranes. Thus, in contrast to primary plastids, which are surrounded by only two membranes, nucleus-encoded proteins of complex plastids face additional barriers, i.e., during evolution, mechanisms had to evolve to transport preproteins across all four membranes. This study reveals that there exist glycoproteins not only in primary but also in complex plastids, making transport issues even more complicated, as most translocation machineries are not believed to be able to transport bulky proteins. We show that plastidal reporter proteins with artificial N-glycosylation sites are indeed glycosylated during transport into the complex plastid of the diatom Phaeodactylum tricornutum. Additionally, we identified five endogenous glycoproteins, which are transported into different compartments of the complex plastid. These proteins get N-glycosylated during transport across the outermost plastid membrane and thereafter are transported across the second, third, and fourth plastid membranes in the case of stromal proteins. The results of this study provide insights into the evolutionary pressure on translocation mechanisms and pose unique questions on the operating mode of well-known transport machineries like the translocons of the outer/inner chloroplast membranes (Toc/Tic).

Effect of genuine non-anthocyanin phenolics and chlorogenic acid on color and stability of black carrot (Daucus carota ssp. sativus var. atrorubens Alef.) anthocyanins.

PubMed

Gras, Claudia C; Bogner, Hanna; Carle, Reinhold; Schweiggert, Ralf M

2016-07-01

This work aimed at studying the color intensity and stability of black carrot anthocyanins as influenced by intermolecular co-pigmentation. For this purpose, purified anthocyanin solutions were supplemented with purified genuine black carrot phenolics, chlorogenic acid, and an aqueous phenolic-rich green coffee bean extract at various anthocyanin:co-pigment ratios (1:0-1:162; pH 3.6). The hyperchromic co-pigmentation effect depended on the concentration of added co-pigments, resulting in an absorbance increase of up to 22% at the absorption maximum. Anthocyanin stability during heating (90°C, 5h) was barely improved unless the concentrations of co-pigments exceeded those of their natural source. When adding co-pigments at ratios above 1:9.4, anthocyanin heat stability was significantly improved. As acylated anthocyanins were most stable, breeders might aim at increasing their content in the future, while breeding for high levels of colorless polyphenols may be unreachable. Nevertheless, we provided proof-of-concept for the successful color enhancement by the addition of a phenolic-rich green coffee bean extract, being useful for food-grade applications. Copyright © 2016 Elsevier Ltd. All rights reserved.

Novel Cross-Reactive Monoclonal Antibodies against Ebolavirus Glycoproteins Show Protection in a Murine Challenge Model.

PubMed

Duehr, James; Wohlbold, Teddy John; Oestereich, Lisa; Chromikova, Veronika; Amanat, Fatima; Rajendran, Madhusudan; Gomez-Medina, Sergio; Mena, Ignacio; tenOever, Benjamin R; García-Sastre, Adolfo; Basler, Christopher F; Munoz-Fontela, Cesar; Krammer, Florian

2017-08-15

Out of an estimated 31,100 cases since their discovery in 1976, ebolaviruses have caused approximately 13,000 deaths. The vast majority (∼11,000) of these occurred during the 2013-2016 West African epidemic. Three out of five species in the genus are known to cause Ebola Virus Disease in humans. Several monoclonal antibodies against the ebolavirus glycoprotein are currently in development as therapeutics. However, there is still a paucity of monoclonal antibodies that can cross-react between the glycoproteins of different ebolavirus species, and the mechanism of these monoclonal antibody therapeutics is still not understood in detail. Here, we generated a panel of eight murine monoclonal antibodies (MAbs) utilizing a prime-boost vaccination regimen with a Zaire ebolavirus glycoprotein expression plasmid followed by infection with a vesicular stomatitis virus expressing the Zaire ebolavirus glycoprotein. We tested the binding breadth of the resulting monoclonal antibodies using a set of recombinant surface glycoproteins from Reston, Taï Forest, Bundibugyo, Zaire, Sudan, and Marburg viruses and found two antibodies that showed pan-ebolavirus binding. An in vivo Stat2 -/- mouse model was utilized to test the ability of these MAbs to protect from infection with a vesicular stomatitis virus expressing the Zaire ebolavirus glycoprotein. Several of our antibodies, including the broadly binding ones, protected mice from mortality despite lacking neutralization capability in vitro , suggesting their protection may be mediated by Fc-FcR interactions. Indeed, three antibodies displayed cellular phagocytosis and/or antibody-dependent cell-mediated cytotoxicity in vitro Our antibodies, specifically the two identified cross-reactive monoclonal antibodies (KL-2E5 and KL-2H7), might add to the understanding of anti-ebolavirus humoral immunity. IMPORTANCE This study describes the generation of a panel of novel anti-ebolavirus glycoprotein monoclonal antibodies, including two

Novel Cross-Reactive Monoclonal Antibodies against Ebolavirus Glycoproteins Show Protection in a Murine Challenge Model

PubMed Central

Duehr, James; Wohlbold, Teddy John; Oestereich, Lisa; Chromikova, Veronika; Amanat, Fatima; Gomez-Medina, Sergio; Mena, Ignacio; tenOever, Benjamin R.; García-Sastre, Adolfo; Basler, Christopher F.

2017-01-01

ABSTRACT Out of an estimated 31,100 cases since their discovery in 1976, ebolaviruses have caused approximately 13,000 deaths. The vast majority (∼11,000) of these occurred during the 2013-2016 West African epidemic. Three out of five species in the genus are known to cause Ebola Virus Disease in humans. Several monoclonal antibodies against the ebolavirus glycoprotein are currently in development as therapeutics. However, there is still a paucity of monoclonal antibodies that can cross-react between the glycoproteins of different ebolavirus species, and the mechanism of these monoclonal antibody therapeutics is still not understood in detail. Here, we generated a panel of eight murine monoclonal antibodies (MAbs) utilizing a prime-boost vaccination regimen with a Zaire ebolavirus glycoprotein expression plasmid followed by infection with a vesicular stomatitis virus expressing the Zaire ebolavirus glycoprotein. We tested the binding breadth of the resulting monoclonal antibodies using a set of recombinant surface glycoproteins from Reston, Taï Forest, Bundibugyo, Zaire, Sudan, and Marburg viruses and found two antibodies that showed pan-ebolavirus binding. An in vivo Stat2−/− mouse model was utilized to test the ability of these MAbs to protect from infection with a vesicular stomatitis virus expressing the Zaire ebolavirus glycoprotein. Several of our antibodies, including the broadly binding ones, protected mice from mortality despite lacking neutralization capability in vitro, suggesting their protection may be mediated by Fc-FcR interactions. Indeed, three antibodies displayed cellular phagocytosis and/or antibody-dependent cell-mediated cytotoxicity in vitro. Our antibodies, specifically the two identified cross-reactive monoclonal antibodies (KL-2E5 and KL-2H7), might add to the understanding of anti-ebolavirus humoral immunity. IMPORTANCE This study describes the generation of a panel of novel anti-ebolavirus glycoprotein monoclonal antibodies

Effect of an aqueous extract of Scoparia dulcis on plasma and tissue glycoproteins in streptozotocin induced diabetic rats.

PubMed

Latha, M; Pari, L

2005-02-01

The influence of Scoparia dulcis, a traditionally used plant for the treatment of diabetes mellitus, was examined in streptozotocin diabetic rats on dearrangement in glycoprotein levels. Diabetes was induced in male Wistar rats by a single intraperitoneal injection of streptozotocin. An aqueous extract of Scoparia dulcis plant was administered orally for 6 weeks. The effect of the Scoparia dulcis extract on blood glucose, plasma insulin, plasma and tissue glycoproteins studied was in comparison to glibenclamide. The levels of blood glucose and plasma glycoproteins were increased significantly whereas the level of plasma insulin was significantly decreased in diabetic rats. There was a significant decrease in the level of sialic acid and elevated levels of hexose, hexosamine and fucose in the liver and kidney of streptozotocin diabetic rats. Oral administration of Scoparia dulcis plant extract (SPEt) to diabetic rats led to decreased levels of blood glucose and plasma glycoproteins. The levels of plasma insulin and tissue sialic acid were increased whereas the levels of tissue hexose, hexosamine and fucose were near normal. The present study indicates that Scoparia dulcis possesses a significant beneficial effect on glycoproteins in addition to its antidiabetic effect.

Role of N-linked polymannose oligosaccharides in targeting glycoproteins for endoplasmic reticulum-associated degradation.