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Sample records for biologically active recombinant

  1. Expression of feline recombinant interferon-gamma in baculovirus and demonstration of biological activity.

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    Argyle, D J; Harris, M; Lawrence, C; McBride, K; Barron, R; McGillivray, C; Onions, D E

    1998-07-08

    We have previously reported the cloning of the coding sequence for feline-specific interferon-gamma. Here, we describe the expression of this sequence in a baculovirus system and demonstrate the biological activity of the recombinant protein. The coding sequence for feline interferon was directionally cloned into the baculovirus transfer vector pAcCL29-1. Transfer vector and linearized wild-type AcMNPV (BacPAK6) were used to co-transfect Sf9 cells by calcium phosphate coprecipitation. Subsequently, wild-type and recombinant viruses were separated by plaque assay. Recombinant plaques were expanded and a master stock of virus is produced. Production of biologically active interferon-gamma from infected Sf9 cells was demonstrated using a standard cytopathic effect reduction assay, utilising vesicular stomatitis virus (VSV), and an MHC class II induction assay.

  2. Refolding techniques for recovering biologically active recombinant proteins from inclusion bodies.

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    Yamaguchi, Hiroshi; Miyazaki, Masaya

    2014-02-20

    Biologically active proteins are useful for studying the biological functions of genes and for the development of therapeutic drugs and biomaterials in a biotechnology industry. Overexpression of recombinant proteins in bacteria, such as Escherichia coli, often results in the formation of inclusion bodies, which are protein aggregates with non-native conformations. As inclusion bodies contain relatively pure and intact proteins, protein refolding is an important process to obtain active recombinant proteins from inclusion bodies. However, conventional refolding methods, such as dialysis and dilution, are time consuming and, often, recovered yields of active proteins are low, and a trial-and-error process is required to achieve success. Recently, several approaches have been reported to refold these aggregated proteins into an active form. The strategies largely aim at reducing protein aggregation during the refolding procedure. This review focuses on protein refolding techniques using chemical additives and laminar flow in microfluidic chips for the efficient recovery of active proteins from inclusion bodies.

  3. Refolding Techniques for Recovering Biologically Active Recombinant Proteins from Inclusion Bodies

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    Hiroshi Yamaguchi

    2014-02-01

    Full Text Available Biologically active proteins are useful for studying the biological functions of genes and for the development of therapeutic drugs and biomaterials in a biotechnology industry. Overexpression of recombinant proteins in bacteria, such as Escherichia coli, often results in the formation of inclusion bodies, which are protein aggregates with non-native conformations. As inclusion bodies contain relatively pure and intact proteins, protein refolding is an important process to obtain active recombinant proteins from inclusion bodies. However, conventional refolding methods, such as dialysis and dilution, are time consuming and, often, recovered yields of active proteins are low, and a trial-and-error process is required to achieve success. Recently, several approaches have been reported to refold these aggregated proteins into an active form. The strategies largely aim at reducing protein aggregation during the refolding procedure. This review focuses on protein refolding techniques using chemical additives and laminar flow in microfluidic chips for the efficient recovery of active proteins from inclusion bodies.

  4. Expression of soluble, biologically active recombinant human endostatin in Escherichia coli.

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    Xu, Han-Mei; Zhang, Guo-Yuan; Ji, Xiao-Dan; Cao, Lin; Shu, Luan; Hua, Zi-Chun

    2005-06-01

    Endostatin, a 20kDa C-terminal fragment of collagen XVIII, is a potent anti-angiogenic protein and inhibitor of tumor growth. Recombinant endostatin was prepared from Escherichia coli deposited as insoluble, inactive inclusion bodies. In the present study, we produced soluble and biologically active recombinant human endostatin (rhEndostatin) in E. coli by employing both co-expression of the molecular chaperones and lower temperature fermentation. Two groups of chaperones Trigger factor and GroEL-GroES (GroEL/ES), DnaK-DnaJ-GrpE and GroEL/ES, were co-expressed, respectively, with rhEndostatin at different temperatures (37, 25, and 16 degrees C). It revealed that low temperature or molecular chaperones alone could enhance the production of active rhEndostatin; meanwhile, combinational employment of low temperature cultivation (16 degrees C) together with co-expression of DnaK-DnaJ-GrpE and GroEL/ES was more effective to prevent aggregation of rhEndostatin. The production of soluble rhEndostatin was about 36 mg/L, and at least 16 mg of rhEndostatin was purified from 1L flask culture. The purified rhEndostatin specifically inhibited the proliferation of endothelial cell-bovine capillary endothelial cell in a dose-dependent manner, and it showed potent anti-angiogenic capability on the chorioallantoic membrane of chick embryo in vivo. Our study provides a feasible and convenient approach to produce soluble and biologically active rhEndostatin.

  5. Generation of biologically active multi-sialylated recombinant human EPOFc in plants.

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    Alexandra Castilho

    Full Text Available Hyperglycosylated proteins are more stable, show increased serum half-life and less sensitivity to proteolysis compared to non-sialylated forms. This applies particularly to recombinant human erythropoietin (rhEPO. Recent progress in N-glycoengineering of non-mammalian expression hosts resulted in in vivo protein sialylation at great homogeneity. However the synthesis of multi-sialylated N-glycans is so far restricted to mammalian cells. Here we used a plant based expression system to accomplish multi-antennary protein sialylation. A human erythropoietin fusion protein (EPOFc was transiently expressed in Nicotiana benthamiana ΔXTFT, a glycosylation mutant that lacks plant specific N-glycan residues. cDNA of the hormone was co-delivered into plants with the necessary genes for (i branching (ii β1,4-galactosylation as well as for the (iii synthesis, transport and transfer of sialic acid. This resulted in the production of recombinant EPOFc carrying bi- tri- and tetra-sialylated complex N-glycans. The formation of this highly complex oligosaccharide structure required the coordinated expression of 11 human proteins acting in different subcellular compartments at different stages of the glycosylation pathway. In vitro receptor binding assays demonstrate the generation of biologically active molecules. We demonstrate the in planta synthesis of one of the most complex mammalian glycoforms pointing to an outstanding high degree of tolerance to changes in the glycosylation pathway in plants.

  6. Purification and characterization of biologically active recombinant human Eppin expressed in Escherichia coli

    Institute of Scientific and Technical Information of China (English)

    ZHU Qing-yi; GU Xiao-jian; YANG Jin; WANG Jun-hong; TANG Bo; WU Hong-fei

    2008-01-01

    Background Eppin(epididymis protease inhibitor)appears to play an important role in primate fertility.However,the function of Eppin and its antibody in men and its relationship with men's infertility are poorly studied.To reveal the significance and possibility of detection of anti-Eppin antibody in clinical infertilty cases,we developed an Escherichia coli expression system for the expression of biologically actire human Eppin.Methods The human Eppin gene was cloned into PET-28a(+)vector after induction with 0.5 mmol/L isopropy-β-D-thiogalactoside(IPTG)at 26℃ for 4 hours,and the expressed fusion protein His6-Eppin was purified by Ni2+ affinity chromatography.Afterwards,six female 8-week-old Balb/c mice were immunized with purified His6-Eppin for three weeks.Their sera were collected and polyclonal antibodies against His6-Eppin were purified,all of which were further verified by Western-blot and immunofluorescence analysis.Results About 18.33 mg His6-Eppin was obtained from 1-L flask culture.The produced polyclonal antibodies against His6-Eppin recognized the Eppin protein both in human epididymis and in HEK293T cells by over-expression of the recombinant human Eppin.Conclusion The purified His6-Eppin protein has biological activity,which might be a candidate for clinical diagnosis of infertility and development of male immuno-contraceptive agents.

  7. Growth hormone from striped catfish (Pangasianodon hypophthalmus): genomic organization, recombinant expression and biological activity.

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    Poen, Sinothai; Pornbanlualap, Somchai

    2013-04-15

    Growth hormone is an essential polypeptide required for normal growth and development of vertebrates. In this report, striped catfish (Pangasianodon hypophthalmus) growth hormone gene and cDNA were isolated by reverse transcriptase-polymerase chain reaction. The striped catfish growth hormone (scGH) encoding gene contains 5 exons and 4 introns. The cDNA sequence of the scGH gene contains a 603bp open reading frame and encodes for a 200-aa protein consisting of a putative 22-aa signal peptide and the mature 178-aa protein. The recombinant histidine-tagged scGH protein which expressed in Escherichia coli as inclusion bodies was unfolded, refolded and purified to near-homogeneity by Ni(2+)-NTA chromatography. Analysis of the secondary structure content by CD spectroscopy showed that the α-helical content of the refolded scGH is 55%. Elucidation of the folding pathway of scGH by fluorescence spectroscopy showed that denaturation transition of scGH is coincident and cooperative, consistent with the two-state denaturation mechanism. The purified scGH was biologically active and exhibited growth-promoting activity in striped catfish, but not tilapia.

  8. Cell biology of mitotic recombination

    DEFF Research Database (Denmark)

    Lisby, Michael; Rothstein, Rodney

    2015-01-01

    Homologous recombination provides high-fidelity DNA repair throughout all domains of life. Live cell fluorescence microscopy offers the opportunity to image individual recombination events in real time providing insight into the in vivo biochemistry of the involved proteins and DNA molecules...... of this review include the stoichiometry and dynamics of recombination complexes in vivo, the choreography of assembly and disassembly of recombination proteins at sites of DNA damage, the mobilization of damaged DNA during homology search, and the functional compartmentalization of the nucleus with respect...... as well as the cellular organization of the process of homologous recombination. Herein we review the cell biological aspects of mitotic homologous recombination with a focus on Saccharomyces cerevisiae and mammalian cells, but will also draw on findings from other experimental systems. Key topics...

  9. Biological Activity of Recombinant Accessory Cholerae Enterotoxin (Ace on Rabbit Ileal Loops and Antibacterial Assay

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    Shaghayegh Anvari

    2012-01-01

    Full Text Available Objective: Vibrio cholerae (V. cholerae causes a potentially lethal disease named cholera. The cholera enterotoxin (CT is a major virulence factor of V. cholerae. In addition to CT, V. cholerae produces other putative toxins, such as the zonula occludens toxin (Zot and accessory cholera enterotoxin (Ace. The ace gene is the third gene of the V. cholerae virulence cassette. The Ace toxin alters ion transport, causes fluid accumulation in ligated rabbit ileal loops, and is a cause of mild diarrhea. The aim of this study is the cloning and overexpression of the ace gene into Escherichia coli (E. coli and determination of some characteristics of the recombinant Ace protein.Materials and Methods: In this experimental study, the ace gene was amplified from V. cholerae strain 62013, then cloned in a pET28a expression vector and transformed into an E. coli (DH5 α host strain. Subsequently, the recombinant vector was retransformed into E. coli BL21 for expression, induced by isopropythio-β-D-galctoside (IPTG at a different concentration, and examined by SDS-PAGE and Western blot. A rabbit ileal loop experiment was conducted. Antibacterial activity of the Ace protein was assessed for E. coli, Stapylococcus aureus (S. aureus, and Pseudomonas aeruginosa (P. aeruginosa.Results: The recombinant Ace protein with a molecular weight of 18 kDa (dimeric form was expressed in E. coli BL21. The Ace protein showed poor staining with Coomassie blue stain, but stained efficiently with silver stain. Western blot analysis showed that the recombinant Ace protein reacted with rabbit anti-V. cholerae polyclonal antibody. The Ace protein had antibacterial activity at a concentration of ≥200 μg/ml and caused significant fluid accumulation in the ligated rabbit ileal loop test.Conclusion: This study described an E. coli cloning and expression system (E. coli BL21- pET-28a-ace for the Ace protein of V. cholerae. We confirmed the antibacterial properties and enterotoxin

  10. Comparison of real time RT-PCR and flow cytometry methods for evaluation of biological activity of recombinant human erythropoietin

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    Sepehrizadeh Z

    2008-05-01

    Full Text Available Background: Evaluation of bioactivity of recombinant erythropoietin is essential for pharmaceutical industry, quality control authorities and researchers. The purpose of this study was to compare real time RT-PCR and flow cytometry for the assay of biological activity of recombinant erythropoietin. Methods: Three concentrations of recombinant erythropoietin BRP (80, 40 and 20 IU/ml were injected subcutaneously to mice. After 4 days the blood was collected and used for reticulocyte counts by flow cytometry and also for the RNA extraction. Real time RT-PCR amplification was carried out for β-globin. Results and conclusion: There was a significant correlation between the total RNA amounts (R2= 0.9995, relative quantity of β-globin mRNA (R2= 0.984 and reticulocyte counts (R2= 0.9742 with rhEpo concentrations. Total RNA and quantitative RT-PCR showed significant dose dependent results as well the reticulocyte counts by flow cytometry for the biological activity assay of rhEpo and so these methods could be considered as alternatives for flow cytometry.

  11. Expression and purification of biologically active recombinant human paraoxonase 1 from inclusion bodies of Escherichia coli.

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    Bajaj, Priyanka; Tripathy, Rajan K; Aggarwal, Geetika; Pande, Abhay H

    2015-11-01

    Human PON1 (h-PON1) is a Ca(2+)-dependent serum enzyme and can hydrolyze (and inactivate) a wide range of substrates. It is a multifaceted enzyme and exhibit anti-inflammatory, anti-oxidative, anti-atherogenic, anti-diabetic, anti-microbial, and organophosphate (OP)-detoxifying properties. Thus, h-PON1 is a strong candidate for the development of therapeutic intervention against these conditions in humans. Insufficient hydrolyzing activity of native h-PON1 against desirable substrate affirms the urgent need to develop improved variant(s) of h-PON1 having enhanced activity. Production of recombinant h-PON1 (rh-PON1) using an Escherichia coli expression system is a key to develop such variant(s). However, generation of rh-PON1 using E. coli expression system has been elusive until now because of the aggregation of over-expressed rh-PON1 protein in inactive form as inclusion bodies (IBs) in the bacterial cells. In this study, we have over-expressed rh-PON1(wt) and rh-PON1(H115W;R192K) proteins as IBs in E. coli, and refolded the inactive enzymes present in the IBs to their active form using in vitro refolding. The active enzymes were isolated from the refolding mixture by ion-exchange chromatography. The catalytic properties of the refolded enzymes were similar to their soluble counterparts. Our results show that the pure and the active variant of rh-PON1 enzyme having enhanced hydrolyzing activity can be produced in large quantities using E. coli expression system. This method can be used for the industrial scale production of rh-PON1 enzymes and will aid in developing h-PON1 as a therapeutic candidate.

  12. Neuronal targeting, internalization, and biological activity of a recombinant atoxic derivative of botulinum neurotoxin A

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    Botulinum neurotoxins (BoNT) have the unique capacity to cross epithelial barriers, target neuromuscular junctions, and translocate active metalloprotease component to the cytosol of motor neurons. We have taken advantage of the molecular carriers responsible for this trafficking to create a family ...

  13. Recombinant hamster oviductin is biologically active and exerts positive effects on sperm functions and sperm-oocyte binding.

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    Xiaojing Yang

    Full Text Available Studies carried out in several mammalian species suggest that oviductin, also known as oviduct-specific glycoprotein or OVGP1, plays a key role in sperm capacitation, fertilization, and development of early embryos. In the present study, we used recombinant DNA technology to produce, for the first time, recombinant hamster OVGP1 (rHamOVGP1 in human embryonic kidney 293 (HEK293 cells. rHamOVGP1 secreted in the culture medium was purified by affinity chromatography. The resulting protein migrated as a poly-dispersed band of 160-350 kDa on SDS-PAGE corresponding to the molecular mass of the native HamOVGP1. Subsequent mass spectrometric analysis of the purified rHamOVGP1 confirmed its identity as HamOVGP1. Immunocytochemistry demonstrated binding of rHamOVGP1 to the mid-piece and head of hamster sperm and to the zona pellucida (ZP of ovarian oocytes. In vitro functional experiments showed that addition of rHamOVGP1 in the capacitation medium further enhanced tyrosine phosphorylation of two sperm proteins of approximately 75 kDa and 83 kDa in a time-dependent manner. After 3 hours of incubation in the presence of rHamOVGP1, a significant increase in acrosome reaction was measured. Pretreatment of either sperm or oocyte with 20 μg/ml of rHamOVGP1 prior to sperm-egg binding assay significantly increased the number of sperm bound to the ZP. Addition of rHamOVGP1 in the medium during sperm-egg binding with either oocyte or sperm pretreated with rHamOVGP1 also saw an increase in the number of sperm bound to ZP. In all experimental conditions, the effect of rHamOVGP1 on sperm-oocyte binding was negated by the addition of monoclonal anti-HamOVGP1 antibody. The successful production and purification of a biologically active rHamOVGP1 will allow further exploration of the function of this glycoprotein in reproductive function.

  14. Recombinant hamster oviductin is biologically active and exerts positive effects on sperm functions and sperm-oocyte binding.

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    Yang, Xiaojing; Zhao, Yuewen; Yang, Xiaolong; Kan, Frederick W K

    2015-01-01

    Studies carried out in several mammalian species suggest that oviductin, also known as oviduct-specific glycoprotein or OVGP1, plays a key role in sperm capacitation, fertilization, and development of early embryos. In the present study, we used recombinant DNA technology to produce, for the first time, recombinant hamster OVGP1 (rHamOVGP1) in human embryonic kidney 293 (HEK293) cells. rHamOVGP1 secreted in the culture medium was purified by affinity chromatography. The resulting protein migrated as a poly-dispersed band of 160-350 kDa on SDS-PAGE corresponding to the molecular mass of the native HamOVGP1. Subsequent mass spectrometric analysis of the purified rHamOVGP1 confirmed its identity as HamOVGP1. Immunocytochemistry demonstrated binding of rHamOVGP1 to the mid-piece and head of hamster sperm and to the zona pellucida (ZP) of ovarian oocytes. In vitro functional experiments showed that addition of rHamOVGP1 in the capacitation medium further enhanced tyrosine phosphorylation of two sperm proteins of approximately 75 kDa and 83 kDa in a time-dependent manner. After 3 hours of incubation in the presence of rHamOVGP1, a significant increase in acrosome reaction was measured. Pretreatment of either sperm or oocyte with 20 μg/ml of rHamOVGP1 prior to sperm-egg binding assay significantly increased the number of sperm bound to the ZP. Addition of rHamOVGP1 in the medium during sperm-egg binding with either oocyte or sperm pretreated with rHamOVGP1 also saw an increase in the number of sperm bound to ZP. In all experimental conditions, the effect of rHamOVGP1 on sperm-oocyte binding was negated by the addition of monoclonal anti-HamOVGP1 antibody. The successful production and purification of a biologically active rHamOVGP1 will allow further exploration of the function of this glycoprotein in reproductive function.

  15. Antiviral cationic peptides as a strategy for innovation in global health therapeutics for dengue virus: high yield production of the biologically active recombinant plectasin peptide.

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    Rothan, Hussin A; Mohamed, Zulqarnain; Suhaeb, Abdulrazzaq M; Rahman, Noorsaadah Abd; Yusof, Rohana

    2013-11-01

    Dengue virus infects millions of people worldwide, and there is no vaccine or anti-dengue therapeutic available. Antimicrobial peptides have been shown to possess effective antiviral activity against various viruses. One of the main limitations of developing these peptides as potent antiviral drugs is the high cost of production. In this study, high yield production of biologically active plectasin peptide was inexpensively achieved by producing tandem plectasin peptides as inclusion bodies in E. coli. Antiviral activity of the recombinant peptide towards dengue serotype-2 NS2B-NS3 protease (DENV2 NS2B-NS3pro) was assessed as a target to inhibit dengue virus replication in Vero cells. Single units of recombinant plectasin were collected after applying consecutive steps of refolding, cleaving by Factor Xa, and nickel column purification to obtain recombinant proteins of high purity. The maximal nontoxic dose (MNTD) of the recombinant peptide against Vero cells was 20 μM (100 μg/mL). The reaction velocity of DENV2 NS2B-NS3pro decreased significantly after increasing concentrations of recombinant plectasin were applied to the reaction mixture. Plectasin peptide noncompetitively inhibited DENV2 NS2B-NS3pro at Ki value of 5.03 ± 0.98 μM. The percentage of viral inhibition was more than 80% at the MNTD value of plectasin. In this study, biologically active recombinant plectasin which was able to inhibit dengue protease and viral replication in Vero cells was successfully produced in E. coli in a time- and cost- effective method. These findings are potentially important in the development of potent therapeutics against dengue infection.

  16. Complexity through Recombination: From Chemistry to Biology

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    Carolina Díaz Arenas

    2010-12-01

    Full Text Available Recombination is a common event in nature, with examples in physics, chemistry, and biology. This process is characterized by the spontaneous reorganization of structural units to form new entities. Upon reorganization, the complexity of the overall system can change. In particular the components of the system can now experience a new response to externally applied selection criteria, such that the evolutionary trajectory of the system is altered. In this work we explore the link between chemical and biological forms of recombination. We estimate how the net system complexity changes, through analysis of RNA-RNA recombination and by mathematical modeling. Our results underscore the importance of recombination in the origins of life on the Earth and its subsequent evolutionary divergence.

  17. Secretory expression and efficient purification of recombinant anthrax toxin lethal factor with full biological activity in E. coli.

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    Liu, Ju; Cai, Chenguang; Guo, Qiang; Zhang, Jun; Dong, Dayong; Li, Guanlin; Fu, Ling; Xu, Junjie; Chen, Wei

    2013-05-01

    Lethal factor (LF), a virulence factor of Bacillus anthracis, plays key roles in anthrax pathogenesis and host-pathogen interactions. The detailed mechanisms by which LF contributes to infection are still under investigation. While these studies require pure, homogeneous and reliable LF preparations, most methods reported for production of recombinant LF (rLF) in B. anthracis or Escherichia coli either are complicated or add extra residues to the protein. In this work, we modified our previous method by codon optimization and chromatograph workflow refinement and developed an improved strategy for efficient production of rLF from the periplasm of E. coli. We were able to obtain fully functional rLF with a purity above 95% and with a considerable yield of 5 mg/L. The preparation was characterized by SDS-PAGE, Western blot, and N-terminal sequencing, and the activity was validated by intoxication of macrophages and Fischer 344 rats. Our final product is suitable for most research involving drug development and mechanism analysis of anthrax pathogenesis.

  18. A simple method for production and purification of soluble and biologically active recombinant human leukemia inhibitory factor (hLIF) fusion protein in Escherichia coli.

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    Imsoonthornruksa, Sumeth; Noisa, Parinya; Parnpai, Rangsun; Ketudat-Cairns, Mariena

    2011-02-20

    Mouse embryonic stem cells (mESCs) rely on a cytokine named leukemia inhibitory factor (LIF) to maintain their undifferentiated state and pluripotency. However, the progress of mESC research is restricted and limited to highly funded laboratories due to the cost of commercial LIF. Here we presented the homemade hLIF which is biologically active. The hLIF cDNA was cloned into two different vectors in order to produce N-terminal His₆-tag and Trx-His₆-tag hLIF fusion proteins in Origami(DE3) Escherichia coli. The His₆-hLIF fusion protein was not as soluble as the Trx-His₆-hLIF fusion protein. One-step immobilized metal affinity chromatography (IMAC) was done to recover high purity (> 95% pure) His₆-hLIF and Trx-His₆-hLIF fusion proteins with the yields of 100 and 200 mg/l of cell culture, respectively. The hLIF fusion proteins were identified by Western blot and verified by mass spectrometry (LC/MS/MS). The hLIF fusion proteins specifically promote the proliferation of TF-1 cells in a dose-dependent manner. They also demonstrate the potency to retain the morphology of undifferentiated mESCs, in that they were positive for mESC markers (Oct-4, Sox-2, Nanog, SSEA-1 and alkaline phosphatase activity). These results demonstrated that the N-terminal fusion tags of the His₆-hLIF and Trx-His₆-hLIF fusion proteins do not interfere with their biological activity. This expression and purification approach to produce recombinant hLIF is a simple, reliable, cost effective and user-friendly method.

  19. Recombinant human nerve growth factor is biologically active and labels novel high-affinity binding sites in rat brain

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    Altar, C.A.; Burton, L.E.; Bennett, G.L.; Dugich-Djordjevic, M. (Genentech, Inc., South San Francisco, CA (USA))

    1991-01-01

    Iodinated recombinant human nerve growth factor (125I-rhNGF) stimulated neurite formation in PC12 cell cultures with a half-maximal potency of 35-49 pg/ml, compared with 39-52 pg/ml for rhNGF. In quantitative ligand autoradiography, the in vitro equilibrium binding of 125I-rhNGF to brain sections showed a 10-fold regional variation in density and was saturable, reversible, and specifically displaced by up to 74% with rhNGF or murine NGF (muNGF). At equilibrium, 125I-rhNGF bound to these sites with high affinity and low capacity (Bmax less than or equal to 13.2 fmol/mg of protein). Calculation of 125I-rhNGF binding affinity by kinetic methods gave average Kd values of 24 and 31 pM. Computer-generated maps revealed binding in brain regions not identified previously with 125I-muNGF, including hippocampus; dentate gyrus; amygdala; paraventricular thalamus; frontal, parietal, occipital, and cingulate cortices; nucleus accumbens; olfactory tubercle; subiculum; pineal gland; and medial geniculate nucleus. NGF binding sites were distributed in a 2-fold increasing medial-lateral gradient in the caudate-putamen and a 2-fold lateral-medial gradient in the nucleus accumbens. 125I-rhNGF binding sites were also found in most areas labeled by 125I-muNGF, including the interpedunucular nucleus, cerebellum, forebrain cholinergic nuclei, caudoventral caudate-putamen, and trigeminal nerve nucleus. 125I-rhNGF binding sites were absent from areas replete with low-affinity NGF binding sites, including circumventricular organs, myelinated fiber bundles, and choroid plexus. The present analysis provides an anatomical differentiation of high-affinity 125I-rhNGF binding sites and greatly expands the number of brain structures that may respond to endogenous NGF or exogenously administered rhNGF.

  20. Recombinant human thrombopoietin: basic biology and evaluation of clinical studies.

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    Kuter, David J; Begley, C Glenn

    2002-11-15

    Thrombocytopenia is a common medical problem for which the main treatment is platelet transfusion. Given the increasing use of platelets and the declining donor population, identification of a safe and effective platelet growth factor could improve the management of thrombocytopenia. Thrombopoietin (TPO), the c-Mpl ligand, is the primary physiologic regulator of megakaryocyte and platelet development. Since the purification of TPO in 1994, 2 recombinant forms of the c-Mpl ligand--recombinant human thrombopoietin (rhTPO) and pegylated recombinant human megakaryocyte growth and development factor (PEG-rHuMGDF)--have undergone extensive clinical investigation. Both have been shown to be potent stimulators of megakaryocyte growth and platelet production and are biologically active in reducing the thrombocytopenia of nonmyeloablative chemotherapy. However, neither TPO has demonstrated benefit in stem cell transplantation or leukemia chemotherapy. Other clinical studies have investigated the use of TPO in treating chronic nonchemotherapy-induced thrombocytopenia associated with myelodysplastic syndromes, idiopathic thrombocytopenic purpura, thrombocytopenia due to human immunodeficiency virus, and liver disease. Based solely on animal studies, TPO may be effective in reducing surgical thrombocytopenia and bleeding, ex vivo expansion of pluripotent stem cells, and as a radioprotectant. Ongoing and future studies will help define the clinical role of recombinant TPO and TPO mimetics in the treatment of chemotherapy- and nonchemotherapy-induced thrombocytopenia.

  1. Overproduction and biological activity of prodigiosin-like pigments from recombinant fusant of endophytic marine Streptomyces species.

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    El-Bondkly, Ahmed M A; El-Gendy, Mervat M A; Bassyouni, Rasha H

    2012-11-01

    Thirty-four endophytic marine Actinomycetes isolates were recovered from the Egyptian marine sponge Latrunculia corticata, out of them 5 isolates (14.7 %) showed red single colonies on yeast-CzAPEK plates. Isolates under the isolation code NRC50 and NRC51 were observed with the strongest red biomass. After application of protoplast fusion between NRC50 and NRC51 isolates, 26 fusants were selected and produced widely different amounts of prodigiosin-like pigments (PLPs) on different fermentation media. Among them fusant NRCF69 produced 79 and 160.4 % PLPs more than parental strains NRC50 and NRC51, respectively. According to the analysis of 16S rDNA sequence (amplified, sequenced, and submitted to GenBank under Accession no. JN232405 and JN232406, respectively), together with their morphological and biochemical characteristics, parental strains NRC50 (P1) and NRC51 (P2) were identified as Streptomyces sp. and designated as Streptomyces sp. NRC50 and Streptomyces sp. NRC51. This study describes a low cost, effective production media by using peanut seed broth, sunflower oil broth or dairy processing wastewater broth alone, or supplemented with 0.5 % mannitol that supports the production of PLPs by the Streptomyces fusant NRCF69 under study (42.03, 40.11, 36.7 and 47 g L(-1), respectively). PLPs compounds exhibited significant cytotoxic activities against three human cancer cell lines: colon cancer cell line (HCT-116), liver cancer cell line (HEPG-2) and breast cancer cell line (MCF-7) and antimycotic activity against clinical dermatophyte isolates of Trichophyton, Microsporum and Epidermophyton.

  2. Production of recombinant orange-spotted grouper (Epinephelus coioides) follicle-stimulating hormone (FSH) in single-chain form and dimer form by Pichia pastoris and their biological activities.

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    Chen, Jun; Zhang, Yanhong; Tang, Zhiguo; Mao, Jiewei; Kuang, Zhonglei; Qin, Chaobin; Li, Wensheng

    2012-09-01

    FSH is a key regulator of steroidogenesis and gonadal growth in teleosts. However, function of FSH is elusive in grouper due to the lack of purified and native FSH. In the present study, we reported production of bioactive orange-spotted grouper (Epinephelus coioides) FSH in dimer form and single-chain form by Pichia pastoris. Dimer form of recombinant grouper FSH (rgFSHba) was accomplished by co-expressing mature FSHb-subunit and a-subunit genes. Fusion of mature FSHb-subunit and a-subunit genes together linking with a polypeptide (4×(Gly-Ser)-Gly-Thr) gene generated single-chain form of recombinant grouper FSH (rgFSHb-a). Recombinant grouper common α-subunit (rgCga) and FSHb-subunit (rgFSHb) were also separately produced. Recombinant proteins were verified by Western blot and mass spectrometry assays, and characterized by deglycosylation analysis. Deglycosylation assay suggested that glycosylation of recombinant FSH mainly occurred on common a-subunit. Bioactivities of recombinant proteins were initially evaluated by activating grouper FSH receptor, and further demonstrated by incubating ovarian fragments of adult grouper and intraperitoneal injection in juvenile female grouper. Two forms of recombinant FSH presented similar biological activities of activating FSH receptor and stimulating in vitro testosterone (T) and estradiol-17β (E2) secretion, though the dimer form functioned slightly weaker than the single-chain form. However, injections of rgFSHb-a or rgFSHba could significantly increase serum T and E2 levels, induce early ovarian development, reduce hypothalamic gnrh1 mRNA level, and increase hypothalamic cyp19a1b mRNA level. Data in this study suggested that recombinant gonadotropin could be produced in dimer form or single-chain form by P. pastoris, and FSH could regulate steroidogenesis and early ovarian development in juvenile grouper.

  3. Cell biology of homologous recombination in yeast

    DEFF Research Database (Denmark)

    Eckert-Boulet, Nadine Valerie; Rothstein, Rodney; Lisby, Michael

    2011-01-01

    Homologous recombination is an important pathway for error-free repair of DNA lesions, such as single- and double-strand breaks, and for rescue of collapsed replication forks. Here, we describe protocols for live cell imaging of single-lesion recombination events in the yeast Saccharomyces...

  4. The biological activity of a recombinantly expressed (His)(6)-tagged peanut allergen (rAra h 1) is unaffected by endotoxin removal

    DEFF Research Database (Denmark)

    Jensen, Louise Bjerremann; Torp, Anna Maria; Andersen, Sven Bode

    2008-01-01

    The application of recombinant (His)(6)-tagged proteins in cell culture assays is associated with problems due to lipopolysaccharide (LPS) contamination. LPS stimulates cells of the immune system, thereby masking antigen-specific activation of T cells. Due to the affinity of LPS for histidine it ...

  5. Biological activities of recombinant equine luteinizing hormone/chorionic gonadotropin (eLH/CG) expressed in Sf9 and Mimic insect cell lines.

    Science.gov (United States)

    Legardinier, Sébastien; Duonor-Cérutti, Martine; Devauchelle, Gérard; Combarnous, Yves; Cahoreau, Claire

    2005-02-01

    Equine luteinizing hormone (eLH) and chorionic gonadotropin (eCG) are composed of identical alpha and beta polypeptide chains, but eCG subunits are much more heavily glycosylated and sialylated. Consequently, eCG exhibits a much longer half-life than eLH in blood. Recombinant eLH/CG, expressed in Sf9 and Mimic insect cells, were compared with one another and to the natural hormones eCG and eLH. Mimic cells are stably-transformed Sf9 cells, expressing five mammalian genes encoding glycosyltransferases involved in the synthesis of complex N-carbohydrate chains. Recombinant eLH/CG expressed in Mimic cells exhibited a higher apparent molecular weight (MW) than that expressed in Sf9 cells, suggesting that its N-glycosylation was, as expected, more complete. Nevertheless, the two recombinant eLH/CG exhibited lower MW than natural eCG from pregnant mare plasma. The two eLH/CG produced in Sf9 and Mimic cells were found to be active in in vitro LH and FSH bioassays, with potencies similar to those of eCG. By contrast, they exhibited no significant in vivo bioactivity, neither in the specific follicle-stimulating hormone (FSH) assay nor in the specific eCG assay. Although recombinant eLH/CG produced in Mimic cells bears more elaborate carbohydrate chains than recombinant eLH/CG from Sf9 cells, it exhibits no significant in vivo bioactivity, probably because of insufficient terminal sialylation of its carbohydrate chains, leading to its rapid removal from blood.

  6. Classification of Recombinant Biologics in the EU

    DEFF Research Database (Denmark)

    Klein, Kevin; De Bruin, Marie L; Broekmans, Andre W;

    2015-01-01

    BACKGROUND AND OBJECTIVE: Biological medicinal products (biologics) are subject to specific pharmacovigilance requirements to ensure that biologics are identifiable by brand name and batch number in adverse drug reaction (ADR) reports. Since Member States collect ADR data at the national level be...

  7. Purification and biological activity of recombinant thymosin alpha 1%重组胸腺素α1的分离纯化和活性测定

    Institute of Scientific and Technical Information of China (English)

    石继红; 张英起; 赵宁; 颜真; 韩苇; 赵永同

    2001-01-01

    *kg-1 of the total bacterial proteins. After CNBr cleava ge of the fusion protein, Tα1 was obtained by ion exchange methods and proved by 2L-Tricine-SDS-PAGE analysis. HPLC showed the purity of Tα1 was 950 g*kg-1. Their biological activity was analysed by 3H-TdR incorparation. They results sh owed that the fusion protein and Tα1 had the similar biological activity to that of the synthesized Tα1. They could increase the p roliferative response of the mitogen ConA stimulated spleen lymphocytes. CONCLUSION Method is convenient and simple for purification and preparation of Tα1 in large scale.

  8. Biological characterization of a recombinant pseudorabies virus

    Energy Technology Data Exchange (ETDEWEB)

    Alvarez, E.; Prieto, C.; Martinez-Lobo, F. J.; Castro, J. M.

    2008-07-01

    In a previous study we obtained and characterized in vitro a novel pseudorabies virus (PRV) variant named gIp2 with a TK, gI/gE, 11k and 28k negative phenotype and a duplication of PK gene. The main objective of the present study was to determine the safety and efficacy, as a vaccine candidate, of this recombinant PRV. For this purpose, we used 24 PRV seronegative three weeks old piglets that were divided into five groups of treatment. Piglets of groups A and B were immunized twice with 10{sup 6}.5 and 10{sup 5}.5 TCID{sub 5}0 of gIp2, respectively; pigs of group C were vaccinated twice with MLV vaccine Auskipra GN and pigs of groups D and E were not immunized and served as infected and uninfected controls, respectively. Four weeks after the second immunization pigs of groups A, B, C and D were challenged by intranasal inoculation of 10{sup 6} TCID{sub 5}0 of the wild type NIA-3 strain of PRV. No adverse reactions or clinical signs were observed in any group after immunization, indicating that the application of up to 10 times the conventional dose included in a commercial vaccine (i.e. 10{sup 5}.5 TCID{sub 5}0) of gIp2 was safe in piglets. Additionally, the inoculation of gIp2 induced an immune response able to provide clinical and virological protection against pseudorabies virus after challenge. In conclusion, the use of gIp2 in piglets as a vaccine virus is safe and induces an immunity comparable to that exerted by commercially available vaccines. (Author) 34 refs.

  9. Palifermin (recombinant keratinocyte growth factor-1): a pleiotropic growth factor with multiple biological activities in preventing chemotherapy- and radiotherapy-induced mucositis.

    NARCIS (Netherlands)

    Blijlevens, N.M.A.; Sonis, S.T.

    2007-01-01

    Oral and intestinal mucositis are among the most significant dose-limiting toxic effects of intensive cancer treatment and are associated with adverse clinical and economic outcomes. Palifermin (Kepivancetrade mark), an N-truncated recombinant human keratinocyte growth factor-1, is the first agent t

  10. A plant cell-expressed recombinant anti-TNF fusion protein is biologically active in the gut and alleviates immune-mediated hepatitis and colitis.

    Science.gov (United States)

    Ilan, Yaron; Gingis-Velitski, Svetlana; Ben Ya'aco, Ami; Shabbat, Yehudit; Zolotarov, Lidya; Almon, Einat; Shaaltiel, Yoseph

    2017-03-01

    The orally administered BY-2 plant cell-expressed recombinant anti-TNF fusion protein (PRX-106) (n=6) consists of the soluble form of the human TNF receptor (TNFR) fused to the Fc component of a human antibody IgG1 domain.

  11. Purification and characterization of biologically active human recombinant 37 kDa soluble CD23 (sFc epsilon RII) expressed in insect cells.

    Science.gov (United States)

    Graber, P; Jansen, K; Pochon, S; Shields, J; Aubonney, N; Turcatti, G; Bonnefoy, J Y

    1992-05-18

    Human recombinant soluble 37 kDa CD23 has been expressed in insect cells and secreted into the culture medium using the IL-2 leader sequence. The 37 kDa CD23 was purified 600-fold to homogeneity by monoclonal antibody affinity chromatography and gel filtration. The pure protein is monomeric, glycosylated, depleted of one N terminal amino acid and contains four disulphide bonds. It degrades into smaller fragments of 33, 29 and 25 kDa if purified in the absence of protease inhibitors. The same pattern of proteolytic fragments is observed when the pure preparation is incubated at room temperature for 3 weeks. Physical characterization of the 37 kDa CD23 by circular dichroism indicates that the protein contains mainly beta sheet and 20% of alpha helical structures. Specific binding of IgE to natural CD23 (low affinity IgE receptor) was inhibited by purified recombinant 37 kDa CD23. Moreover, purified recombinant 37kDa CD23 and interleukin-1 promoted the survival of germinal centre B cells.

  12. The relationship between the connecting peptide of recombined single chain insulin and its biological function

    Institute of Scientific and Technical Information of China (English)

    黄一丁; 梁镇和; 冯佑民

    2001-01-01

    To investigate the relationship between the biological activity of recombined single chain insulin and the length of the connecting peptide, we designed and prepared three single chain insulin molecules, namely, PIP, [A]5PIP and [A]10PIP, by site-directed mutagenesis, in which B30 and A1 were linked through dipeptide A-K, heptapeptide A-A-A-A-A-A-K, and dodecapeptide A-A-A-A-A-A-A-A-A-A-A-K, respectively. Their receptor binding capacities were 0.14%, 14.3% and 11.1% of that of insulin respectively and their in vivo biological activities were in consistence with their receptor binding capacity; whereas their growth promoting activities were 17%, 116.3% and 38% of that of insulin. These results suggested the following conclusions. (i) The recombined single chain insulin could also possess the same metabolic and mitogenic function as insulin. (ii) The receptor binding capacity of recombined single chain insulin to insulin receptor was closely related to the length and amino acid composition of the connecting peptide and could change from 0 to 100% of insulin depending on the different connecting peptides. This result further illustrated the necessity of B chain C-terminus swaying away from A chain N-terminus when insulin binds to its receptor. (iii) The mitogenic activity of recombined single chain insulin also depended on the length and the amino acid composition of the connecting peptide and was higher than its metabolic activity.

  13. Sublingual immunotherapy: from biological extracts to recombinant allergens.

    Science.gov (United States)

    Moingeon, P

    2006-01-01

    Sublingual vaccines based on biological extracts from various natural allergen sources are effective in the treatment of respiratory allergies. These vaccines comprise a complex mixture of proteins and glycoproteins that require dedicated standardization procedures to ensure batch-to-batch consistency. Because of the lack of correlation between the potency of an allergen extract and the quantity of major allergen content, standardization is achieved predominantly by determining the global IgE binding capacity of the extract in vitro. New proteomic technologies can be used to further characterize the most abundant proteins present in an extract. Second-generation sublingual vaccines based on recombinant allergens are under development. The aim is to produce molecularly defined vaccines that exhibit superior efficacy, while allowing for simplified immunization schedules. In this approach, recombinant DNA technology is used to express highly purified allergens in their native (i.e. wild-type) conformation. The recombinant allergens are then formulated with ad hoc adjuvants and/or mucoadhesive galenic excipients so that they specifically target oral Langerhans cells and induce allergen-specific regulatory T cells.

  14. Cloning, bacterial expression and biological characterization of recombinant human granulocyte chemotactic protein-2 and differential expression of granulocyte chemotactic protein-2 and epithelial cell-derived neutrophil activating peptide-78 mRNAs.

    Science.gov (United States)

    Froyen, G; Proost, P; Ronsse, I; Mitera, T; Haelens, A; Wuyts, A; Opdenakker, G; Van Damme, J; Billiau, A

    1997-02-01

    Human osteosarcoma cells secrete a novel C-X-C chemokine called granulocyte chemotactic protein-2 (GCP-2), which was previously identified by amino acid sequencing of the purified natural protein. In order to understand the role of this new protein in inflammatory reactions, we cloned GCP-2 DNA sequences to generate recombinant protein and specific DNA probes and primers. By means of PCR on cloned cDNA of osteosarcoma cells induced by interleukin-1 beta and fibroblasts induced by lipopolysaccharide plus dsRNA, the complete coding domain of GCP-2 was isolated. This sequence was cloned into the bacterial expression vector pHEN1 and, after induction, GCP-2 was secreted into the periplasm of Escherichia coli. Recombinant GCP-2 (rGCP-2) was purified and characterized by SDS/PAGE as a monomeric 6.5-kDa protein and by amino-terminal sequencing. The chemoattractive potency of GCP-2 for neutrophilic granulocytes was about 10-times less than that of interleukin-8 and the minimal effective dose was 10 ng/ml. However, at optimal dose (100 ng/ml) the maximal chemotactic response was comparable with that of interleukin-8. Both characteristics correspond with those of natural GCP-2. In addition, intracellular calcium release in neutrophils by recombinant GCP-2 was achieved with as little as 10 ng/ml. Quantitation studies using reverse transcriptase and the polymerase chain reaction revealed higher GCP-2 mRNA production in normal fibroblasts than in tumor cells. When compared with epithelial-cell-derived neutrophil-activating peptide-78 (ENA-78) mRNA, the GCP-2 mRNA levels were higher in all cell lines tested. In addition, GCP-2 and ENA-78 expression seem to be differentially regulated in that phorbol ester and lipopolysaccharide have opposing effects on their mRNA induction in diploid fibroblasts and epithelial cells, respectively. Interleukin-1 was demonstrated to be a general inducer for both chemokines, while interferon-gamma down-regulates their mRNA expression. The

  15. Soluble Fc gamma R (sFc gamma R): detection in biological fluids and production of a murine recombinant sFc gamma R biologically active in vitro and in vivo.

    Science.gov (United States)

    Sautès, C; Teillaud, C; Mazières, N; Tartour, E; Bouchard, C; Galinha, A; Jourde, M; Spagnoli, R; Fridman, W H

    1992-08-01

    Soluble forms of receptors for the Fc portion of IgG (sFc gamma R) were detected in biological fluids from mice and humans. In mouse bearing tumors, circulating amounts of sFc gamma R increased concurrently with tumor growth. Tumors secreting IgG2a, IgG2b or IgG3 led to a 5- to 10-fold increase in serum sFc gamma R levels whereas tumors secreting IgG1, IgGA or other types of tumors (non Ig B cell tumors, T cell lymphoma and a melanoma) increased 2- to 3-fold the levels of circulating sFc gamma R. In the human, sFc gamma R were also detected in whole unstimulated saliva. Levels of sFc gamma RII and of sFc gamma RIII were variable and did not seem to depend on the dental status of the individuals. Finally, a murine recombinant sFc gamma R (rsFc gamma R) composed of the two extracellular domains of Fc gamma RII was produced by culture of transfected L cells in bioreactors. The purified rsFc gamma R was found to inhibit antibody production in vitro in anti-SRBC responses and by cultures of small B cells stimulated by anti-IgM antibodies in the presence of IL-4 and IL-5. Moreover, the i.p. injection of this material into adult mice immunized with SRBC led to a decrease of IgG antibody production by splenocytes, as measured by a hemolytic plaque assay, and in serum, as measured by antigen-specific ELISA.

  16. CONSTRUCTION,EXPRESSION AND BIOLOGICAL ASSESSMENT OF BPI23—Fcγ1 RECOMBINANT PROTEIN PROKARYOTIC EXPRESSION VECTOR

    Institute of Scientific and Technical Information of China (English)

    安云庆; 管远志; 等

    2002-01-01

    Objective:To construct pBV-BPI600-Fcγ1700 recombinant expression vector,to transform it into Escherichia coli DH5α,and to induce the expression of BPI23-Fcγ1 anti-bacterial recombinant protein.Methods:Genes coding for BPI23 and Fcγ1 were amplified by RT-PCR from mRNA extracted from Hl-60 cell and normal human leukocytes;recombinant cloning vector and recombinant expression vector were then constructed.pBV-BPI600-Fcγ1700 recombinant expression vector was transformed into the competent Escherichia coli DH5α and BPI23-Fcγ1 recombinant protein was expressed by a temperature-induced method.Results:(1)Expected amplified products BPI600bp and Fcγ1700bp were obtained by RT-PCR method.(2)pUC18-BPI180,pUC18-BPI420 and pUC18-Fcγ1700 recombinant cloning vectors were successfully constructed, and sequences were identical with the reported ones.(3)pBV-BPI600-Fcγ1700 recombinant expression vector was successfully constructed,and the enzyme digestion analysis showed an expected result.(4)The expression level of BPI23-Fcγ1 recombinant protein accounted for 20% of total bacterial proteins.(5)The renatured BPI23-Fcγ1 recombinant protein showed bacteriocidal activity and biological function of complement fixation,and opsonization.Conclusion:pBV-BPI600-Fcγ1700 recombinant expression vector was successfully constructed,and BPI23-Fcγ1 recombinant protein with double biological activity of BPI and IgGFc was expressed in Escherichia coli.

  17. CONSTRUCTION, EXPRESSION AND BIOLOGICAL ASSESSMENT OF BPI23-Fcγ 1 RECOMBINANT PROTEIN PROKARYOTIC EXPRESSION VECTOR

    Institute of Scientific and Technical Information of China (English)

    安云庆; 管远志; 柯岩; 杨贵贞

    2002-01-01

    Objective. To construct pBV-BPI600-Fcγ 1700 recombinant expression vector, to transform it into Escherichia coli DH5α , and to induce the expression of BPI23-Fcγ 1 anti-bacterial recombinant protein. Methods. Genes coding for BPI23 and Fcγ 1 were amplified by RT-PCR from mRNA extracted from HL-60 cell and normal human leukocytes; recombinant cloning vector and recombinant expression vector were then constructed. pBV-BPI600-Fcγ 1700 recombinant expression vector was transformed into the competent Escherichia coli DH5α and BPI23-Fcγ 1 recombinant protein was expressed by a temperature-induced method. Results. (1) Expected amplified products BPI600bp and Fcγ 1700bp were obtained by RT-PCR method. (2) pUC18-BPI180, pUC18-BPI420 and pUC18-Fcγ 1700 recombinant cloning vectors were successfully constructed, and sequences were identical with the reported ones. (3) pBV-BPI600-Fcγ 1700 recombinant expression vector was successfully constructed, and the enzyme digestion analysis showed an expected result. (4) The expression level of BPI23-Fcγ 1 recombinant protein accounted for 20% of total bacterial proteins. (5) The renatured BPI23-Fcγ 1 recombinant protein showed bacteriocidal activity and biological function of complement fixation, and opsonization. Conclusion. pBV-BPI600-Fcγ 1700 recombinant expression vector was successfully constructed, and BPI23-Fcγ 1 recombinant protein with double biological activity of BPI and IgGFc was expressed in Escherichia coli.

  18. Recombinant protein expression plasmids optimized for industrial E. coli fermentation and plant systems produce biologically active human insulin-like growth factor-1 in transgenic rice and tobacco plants.

    Science.gov (United States)

    Panahi, Mitra; Alli, Zaman; Cheng, Xiongying; Belbaraka, Loubaba; Belgoudi, Jaafar; Sardana, Ravinder; Phipps, Jenny; Altosaar, Illimar

    2004-06-01

    Human insulin-like growth factor-1 (hIGF-1) is a growth factor with clinical significance in medicine. The therapeutic potential of recombinant hIGF-1 (rthIGF-1) stems from the fact that hIGF-1 resembles insulin in many aspects of physiology. The expression of hIGF-1 in transgenic tobacco and rice plants using different expression cassettes is reported here. In the present study, two coding sequences were tested, one with the original human sequence, but partially optimized for expression in E. coli and the other with a plant-codon-optimized sequence that was expected to give a higher level of expression in plant systems. Three different hIGF-1 recombinant expression constructs were generated. All expression constructs utilized the maize ubiquitin 1 promoter with or without a signal sequence. Analyses conducted using a hIGF-1 specific ELISA kit showed all transgenic plants produced hIGF-1 and the accumulated hIGF-1 increased from the E. coli codon bias to higher levels when the hIGF-1 coding sequence was codon-optimized to match that of the maize zeamatin protein--the most transcribed gene in maize endosperm suspension cells. Further analyses that compared the functionality of the bacterial signal peptide Lam B in plants showed that this leader peptide led to lower expression levels when compared to transgenic plants that did not contain this sequence. This indicated that this expression construct was functional without removal of the bacterial signal sequence. The maize ubiquitin 1 promoter was found to be more active in rice plants than tobacco plants indicating that in this case, there was a class preference that was biased towards a monocot host. Biological analyses conducted using protein extracts from transgenic plants showed that the rthIGF-1 was effective in stimulating the in vitro growth and proliferation of human SH-SY5Y neuroblastoma cells. This indicated that the plant-produced rthIGF-1 was stable and biologically active. As some plants have been

  19. [Biological activity of Spirulina].

    Science.gov (United States)

    Blinkova, L P; Gorobets, O B; Baturo, A P

    2001-01-01

    In this review information of Spirulina platensis (SP), a blue-green alga (photosynthesizing cyanobacterium) having diverse biological activity is presented. Due to high content of highly valuable proteins, indispensable amino acids, vitamins, beta-carotene and other pigments, mineral substances, indispensable fatty acids and polysaccharides, PS has been found suitable for use as bioactive additive. SP produces an immunostimulating effect by enhancing the resistance of humans, mammals, chickens and fish to infections, the capacity of influencing hemopoiesis, stimulating the production of antibodies and cytokines. Under the influence of SP macrophages, T and B cells are activated. SP sulfolipids have proved to be effective against HIV. Preparations obtained from SP biomass have also been found active against herpesvirus, cytomegalovirus, influenza virus, etc. SP extracts are capable in inhibiting cancerogenesis. SP preparations are regarded as functional products contributing to the preservation of the resident intestinal microflora, especially lactic acid bacilli and bifidobacteria, and to a decrease in the level of Candida albicans. The biological activity of SP with respect to microorganisms holds good promise for using these microalgae as components of culture media.

  20. Cold Spring Harbor symposia on quantitative biology: Volume 49, Recombination at the DNA level

    Energy Technology Data Exchange (ETDEWEB)

    1984-01-01

    This volume contains full papers prepared by the participants to the 1984 Cold Springs Harbor Symposia on Quantitative Biology. This year's theme is entitled Recombination at the DNA level. The volume consists of 93 articles grouped into subject areas entitled chromosome mechanics, yeast systems, mammalian homologous recombination, transposons, mu, plant transposons/T4 recombination, topoisomerase, resolvase and gyrase, Escherichia coli general recombination, RecA, repair, leukaryotic enzymes, integration and excision of bacteriophage, site-specific recombination, and recombination in vitro.

  1. Biological Activity of Masked Endotoxin

    Science.gov (United States)

    Schwarz, Harald; Gornicec, Jan; Neuper, Theresa; Parigiani, Maria Alejandra; Wallner, Michael; Duschl, Albert; Horejs-Hoeck, Jutta

    2017-01-01

    Low endotoxin recovery (LER) is a recently discovered phenomenon describing the inability of limulus amebocyte lysate (LAL)-based assays to detect lipopolysaccharide (LPS) because of a “masking effect” caused by chelators or detergents commonly used in buffer formulations for medical products and recombinant proteins. This study investigates the masking capacities of different buffer formulations and whether masked endotoxin is biologically active. We show that both naturally occurring endotoxin as well as control standard endotoxin can be affected by LER. Furthermore, whereas masked endotoxin cannot be detected in Factor C based assays, it is still detectable in a cell-based TLR4-NF-κB-luciferase reporter gene assay. Moreover, in primary human monocytes, masked LPS induces the expression of pro-inflammatory cytokines and surface activation markers even at very low concentrations. We therefore conclude that masked LPS is a potent trigger of immune responses, which emphasizes the potential danger of masked LPS, as it may pose a health threat in pharmaceutical products or compromise experimental results. PMID:28317862

  2. BIOLOGICAL EFFECTS OF TNF-BINDING VARIOLAVIRUS RECOMBINANT PROTEIN

    Directory of Open Access Journals (Sweden)

    I. A. Orlovskaya

    2012-01-01

    Full Text Available Abstract. This review presents a summary of our data concerning in vivo and in vitro effects of recombinant TNF-binding protein from variola virus (VARVCrmB upon TNF-induced functional changes of human and murine cells. VARV-CrmB protein blocks TNF-induced production of IL-1β and IL-6 by human mononuclear cells, and their in vitro oxidation-related metabolic (OM activity. VARV-CrmB protein restores TNF-induced reduction of BFU-E+CFU-E colony-forming activity and normalizes TNF-induced effects upon CFU-GM formation in a colony-forming model of human and murine bone morrow cells (BMC. VARV-CrmB protein displays a pronounced in vivo alleviation of LPS-induced endotoxic shock symptoms in SPF BALB/C mice thus significantly increasing survival of experimental animals. Moreover, VARV-CrmBprotein decreases intensity of collagen-induced arthritis at early terms. Application of VARV-CrmB protein results in normalization of TNF-induced increase of migratory and OM activity of murine leukocytes, and exerts corrective effects upon colony-forming ability of murine hematopoietic precursors. Skin application of VARV-CrmB protein decreases leukocyte migration from a skin scrap in afferent phase of DNCB-induced contact reaction, as well as “ear oedema” index. Our results demonstrate TNF-blocking properties of VARVCrmB protein. In summary, our data allow to consider a recombinant variola virus VARV-CrmB as a new potential TNF-antagonist. Its effects can be explained by its ability of neutralizing TNF-induced activation of oxidation-related metabolic, cytokine-producing and migratory functions of effector cells in therapy of pathological inflammatory processes.

  3. 一种重组β-内酰胺酶的原核表达,纯化及其活性检测%Prokaryotic expression,purification and biological activity analysis of recombinant β-Lactamase protein

    Institute of Scientific and Technical Information of China (English)

    周晓靓; 施培基; 王浩

    2011-01-01

    目的:利用原核表达系统诱导表达一种可以用于肿瘤生物治疗ADEPT(单抗导向酶活化前药)的重组蛋白RGD4CβL,并对该蛋白进行纯化以及活性检测.方法:对合成的DNA序列进行测序,将重组载体pColdⅡ-RGD4CβL转化人大肠杆菌E.coli BL(DE3),在大肠杆菌细胞中诱导表达目的蛋白,用His标签纯化树脂Ni-NTA纯化目的蛋白,以流式细胞术(FCM)检测纯化所得蛋白的活性.结果:合成的DNA全长为1125 bp,其在E.coli BL(DE3)中经IPTG诱导后表达出相对分子质量(Mr)约为42 000的目的蛋白,经纯化后获得了产率和纯度均很高的重组蛋白RGD4CβL,流式细胞检测证明该蛋白具有很好的肿瘤细胞靶向性.结论:成功地对重组蛋白RGD4CβL进行了诱导表达,得到的纯化蛋白具有较好的肿瘤细胞靶向性,该重组蛋白将可能成为一种免疫原性更低的新型双功能蛋白用于ADEPT.%AIM: To prepare RGD4CβL fusion proteinusing prokaryotic expression system and evaluate the biological activity of the RGD4CβL. METHODS: RGD4CβL gene was cloned into pColdll to contruct β-Lactamase prokaryotic expression vector. After transformation, the recombinant vector was induced to express recombinant protein RGD4CβL by IPTG in E. coli BL ( DE3 ). The recombinant protein was purified by Ni-NTA resin under denaturing condition and then dialyzed to renature. The tumor cell targeting ability of the recombinant protein was analyzed by flow cytometric analysis. RESULTS: After cleavage and purification,β-Lactamase moiety showed the expected size of 42 000 on Tricine-SDS-PAGE, and was further confirmed by Western blotting. Based on flow cytometric analysis, the purified protein specially targeted breast cancer cell line MCF-7. CONCLUSION: This research successfully estiblished a method for prokaryotic expression and purification of β-lactamase.These results suggest the potential use of the protein as an agent for ADEPT.

  4. Using synthetic biology to increase nitrogenase activity

    OpenAIRE

    Li, Xin-Xin; Liu, Qi; Liu, Xiao-Meng; Shi, Hao-Wen; Chen, San-feng

    2016-01-01

    Background Nitrogen fixation has been established in protokaryotic model Escherichia coli by transferring a minimal nif gene cluster composed of 9 genes (nifB, nifH, nifD, nifK, nifE, nifN, nifX, hesA and nifV) from Paenibacillus sp. WLY78. However, the nitrogenase activity in the recombinant E. coli 78-7 is only 10 % of that observed in wild-type Paenibacillus. Thus, it is necessary to increase nitrogenase activity through synthetic biology. Results In order to increase nitrogenase activity ...

  5. Dielectronic Recombination In Active Galactic Nuclei

    Science.gov (United States)

    Lukić, D.; Savin, D. W.; Schnell, M.; Brandau, C.; Schmidt, E.; Schippers, S.; Müller, A.; Lestinsky, M.; Sprenger, F.; Wolf, A.; Altun, Z.; Badnell, N. R.

    2006-05-01

    Recent X-ray satelitte observations of active galactic nuclei point out shortcomings in our understanding of low temperature dielectronic recombination (DR) for iron M- shell ions. In order to resolve this issue and to provide reliable iron M-shell DR data for modeling astrophysical plasmas, we are carrying out a series of laboratory measurements using the heavy-ion Test Storage Ring at the Max- Plank-Institute for Nuclear Physics in Heidelberg, Germany. Storage rings are currently the only laboratory method capable of studying low temperature DR. We use our results to produce experimentally- derived DR rate coefficients. We are also providing our data to atomic theorist to benchmark their DR calculations. Here we will report our recent DR results for selected Fe M-shell ions. At temperatures where these ions are predicted to form in photoionized gas, we find a significant discrepancy between our experimental results and previously recommended DR rate coefficients.

  6. How-to-Do-It: Teaching Recombinant DNA Technology in High School Biology Courses.

    Science.gov (United States)

    Dixon, Linda

    1988-01-01

    Reports on the teaching of recombinant DNA technology in high school biology courses. Explains reactions of the public, students, and colleagues to the molecular genetics unit. Indicates equipment, curricular materials, training, workshops, and availability. (RT)

  7. A recombinant wheat serpin with inhibitory activity

    DEFF Research Database (Denmark)

    Rasmussen, Søren K; Dahl, Søren Weis; Nørgård, Anette

    1996-01-01

    A full-length clone encoding the wheat (Triticum aestivum L.) serpin WSZ1 was isolated from a cDNA library based on mRNA from immature grain. The 398 amino acid sequence deduced from the cDNA was corroborated by sequencing CNBr peptides of WSZ1 purified from resting grain. WSZ1 belongs...... to the subfamily of protein Z-type serpins and the amino acid sequence is 70%, identical with the barley serpins BSZ4 and BSZx and 27-33% identical with human serpins such as alpha(1)-proteinase inhibitor, antithrombin III, and plasminogen activator inhibitor. The cDNA was subcloned in the pET3d expression vector......, equipped with a histidine affinity tag at the N-terminus and expressed in Escherichia coli BL(21) DE3 pLysS. Recombinant WSZ1 from the soluble fraction was partially purified on Ni-NTA agarose and MonoQ columns and shown to form SDS-stable complexes with sc-chymotrypsin. Southern blots and amino acid...

  8. Biological Activities of Hydrazone Derivatives

    Directory of Open Access Journals (Sweden)

    S. Güniz Küçükgüzel

    2007-08-01

    Full Text Available There has been considerable interest in the development of novel compounds with anticonvulsant, antidepressant, analgesic, antiinflammatory, antiplatelet, antimalarial, antimicrobial, antimycobacterial, antitumoral, vasodilator, antiviral and antischistosomiasis activities. Hydrazones possessing an azometine -NHN=CH- proton constitute an important class of compounds for new drug development. Therefore, many researchers have synthesized these compounds as target structures and evaluated their biological activities. These observations have been guiding for the development of new hydrazones that possess varied biological activities.

  9. Recent advances in yeast molecular biology: recombinant DNA. [Lead abstract

    Energy Technology Data Exchange (ETDEWEB)

    1982-09-01

    Separate abstracts were prepared for the 25 papers presented at a workshop focusing on chromosomal structure, gene regulation, recombination, DNA repair, and cell type control, that have been obtained by experimental approaches incorporating the new technologies of yeast DNA transformation, molecular cloning, and DNA sequence analysis. (KRM)

  10. Biological activities of alginate.

    Science.gov (United States)

    Ueno, Mikinori; Oda, Tatsuya

    2014-01-01

    To gain insight into the structure-activity relationship of alginate, we examined the effect of alginates with varying molecular weights and M/G ratio on murine macrophage cell line, RAW264.7 cells in terms of induction of tumor necrosis factor-α (TNF-α) secretion. Among the alginates tested, alginate with the highest molecular weight (MW 38,000, M/G 2.24) showed the most potent TNF-α-inducing activity. Alginates having higher M/G ratio tended to show higher activity. These results suggest that molecular size and M/G ratio are important structural parameters influencing the TNF-α-inducing activity. Interestingly, enzymatic depolymerization of alginate with bacterial alginate lyase resulted in dramatic increase in the TNF-α-inducing activity. The higher activity of enzymatically digested alginate oligomers to induce nitric oxide production from RAW264.7 cells than alginate polymer was also observed. On the other hand, alginate polymer and oligomer showed nearly equal hydroxyl radical scavenging activities.

  11. Recombinant production of Epstein-Barr virus BZLF1 trans-activator and characterization of its DNA-binding specificity.

    Science.gov (United States)

    Lim, Chun Shen; Goh, Siang Ling; Krishnan, Gopala; Ng, Ching Ching

    2014-03-01

    This paper describes the recombinant production of a biologically active Epstein-Barr virus BZLF1 trans-activator, i.e., Z-encoded broadly reactive activator (ZEBRA), that recognized specific DNA motifs. We used auto-induction for histidine-tagged BZLF1 expression in Escherichia coli and immobilized cobalt affinity membrane chromatography for protein purification under native conditions. We obtained the purified BZLF1 at a yield of 5.4mg per gram of wet weight cells at 75% purity, in which 27% of the recombinant BZLF1 remained biologically active. The recombinant BZLF1 bound to oligonucleotides containing ZEBRA response elements, either AP-1 or ZIIIB, but not a ZIIIB mutant. The recombinant BZLF1 showed a specific DNA-binding activity which could be useful for functional studies.

  12. Recombinant biologic products versus nutraceuticals from plants - a regulatory choice?

    Science.gov (United States)

    Drake, Pascal M W; Szeto, Tim H; Paul, Mathew J; Teh, Audrey Y-H; Ma, Julian K-C

    2017-01-01

    Biotechnology has transformed the potential for plants to be a manufacturing source of pharmaceutical compounds. Now, with transgenic and transient expression techniques, virtually any biologic, including vaccines and therapeutics, could be manufactured in plants. However, uncertainty over the regulatory path for such new pharmaceuticals has been a deterrent. Consideration has been given to using alternative regulatory paths, including those for nutraceuticals or cosmetic agents. This review will consider these possibilities, and discuss the difficulties in establishing regulatory guidelines for new pharmaceutical manufacturing technologies.

  13. Fast and easy protocol for the purification of recombinant S-layer protein for synthetic biology applications

    KAUST Repository

    Norville, Julie E.

    2011-06-17

    A goal of synthetic biology is to make biological systems easier to engineer. One of the aims is to design, with nanometer-scale precision, biomaterials with well-defined properties. The surface-layer protein SbpA forms 2D arrays naturally on the cell surface of Lysinibacillus sphaericus, but also as the purified protein in solution upon the addition of divalent cations. The high propensity of SbpA to form crystalline arrays, which can be simply controlled by divalent cations, and the possibility to genetically alter the protein, make SbpA an attractive molecule for synthetic biology. To be a useful tool, however, it is important that a simple protocol can be used to produce recombinant wild-type and modified SbpA in large quantities and in a biologically active form. The present study addresses this requirement by introducing a mild and non-denaturing purification protocol to produce milligram quantities of recombinant, active SbpA.

  14. A synthetic biology approach to self-regulatory recombinant protein production in Escherichia coli

    Directory of Open Access Journals (Sweden)

    Dragosits Martin

    2012-03-01

    Full Text Available Abstract Background Recombinant protein production is a process of great industrial interest, with products that range from pharmaceuticals to biofuels. Since high level production of recombinant protein imposes significant stress in the host organism, several methods have been developed over the years to optimize protein production. So far, these trial-and-error techniques have proved laborious and sensitive to process parameters, while there has been no attempt to address the problem by applying Synthetic Biology principles and methods, such as integration of standardized parts in novel synthetic circuits. Results We present a novel self-regulatory protein production system that couples the control of recombinant protein production with a stress-induced, negative feedback mechanism. The synthetic circuit allows the down-regulation of recombinant protein expression through a stress-induced promoter. We used E. coli as the host organism, since it is widely used in recombinant processes. Our results show that the introduction of the self-regulatory circuit increases the soluble/insoluble ratio of recombinant protein at the expense of total protein yield. To further elucidate the dynamics of the system, we developed a computational model that is in agreement with the observed experimental data, and provides insight on the interplay between protein solubility and yield. Conclusion Our work introduces the idea of a self-regulatory circuit for recombinant protein products, and paves the way for processes with reduced external control or monitoring needs. It demonstrates that the library of standard biological parts serves as a valuable resource for initial synthetic blocks that needs to be further refined to be successfully applied in practical problems of biotechnological significance. Finally, the development of a predictive model in conjunction with experimental validation facilitates a better understanding of the underlying dynamics and can be

  15. Imidazole: Having Versatile Biological Activities

    Directory of Open Access Journals (Sweden)

    Amita Verma

    2013-01-01

    Full Text Available Imidazoles have occupied a unique position in heterocyclic chemistry, and its derivatives have attracted considerable interests in recent years for their versatile properties in chemistry and pharmacology. Imidazole is nitrogen-containing heterocyclic ring which possesses biological and pharmaceutical importance. Thus, imidazole compounds have been an interesting source for researchers for more than a century. The imidazole ring is a constituent of several important natural products, including purine, histamine, histidine, and nucleic acid. Being a polar and ionisable aromatic compound, it improves pharmacokinetic characteristics of lead molecules and thus is used as a remedy to optimize solubility and bioavailability parameters of proposed poorly soluble lead molecules. There are several methods used for the synthesis of imidazole-containing compounds, and also their various structure reactions offer enormous scope in the field of medicinal chemistry. The imidazole derivatives possess extensive spectrum of biological activities such as antibacterial, anticancer, antitubercular, antifungal, analgesic, and anti-HIV activities. This paper aims to review the biological activities of imidazole during the past years.

  16. Evaluation of the effectiveness and safety of the thermo-treatment process to dispose of recombinant DNA waste from biological research laboratories.

    Science.gov (United States)

    Li, Meng-Nan; Zheng, Guang-Hong; Wang, Lei; Xiao, Wei; Fu, Xiao-Hua; Le, Yi-Quan; Ren, Da-Ming

    2009-01-01

    The discharge of recombinant DNA waste from biological laboratories into the eco-system may be one of the pathways resulting in horizontal gene transfer or "gene pollution". Heating at 100 degrees C for 5-10 min is a common method for treating recombinant DNA waste in biological research laboratories in China. In this study, we evaluated the effectiveness and the safety of the thermo-treatment method in the disposal of recombinant DNA waste. Quantitative PCR, plasmid transformation and electrophoresis technology were used to evaluate the decay/denaturation efficiency during the thermo-treatment process of recombinant plasmid, pET-28b. Results showed that prolonging thermo-treatment time could improve decay efficiency of the plasmid, and its decay half-life was 2.7-4.0 min during the thermo-treatment at 100 degrees C. However, after 30 min of thermo-treatment some transforming activity remained. Higher ionic strength could protect recombinant plasmid from decay during the treatment process. These results indicate that thermo-treatment at 100 degrees C cannot decay and inactivate pET-28b completely. In addition, preliminary results showed that thermo-treated recombinant plasmids were not degraded completely in a short period when they were discharged into an aquatic environment. This implies that when thermo-treated recombinant DNAs are discharged into the eco-system, they may have enough time to re-nature and transform, thus resulting in gene diffusion.

  17. Activity of recombinant factor VIIa under different conditions in vitro

    DEFF Research Database (Denmark)

    Bladbjerg, Else-Marie; Jespersen, Jørgen

    2008-01-01

    Recombinant activated factor VII (NovoSeven; Novo Nordisk A/S, Måløv, Denmark) is an effective drug for treatment of bleeding in patients with haemophilia A or B and inhibitors. Little is known about physiological conditions influencing the efficacy of recombinant activated factor VII. We......, 33, 37, and 40 degrees C, or diluted 0, 10, 20, 40, and 60% with dextran before analysis. Samples were analysed as rotational thromboelastometry in whole blood (clotting time, clot formation time, and maximum clot firmness) with and without Innovin (tissue factor), and as factor VII coagulant...... activity in plasma. Significant effects of pH were observed for clotting time, clot formation time, maximum clot firmness, and factor VII coagulant activity in the direction of longer clot formation times and less firm clots with decreasing pH. Temperature had significant effects on clotting time, clot...

  18. Staphylococcus simulans Recombinant Lysostaphin: Production, Purification, and Determination of Antistaphylococcal Activity.

    Science.gov (United States)

    Boksha, I S; Lavrova, N V; Grishin, A V; Demidenko, A V; Lyashchuk, A M; Galushkina, Z M; Ovchinnikov, R S; Umyarov, A M; Avetisian, L R; Chernukha, M Iu; Shaginian, I A; Lunin, V G; Karyagina, A S

    2016-05-01

    Staphylococcus simulans lysostaphin is an endopeptidase lysing staphylococcus cell walls by cleaving pentaglycine cross-bridges in their peptidoglycan. A synthetic gene encoding S. simulans lysostaphin was cloned in Escherichia coli cells, and producer strains were designed. The level of produced biologically active lysostaphin comprised 6-30% of total E. coli cell protein (depending on E. coli M15 or BL21 producer) under batch cultivation conditions. New methods were developed for purification of lysostaphin without affinity domains and for testing its enzymatic activity. As judged by PAGE, the purified recombinant lysostaphin is of >97% purity. The produced lysostaphin lysed cells of Staphylococcus aureus and Staphylococcus haemolyticus clinical isolates. In vitro activity and general biochemical properties of purified recombinant lysostaphin produced by M15 or BL21 E. coli strains were identical to those of recombinant lysostaphin supplied by Sigma-Aldrich (USA) and used as reference in other known studies. The prepared recombinant lysostaphin represents a potential product for development of enzymatic preparation for medicine and veterinary due to the simple purification scheme enabling production of the enzyme of high purity and antistaphylococcal activity.

  19. 重组人β-神经生长因子在昆虫细胞中的表达、纯化及其生物学活性%Expression of recombinant human β-nerve growth factor in insect cells and purification and biological activity of expressed product

    Institute of Scientific and Technical Information of China (English)

    李佳楠; 王玲; 吕新凯; 吴红梅; 汤华东

    2013-01-01

    Objective To express recombinant human B nerve growth factor (rhB-NGF) in insect cells, purify the expressed product and determine its biological activity. Methods Recombinant plasmid Bacmid + [rhβ-NGF] was constructed and transfected to insect cell line Sf9 in mediation of liposome cellfection Ⅱ. Sf9 cells were repeatedly infected with the supernatant of transfected cells, of which the culture supernatant containing target protein was collected in a large quantity. The rhB-NGF was purified by ion exchange and molecular sieve chromatography, identified by SDS-PAGE and Western blot and determined for biological activity by chick embryo dorsal root nervous ganglion culture method. Results Restriction analysis and sequencing proved that recombinant plasmid Bacmid + [rhB-NGF] was constructed correctly. The rhB-NGF was expressed in Sf9 cells infected with recombinant virus, which showed a single band with relative molecular mass of about 14 200 on SDS-PAGE profile and reached a purity of more than 98% after purification. The expressed protein showed specific binding to rabbit anti-human monoclonal antibody against NGF and stimulated the growth of nerve ganglion process, of which the specific activity was about 600 000 AU / mg. Conclusion The rhB-NGF with biological activity was expressed successfully in insect cells, which laid a foundation of preparation of rhB-NGF in a large quantity.%目的 在昆虫细胞中表达重组人β-神经生长因子(Recombinant human β nerve growth factor,rhβ-NGF),并对表达产物进行纯化和生物学活性鉴定.方法 构建rhβ-NGF杆状病毒表达质粒Bacmid+[rhβ-NGF],利用脂质体cellfectionⅡ将质粒转染至昆虫细胞Sf9中,用转染细胞的上清反复感染Sf9细胞,大量收获含目的蛋白的培养上清,利用离子交换层析和分子筛层析纯化rhβ-NGF,纯化产物经SDS-PAGE和Western blot分析,并通过鸡胚背根神经节培养法检测rhβ-NGF的生物学活性.结果

  20. Detection of contaminating enzymatic activity in plant-derived recombinant biotechnology products.

    Science.gov (United States)

    Brinson, Robert G; Giulian, Gary G; Kelman, Zvi; Marino, John P

    2014-12-02

    Residual impurities in recombinantly produced protein biologics, such as host cell proteins (HCP), can potentially cause unwanted toxic or immunogenic responses in patients. Additionally, undetected impurities found in recombinant proteins used in cell culture may adversely impact basic research and biotechnology applications. Currently, the enzyme-linked immunosorbent assay (ELISA) is the standard for detection of residual HCP contamination in recombinantly produced biologics. Alternatively, two-dimensional liquid chromatography coupled to mass spectrometry is being developed as a tool for assessing this critical quality attribute. Both of these methods rely on the direct detection of HCPs and some previous knowledge of the contaminant. For contaminating enzymes, the mass level of the impurity may fall below the threshold of detection of these methods and underestimate the true impact. To address this point, here we demonstrate facile detection and characterization of contaminating phytase activity in rice-derived recombinant human serum albumin (rHSA) using a sensitive, label-free nuclear magnetic resonance (NMR) spectroscopy assay. We observed varying degrees of phytase contamination in biotechnology-grade rHSA from various manufacturers by monitoring the degradation of adenosine-5'-triphosphate and myo-inositol-1,2,3,4,5,6-hexakisphosphate by (31)P NMR. The observed lot-to-lot variability may result in irreproducible cell culture results and should be evaluated as a possible critical quality attribute in plant-derived biotherapeutics.

  1. Molecular characteristics versus biological activity

    Science.gov (United States)

    Applegate, Vernon C.; Smith, Manning A.; Willeford, Bennett R.

    1967-01-01

    The molecular characteristics of mononitrophenols containing halogens not only play a key role in their biological activity but provide a novel example of selective toxicity among vertebrate animals. It has been reported that efforts to control the parasitic sea lamprey in the Great Lakes are directed at present to the applications of a selective toxicant to streams inhabited by lamprey larvae. Since 1961, the larvicide that has been used almost exclusively in the control program has been 3-trifluoromethyl-4-nitrophenol (TFM). However, this is only one of about 15 closely related compounds, all halogen-containing mononitrophenols, that display a selectively toxic action upon lampreys. Although not all of the halogenated mononitrophenols are selectively toxic to lampreys (in fact, fewer than half of those tested), no other group of related compounds has displayed any useful larvicidal activity except for the substituted nitrosalicylanilides.

  2. Detection of recombinant and cellular MALT1 paracaspase activity.

    Science.gov (United States)

    Nagel, Daniel; Krappmann, Daniel

    2015-01-01

    MALT1 (mucosa-associated lymphoid tissue protein 1) is a key regulator of antigen-induced NF-κB activation in the adaptive immune response. Activation of proteolytic activity of the MALT1 paracaspase was shown to boost the immune response. Additionally, MALT1 proteolytic activity is essential for the survival of MALT1-dependent lymphoma, such as the activated B-cell type (ABC) of diffuse large B-cell lymphoma (DLBCL) or MALT lymphoma. The functional impact of MALT1 paracaspase on T-cell activation and lymphomagenesis suggests that MALT1 is a promising therapeutic target for the treatment of autoimmune diseases and distinct lymphoma entities. To evaluate the requirement of MALT1 in further detail, direct measurement of its activity status is of great importance. We have established a fluorogenic cleavage assay which can be used to measure activity of recombinant and cellular MALT1. Here we describe the basis of the cleavage assay and include a detailed protocol for recombinant production of MALT1 and also the cellular immunoprecipitation of endogenous MALT1 to determine its proteolytic activity.

  3. Histidine tag fusion increases expression levels of active recombinant amelogenin in Escherichia coli.

    Science.gov (United States)

    Svensson, Johan; Andersson, Christer; Reseland, Janne E; Lyngstadaas, Petter; Bülow, Leif

    2006-07-01

    Amelogenin is a dental enamel matrix protein involved in formation of dental enamel. In this study, we have expressed two different recombinant murine amelogenins in Escherichia coli: the untagged rM179, and the histidine tagged rp(H)M180, identical to rM179 except that it carries the additional N-terminal sequence MRGSHHHHHHGS. The effects of the histidine tag on expression levels, and on growth properties of the amelogenin expressing cells were studied. Purification of a crude protein extract containing rp(H)M180 was also carried out using IMAC and reverse-phase HPLC. The results of this study showed clearly that both growth properties and amelogenin expression levels were improved for E. coli cells expressing the histidine tagged amelogenin rp(H)M180, compared to cells expressing the untagged amelogenin rM179. The positive effect of the histidine tag on amelogenin expression is proposed to be due to the hydrophilic nature of the histidine tag, generating a more hydrophilic amelogenin, which is more compatible with the host cell. Human osteoblasts treated with the purified rp(H)M180 showed increased levels of secreted osteocalcin, compared to untreated cells. This response was similar to cells treated with enamel matrix derivate, mainly composed by amelogenin, suggesting that the recombinant protein is biologically active. Thus, the histidine tag favors expression and purification of biologically active recombinant amelogenin.

  4. Three faces of recombination activating gene 1 (RAG1) mutations.

    Science.gov (United States)

    Patiroglu, Turkan; Akar, Himmet Haluk; Van Der Burg, Mirjam

    2015-12-01

    Severe combined immune deficiency (SCID) is a group of genetic disorder associated with development of T- and/or B-lymphocytes. Recombination-activating genes (RAG1/2) play a critical role on VDJ recombination process that leads to the production of a broad T-cell receptor (TCR) and B-cell receptor (BCR) repertoire in the development of T and B cells. RAG1/2 genes mutations result in various forms of primary immunodeficiency, ranging from classic SCID to Omenn syndrome (OS) to atypical SCID with such as granuloma formation and autoimmunity. Herein, we reported 4 patients with RAG1 deficiency: classic SCID was seen in two patients who presented with recurrent pneumonia and chronic diarrhoea, and failure to thrive. OS was observed in one patient who presented with chronic diarrhoea, skin rash, recurrent lower respiratory infections, and atypical SCID was seen in one patient who presented with Pyoderma gangrenosum (PG) and had novel RAG1 mutation.

  5. Intein-mediated Rapid Purification of Recombinant Human Pituitary Adenylate Cyclase Activating Polypeptide

    Institute of Scientific and Technical Information of China (English)

    Rong-jie YU; An HONG; Yun DAI; Yuan GAO

    2004-01-01

    In order to obtain the recombinant human PACAP efficiently by intein-mediated single column purification, a gene encoding human PACAP was synthesized and cloned into Escherichia coli expression vector pKYB. The recombinant vector pKY-PAC was transferred into E. coli ER2566 cells and the target protein was over-expressed as a fusion to the N-terminus of a self-cleavable affinity tag. After the PACAPintein-CBD fusion protein was purified by chitin-affinity chromatography, the self-cleavage activity of the intein was induced by DTT and the rhPACAP was released from the chitin-bound intein tag. The activity of the rhPACAP to stimulate cyclic AMP accumulation was detected using the human pancreas carcinoma cells SW1990. Twenty-two milligrams of rhPACAP with the purity over 98% was obtained by single column purification from 1 liter of induced culture. The preliminary biological assay indicated that the rhPACAP, which has an extra Met at its N-terminus compared with the native human PACAP, had the similar activity of stimulating cAMP accumulation with the standard PACAP38 in the SW1990 cells. A new efficient production procedure of the active recombinant human PACAP was established.

  6. A review exploring biological activities of hydrazones

    Directory of Open Access Journals (Sweden)

    Garima Verma

    2014-01-01

    Full Text Available The development of novel compounds, hydrazones has shown that they possess a wide variety of biological activities viz. antimicrobial, anticonvulsant, antidepressant, anti-inflammatory, analgesic, antiplatelet, antimalarial, anticancer, antifungal, antitubercular, antiviral, cardio protective etc., Hydrazones/azomethines/imines possess-NHN = CH- and constitute an important class of compounds for new drug development. A number of researchers have synthesized and evaluated the biological activities of hydrazones. This review aims at highlighting the diverse biological activities of hydrazones.

  7. Biological properties of purified recombinant HCV particles with an epitope-tagged envelope

    Energy Technology Data Exchange (ETDEWEB)

    Takahashi, Hitoshi; Akazawa, Daisuke [Department of Virology II, National Institute of Infectious Diseases, Tokyo (Japan); Toray Industries, Inc., Kanagawa (Japan); Kato, Takanobu; Date, Tomoko [Department of Virology II, National Institute of Infectious Diseases, Tokyo (Japan); Shirakura, Masayuki [Department of Virology II, National Institute of Infectious Diseases, Tokyo (Japan); Toray Industries, Inc., Kanagawa (Japan); Nakamura, Noriko; Mochizuki, Hidenori [Toray Industries, Inc., Kanagawa (Japan); Tanaka-Kaneko, Keiko; Sata, Tetsutaro [Department of Pathology, National Institute of Infectious Diseases, Tokyo (Japan); Tanaka, Yasuhito [Department of Clinical Molecular Informative Medicine, Nagoya City University Graduate School of Medicine, Nagoya (Japan); Mizokami, Masashi [Research Center for Hepatitis and Immunology, Kohnodai Hospital, International Medical Center of Japan, Chiba (Japan); Suzuki, Tetsuro [Department of Virology II, National Institute of Infectious Diseases, Tokyo (Japan); Wakita, Takaji, E-mail: wakita@nih.go.jp [Department of Virology II, National Institute of Infectious Diseases, Tokyo (Japan)

    2010-05-14

    To establish a simple system for purification of recombinant infectious hepatitis C virus (HCV) particles, we designed a chimeric J6/JFH-1 virus with a FLAG (FL)-epitope-tagged sequence at the N-terminal region of the E2 hypervariable region-1 (HVR1) gene (J6/JFH-1/1FL). We found that introduction of an adaptive mutation at the potential N-glycosylation site (E2N151K) leads to efficient production of the chimeric virus. This finding suggests the involvement of glycosylation at Asn within the envelope protein(s) in HCV morphogenesis. To further analyze the biological properties of the purified recombinant HCV particles, we developed a strategy for large-scale production and purification of recombinant J6/JFH-1/1FL/E2N151K. Infectious particles were purified from the culture medium of J6/JFH-1/1FL/E2N151K-infected Huh-7 cells using anti-FLAG affinity chromatography in combination with ultrafiltration. Electron microscopy of the purified particles using negative staining showed spherical particle structures with a diameter of 40-60 nm and spike-like projections. Purified HCV particle-immunization induced both an anti-E2 and an anti-FLAG antibody response in immunized mice. This strategy may contribute to future detailed analysis of HCV particle structure and to HCV vaccine development.

  8. Recombinant paracoccin reproduces the biological properties of the native protein and induces protective Th1 immunity against Paracoccidioides brasiliensis infection.

    Directory of Open Access Journals (Sweden)

    Ana Claudia Paiva Alegre

    2014-04-01

    Full Text Available BACKGROUND: Paracoccin is a dual-function protein of the yeast Paracoccidioides brasiliensis that has lectin properties and N-acetylglucosaminidase activities. Proteomic analysis of a paracoccin preparation from P. brasiliensis revealed that the sequence matched that of the hypothetical protein encoded by PADG-3347 of isolate Pb-18, with a polypeptide sequence similar to the family 18 endochitinases. These endochitinases are multi-functional proteins, with distinct lectin and enzymatic domains. METHODOLOGY/PRINCIPAL FINDINGS: The multi-exon assembly and the largest exon of the predicted ORF (PADG-3347, was cloned and expressed in Escherichia coli cells, and the features of the recombinant proteins were compared to those of the native paracoccin. The multi-exon protein was also used for protection assays in a mouse model of paracoccidioidomycosis. CONCLUSIONS/SIGNIFICANCE: Our results showed that the recombinant protein reproduced the biological properties described for the native protein-including binding to laminin in a manner that is dependent on carbohydrate recognition-showed N-acetylglucosaminidase activity, and stimulated murine peritoneal macrophages to produce high levels of TNF-α and nitric oxide. Considering the immunomodulatory potential of glycan-binding proteins, we also investigated whether prophylactic administration of recombinant paracoccin affected the course of experimental paracoccidioidomycosis in mice. In comparison to animals injected with vehicle (controls, mice treated with recombinant paracoccin displayed lower pulmonary fungal burdens and reduced pulmonary granulomas. These protective effects were associated with augmented pulmonary levels of IL-12 and IFN-γ. We also observed that injection of paracoccin three days before challenge was the most efficient administration protocol, as the induced Th1 immunity was balanced by high levels of pulmonary IL-10, which may prevent the tissue damage caused by exacerbated

  9. Recombination activating activity of XRCC1 analogous genes in X-ray sensitive and resistant CHO cell lines

    Science.gov (United States)

    Golubnitchaya-Labudová, O.; Portele, A.; Vaçata, V.; Lubec, G.; Rink, H.; Höfer, M.

    1997-10-01

    The XRCC1 gene (X-ray repair cross complementing) complements the DNA repair deficiency of the radiation sensitive Chinese hamster ovary (CHO) mutant cell line EM9 but the mechanism of the correction is not elucidated yet. XRCC1 shows substantial homology to the RAG2 gene (recombination activating gene) and we therefore tried to answer question, whether structural similarities (sequence of a putative recombination activating domain, aa 332-362 for XRCC1 and aa 286-316 in RAG2) would reflect similar functions of the homologous, putative recombination activating domain. PCR experiments revealed that no sequence homologous to the structural part of human XRCC1 was present in cDNA of CHO. Differential display demonstrated two putative recombination activating domains in the parental CHO line AA8 and one in the radiosensitive mutant EM9. Southern blot experiments showed the presence of several genes with partial homology to human XRCC1. Recombination studies consisted of expressing amplified target domains within chimeric proteins in recA - bacteria and subsequent detection of recombination events by sequencing the recombinant plasmids. Recombination experiments demonstrated recombination activating activity of all putative recombination activating domains amplified from AA8 and EM9 genomes as reflected by deletions within the insert of the recombinant plasmids. The recombination activating activity of XRCC1 analogues could explain a mechanism responsible for the correction of the DNA repair defect in EM9.

  10. Insect immune activation by recombinant Galleria mellonella apolipophorin III(1).

    Science.gov (United States)

    Niere, M; Meisslitzer, C; Dettloff, M; Weise, C; Ziegler, M; Wiesner, A

    1999-08-17

    Apolipophorin III (apoLp-III) is an exchangeable insect apolipoprotein. Its function, as currently understood, lies in the stabilization of low-density lipophorin particles (LDLp) crossing the hemocoel in phases of high energy consumption to deliver lipids from the fat body to the flight muscle cells. Recent studies with native Galleria mellonella-apoLp-III gave first indications of an unexpected role of that protein in insect immune activation. Here we report the immune activation by the recombinant protein, documenting a newly discovered correlation between lipid physiology and immune defense in insects. The complete cDNA sequence of G. mellonella-apoLp-III was identified by mixed oligonucleotide-primed amplification of cDNA (MOPAC), 3'-RACE-PCR, and cRACE-PCR. The sequence coding for the native protein was ligated into a pET-vector; this construct was transfected into Escherichia coli and overexpressed in the bacteria. Photometric turbidity assays with human low density lipoprotein (LDL) and transmission electron microscopy studies on apoLp-III-stabilized lipid discs revealed the full functionality of the isolated recombinant apoLp-III with regard to its lipid-association ability. For proving its immune-stimulating capacity, apoLp-III was injected into the hemocoel of last instar G. mellonella larvae and the antibacterial activity in cell-free hemolymph was determined 24 h later. As a result, the hemolymph samples of injected insects contained strongly increased antibacterial activities against E. coli as well as clearly enhanced lysozyme-like activities. From Northern blot analysis of total RNA from insects injected with apoLp-III or the bacterial immune provocator lipopolysaccharide, it could be concluded that the transcription rate of apoLp-III mRNA does not vary in comparison to untreated last instar larvae.

  11. Activity of recombinant and natural defensins from Vigna unguiculata seeds against Leishmania amazonensis.

    Science.gov (United States)

    Souza, Géssika Silva; do Nascimento, Viviane Veiga; de Carvalho, Laís Pessanha; de Melo, Edésio José Tenório; Fernandes, Keysson Vieira; Machado, Olga Lima Tavares; Retamal, Claudio Andres; Gomes, Valdirene Moreira; Carvalho, André de Oliveira

    2013-09-01

    Antimicrobial peptides (AMPs), which are differentiated from other antibiotic peptides, such as gramicidins and polymyxins, because they are synthesized by large enzymatic complex and bear modified amino acids including d-amino acids, are short polymers of l-amino acids synthesized by ribosomes upon which all living organisms rely to defend themselves from invaders or competitor microorganisms. AMPs have received a great deal of attention from the scientific community as potential new drugs for neglected diseases such as Leishmaniasis. In plants, they include several families of compounds, including the plant defensins. The aim of the present study was to improve the expression of recombinant defensin from Vigna unguiculata seeds (Vu-Defr) and to test its activity against Leishmania amazonensis promatigotes. Recombinant expression was performed in LB and TB media and under different conditions. The purification of Vu-Defr was achieved by immobilized metal ion affinity and reversed-phase chromatography. The purified Vu-Defr was analyzed by circular dichroism (CD), and its biological activity was tested against L. amazonenis promastigotes. To demonstrate that the recombinant production of Vu-Defr did not interfere with its fold and biological activity, the results of all experiments were compared with the results from the natural defensin (Vu-Def). The CD spectra of both peptides presented good superimposition indicating that both peptides present very similar secondary structure and that the Vu-Defr was correctly folded. L. amazonensis treated with Vu-Defr led to the elimination of 54.3% and 46.9% of the parasites at 24 and 48h of incubation time, respectively. Vu-Def eliminated 50% and 54.8% of the parasites at 24 and 48 h, respectively. Both were used at a concentration of 100 μg/mL. These results suggested the potential for plant defensins to be used as new antiparasitic substances.

  12. 冻干重组人三突变型HIF-1α腺病毒生物学效应考察%Characterization of the biological activities of lyophilized recombinant adenovirus expressing the triple mutant of hypoxia inducible factor-lα

    Institute of Scientific and Technical Information of China (English)

    陶宇; 王月刚; 魏旋; 刘城; 陈冬冬; 吴平生

    2011-01-01

    To investigate the influence of lyophilization on the biological activity of recombinant adenovirus-mediated triple mutant of hypoxia inducible factor-la (Ad-HIF-la-564/402/803). Methods Ad-HIF-la-564/ 402/803 was amplified from HEK293A cells and purified by ultracentrifugation in CsCl gradient solutions. The infection efficiency was observed by X-gal staining. The lyophilized adenovirus was prepared under appropriate conditions. Before and after lyophilization, the effect of Ad-HIF-la-564/402/803 on hMVEC proliferation was evaluated by MTS assay. The recombinant adenovirus was confirmed by PCR and DNA sequence analysis before and 1 day, 6 months and 12 months after lyophilization, and hMVECs infected with Ad-HIF-la-564/402/803 at these time points were examined for HIF-la protein expression using Western blotting. Results No significant changes were observed in the effect of lyophilized Ad-HIF-la-564/402/803 on hMVECs proliferation at the optimal multiplicity of infection of 100 pfu/cell (P>0.05). At the 4 time points, the recombinant adenovirus HIF-la showed no structural alterations or significant changes in the expression level of HIF-la protein in the transfected hMVECs (P>0.05). Conclusion Lyophilized Ad-HIF-la-564/402/803 can maintain its biological activities for a long time.%目的 研究冻干对重组人三突变型HIF-1α病毒(Ad-HIF-1α-564/402/803)生物学效应的影响.方法 将前期构建的Ad-HIF-1α-564/402/803在HEK293A细胞中进行扩增,用氯化铯浓度梯度离心法进行腺病毒纯化,X-Gal染色法测定重组腺病毒转染效率,在适宜的条件下制成冻于剂.采用MTS试剂盒观察冻干前后Ad-HIF-1α-564/402/803对hMvECs增殖的影响.分别在冻干前、冻干后1 d、6月、12月四个时间点提取病毒DNA,进行PCR及PCR产物测序鉴定重组人三突变型HIF-1α腺病毒基因;并在4个时间点以Ad-HIF-1α-564/402/803转染hMVECs,提取蛋白进行western blot检测HIF-1α蛋白的表达.结果 X

  13. The Recombinant ACTH (4-10) -GDNF Fusion Protein and Study ofIts Biological Activity%重组ACTH(4-10)与GDNF融合蛋白及其生物活性研究

    Institute of Scientific and Technical Information of China (English)

    陈哲宇; 张勇; 何成; 路长林; 吴祥甫

    2001-01-01

    通过PCR方法构建了促肾上腺皮质激素4-10(ACTH(4-10))与胶质细胞源性神经营养因子(GDNF)的 融合基因,并将它重组克隆到表达载体pET-28a(+)中,构建表达质粒pET-ACTH(4-10)-GDNF,转化大肠 杆菌BL21(DE3),经IPTG诱导可高效表达ACTH(4-10)-GDNF融合蛋白.用Ni2+-NTA树脂一步法纯化目 的蛋白,纯度达85%以上.纯化和复性的ACTH(4-10)-GDNF融合蛋白能显著促进脊髓神经元存活,作用强 于ACTH(4-10)及GDNF蛋白.%The chimeric gene of ACTH (4-10) with GDNF was constructed by PCR amplification. The fused gene was inserted into the expression vector pET-28a ( + ) and expressed in E. coli . with a level of 30% of the total bacterial proteins. The expressed product was purified by Ni2+ -NTA resin, up to 85% purity. The results of activity assays showed that the chimeric protein could significantly promote the survival of spinal cord neurons and had a higher neurotrophic activity than ACTH (4-10) and GDNF respectively.

  14. Expression of recombination-activating genes and T cell receptor gene recombination in the human T cell leukemia cell line

    Institute of Scientific and Technical Information of China (English)

    ZOU Hong-yun; MA Li; MENG Min-jie; YAO Xin-sheng; LIN Ying; WU Zhen-qiang; HE Xiao-wei; WANG Ju-fang; WANG Xiao-ning

    2007-01-01

    Background Recent studies have suggested that mature T cells can change their specificity through reexpression of recombination-activating genes (RAG) and RAG-mediated V(D)J recombination. This process is named receptor revision and has been observed in mature peripheral T cells from transgenic mice and human donors. However, whether the receptor revision in mature T cells is a random or orientated process remains poorly understood. Here we used the Jurkat human T cell line, which represents a mature stage of T cell development, as a model to investigate the regulation of T cell receptor (TCR) gene recombination.Methods TCR Dβ-Jβ signal joint T cell receptor excision DNA circles (sjTRECs) were determined by nested and seminested PCR. Double-strand DNA breaks at recombination signal sequences (RSSs) in the TCRVβ chain locus were detected by ligation-mediated-PCR. Further analysis of the complementarity-determining region 3 (CDR3) size of the TCRVβ chain was examined by the TCR GeneScan technique.Results RAG1, RAG2, and three crucial components of the nonhomologous DNA end-joining (NHEJ) pathway were readily detected in Jurkat. Characteristics of junctional diversity of Dβ2-Jβ2 signal joints and ds RSS breaks associated with the Dβ25' and Dβ 23' sites were detected in DNA from Jurkat cells. CDR3 size and the gene sequences of the TCRVβ chain did not change during cell proliferation.Conclusions RAG1 and RAG2 and ongoing TCR gene recombination are coexpressed in Jurkat cells, but the ongoing recombination process may not play a role in modification of the TCR repertoire. However, the results suggest that Jurkat could be used as a model for studying the regulation of RAGs and V(D)J recombination and as a "special" model of the coexistence of TCR gene rearrangements and "negative" receptor revision.

  15. 78 FR 27977 - Office of Biotechnology Activities; Recombinant DNA Research: Proposed Actions Under the NIH...

    Science.gov (United States)

    2013-05-13

    ... HUMAN SERVICES National Institutes of Health Office of Biotechnology Activities; Recombinant DNA... Acid Molecules (NIH Guidelines) SUMMARY: The NIH Office of Biotechnology Activities (NIH OBA) proposes... by mail to the NIH Office of Biotechnology Activities, National Institutes of Health, 6705...

  16. 75 FR 28811 - Office of Biotechnology Activities; Recombinant DNA Research: Proposed Actions Under the NIH...

    Science.gov (United States)

    2010-05-24

    ... HUMAN SERVICES National Institutes of Health Office of Biotechnology Activities; Recombinant DNA... Office of Biotechnology Activities (OBA) by the Institutional Biosafety Committee at Lawrence Livermore... Biotechnology Activities, National Institutes of Health. BILLING CODE 4140-01-P...

  17. Biological activity of liposomal vanillin.

    Science.gov (United States)

    Castan, Leniher; Del Toro, Grisel; Fernández, Adolfo A; González, Manuel; Ortíz, Emilia; Lobo, Daliana

    2013-06-01

    This article presents a study of vanillin encapsulation inside multilamellar liposomes, with emphasis on the evaluation of antioxidant activity, the hemolytic effect, and the antisickling properties of these products. Egg phosphatidylcholine-cholesterol and egg phosphatidylcholine-cholesterol-1-O-decylglycerol liposomes were prepared by mechanical dispersion, all with vanillin included. Vesicles were characterized by determination of encapsulation efficiency and vanillin retention capacity. Antioxidant activity was determined by the 2,2-diphenyl-1-picrylhydrazyl (DPPH) method. The hemolytic effect of liposomes was also evaluated by spectrophotometry, as well as the antisickling activity by the Huck test using optical microscopy. Results showed that the lipid composition of liposomes did not significantly affect the encapsulation efficiency. Stable vesicles were obtained with a high retention percentage of vanillin. Liposomes exhibited a high capture of the DPPH radical compared to free vanillin and 1-O-decylglycerol (C10) in solution. Vesicles caused no significant hemolisys in normal erythrocytes, nor in those coming from patients with sickle cell anemia. Vanillin encapsulated in liposomes retained its antisickling activity, with a greater effect for C10-containing vesicles. Our results show that vanillin encapsulation in liposomes is a way to enhance the pharmacologic properties of this molecule using a suitable vehicle.

  18. Biologically active sesquiterpene coumarins from Ferula species.

    Science.gov (United States)

    Nazari, Zeinab Esmail; Iranshahi, Mehrdad

    2011-03-01

    Extracts from different species of the genus Ferula (Apiaceae) have had various biomedical applications for many centuries. Many biological features of this genus such as cytotoxicity, antibacterial, antiviral, P-glycoprotein (P-gp) inhibitory and antiinflammatory activity have been attributed to sesquiterpene coumarins; structures containing a common coumarin group and a sesquiterpene moiety. This both highlights the importance of sesquiterpene coumarins as biologically active natural products and necessitates further studies on these compounds. Taking into account the versatile biological properties of compounds isolated from Ferula and the unprecedented interest in the application of natural products as a new generation of therapeutics, the present review will discuss reports on biological activities of sesquiterpene coumarins of the genus Ferula, from 1990 onwards.

  19. Recombinant yeast technology at the cutting edge: robust tools for both designed catalysts and new biologicals.

    Science.gov (United States)

    Kovar, Karin; Looser, Verena; Hyka, Petr; Merseburger, Tobias; Meier, Christian

    2010-01-01

    Health and safety concerns, enhanced quality criteria, and environmental sustainability, have prompted investigations into production using recombinant yeasts as a feasible alternative for isolation of proteins from natural animal or plant sources, as well as for processes utilising either mammalian cell cultures or bacterial systems. An overview of recent research papers and review articles provides readers with a comprehensive insight into the field of next-generation yeast expression systems. Major breakthroughs in recombinant yeast technology linked to Pichia pastoris are (i) the public availability of tools to generate proteins with tailored and highly homogenous N-glycan structures, similar to the forms assembled in humans, (ii) the recent accomplishment of the annotation of its genome sequence, and finally, (iii) the presence of the first few (non-glycosylated) therapeutic proteins in Pichia on the market. The P. pastoris expression platform is now well developed, as proven by multiple products used in human and veterinary medicine and in industry (e.g., enzymes for chemical synthesis and for the modification/synthesis of pharmaceuticals, drug target proteins used for structural analysis or for high throughput screening, proteins for diagnostics, proteinous biomaterials, vaccines, and therapeutic proteins). Nevertheless, the complexity of protein analysis (monitoring) continues to restrict process development for recombinant products. Drawing on combined expertise in molecular biology and process technology, the Institute of Biotechnology (IBT) at the Zurich University of Applied Science (ZHAW) and its international partners have developed solutions which (i) fully eliminate (or partially reduce) the use of methanol, which is undesirable in high-cell-density and high-productivity processes, (ii) match both strain construction and process design with the target protein characteristics to the benefit of the cells' physiological shape, and (iii) allow multi

  20. Expression of biologically active murine interleukin-18 in Lactococcus lactis.

    Science.gov (United States)

    Feizollahzadeh, Sadegh; Khanahmad, Hossein; Rahimmanesh, Ilnaz; Ganjalikhani-Hakemi, Mazdak; Andalib, Alireza; Sanei, Mohammad Hossein; Rezaei, Abbas

    2016-11-01

    The food-grade bacterium Lactococcus lactis is increasingly used for heterologous protein expression in therapeutic and industrial applications. The ability of L. lactis to secrete biologically active cytokines may be used for the generation of therapeutic cytokines. Interleukin (IL)-18 enhances the immune response, especially on mucosal surfaces, emphasizing its therapeutic potential. However, it is produced as an inactive precursor and has to be enzymatically cleaved for maturation. We genetically manipulated L. lactis to secrete murine IL-18. The mature murine IL-18 gene was inserted downstream of a nisin promoter in pNZ8149 plasmid and the construct was used to transform L. lactis NZ3900. The transformants were selected on Elliker agar and confirmed by restriction enzyme digestion and sequencing. The expression and secretion of IL-18 protein was verified by SDS-PAGE, western blotting and ELISA. The biological activity of recombinant IL-18 was determined by its ability to induce interferon (IFN)-γ production in L. lactis co-cultured with murine splenic T cells. The amounts of IL-18 in bacterial lysates and supernatants were 3-4 μg mL(-1) and 0.6-0.7 ng mL(-1), respectively. The successfully generated L. lactis strain that expressed biologically active murine IL-18 can be used to evaluate the possible therapeutic effects of IL-18 on mucosal surfaces.

  1. Marine Pyridoacridine Alkaloids: Biosynthesis and Biological Activities.

    Science.gov (United States)

    Ibrahim, Sabrin R M; Mohamed, Gamal A

    2016-01-01

    Pyridoacridines are a class of strictly marine-derived alkaloids that constitute one of the largest chemical families of marine alkaloids. During the last few years, both natural pyridoacridines and their analogues have constituted excellent targets for synthetic works. They have been the subject of intense study due to their significant biological activities; cytotoxic, antibacterial, antifungal, antiviral, insecticidal, anti-HIV, and anti-parasitic activities. In the present review, 95 pyridoacridine alkaloids isolated from marine organisms are discussed in term of their occurrence, biosynthesis, biological activities, and structural assignment.

  2. [Comparison of expression and antibacterial activities of recombinant porcine lactoferrin expressed in four Lactobacillus species].

    Science.gov (United States)

    Yu, Hui; Jiang, Yanping; Cui, Wen; Wu, Xiao; He, Jia; Qiao, Xinyuan; Li, Yijing; Tang, Lijie

    2014-09-01

    The coding sequence for the mature peptide of porcine lactoferrin (Plf) was synthesized according to the codon usage of lactobacillus, to establish optimized porcine lactoferrin Lactobacillus expression system. The gene was ligated into the Xho I/BamH I site of Lactobacillus expression vector pPG612.1 and the recombinant plasmid pPG612.1-plf was transformed individually into Lactobacillus casei ATCC393, Lactobacillus pentosus KLDS1.0413, Lactobacillus plantarum KLDS1.0344 or Lactobacillus paracasei KLDS1.0652 by electroporation. After induction with xylose, expression of the recombinant proteins was detected by Western blotting and confocal laser scanning microscopy. Secretion of recombinant Plf proteins from four recombinant species was determined quantitatively by ELISA. The antibacterial activities of recombinant proteins were measured by agar diffusion method. The result shows that Plf was correctly expressed in four species of recombinant lactobacillus, with molecular weight of about 73 kDa. The expression levels in recombinant Lactobacillus casei, Lactobacillus pentosus, Lactobacillus plantarum, Lactobacillus paracasei were 9.6 μg/mL, 10.8 μg/mL, 12.5 μg/mL and 9.9 μg/mL, respectively. Antimicrobial activity experiment shows that the recombinant proteins were active against E. coli, Staphylococcus aureus, Salmonella typhimurium, Listeria, Pasteurella. The recombinant Plf expressed by recombinant Lactobacillus plantarum showed the best antibacterial activity among all recombinant lactobacillus species. These data represent a basis for the development and application of porcine lactoferrin from recombinant lactobacillus.

  3. Marine Biology Activities. Ocean Related Curriculum Activities.

    Science.gov (United States)

    Pauls, John

    The ocean affects all of our lives. Therefore, awareness of and information about the interconnections between humans and oceans are prerequisites to making sound decisions for the future. Project ORCA (Ocean Related Curriculum Activities) has developed interdisciplinary curriculum materials designed to meet the needs of students and teachers…

  4. Biological activity of acetylated phenolic compounds.

    Science.gov (United States)

    Fragopoulou, Elizabeth; Nomikos, Tzortzis; Karantonis, Haralabos C; Apostolakis, Constantinos; Pliakis, Emmanuel; Samiotaki, Martina; Panayotou, George; Antonopoulou, Smaragdi

    2007-01-10

    In recent years an effort has been made to isolate and identify biologically active compounds that are included in the Mediterranean diet. The existence of naturally occurring acetylated phenolics, as well as studies with synthetic ones, provide evidence that acetyl groups could be correlated with their biological activity. Platelet activating factor (PAF) is implicated in atherosclerosis, whereas its inhibitors seem to play a protective role against cardiovascular disease. The aim of this study was to examine the biological activity of resveratrol and tyrosol and their acetylated derivatives as inhibitors of PAF-induced washed rabbit platelet aggregation. Acetylation of resveratrol and tyrosol was performed, and separation was achieved by HPLC. Acetylated derivatives were identified by negative mass spectrometry. The data showed that tyrosol and its monoacetylated derivatives act as PAF inhibitors, whereas diacetylated derivatives induce platelet aggregation. Resveratrol and its mono- and triacetylated derivatives exert similar inhibitory activity, whereas the diacetylated ones are more potent inhibitors. In conclusion, acetylated phenolics exert the same or even higher antithrombotic activity compared to the biological activity of the initial one.

  5. 75 FR 31795 - Office of Biotechnology Activities; Recombinant DNA Research: Amended Notice of Meeting

    Science.gov (United States)

    2010-06-04

    ... From the Federal Register Online via the Government Publishing Office DEPARTMENT OF HEALTH AND HUMAN SERVICES National Institutes of Health Office of Biotechnology Activities; Recombinant DNA...-Curay, Acting Director, Office of Biotechnology Activities, National Institutes of Health. BILLING...

  6. 76 FR 27653 - Office of Biotechnology Activities; Recombinant DNA Research: Action Under the NIH Guidelines for...

    Science.gov (United States)

    2011-05-12

    ... HUMAN SERVICES National Institutes of Health Office of Biotechnology Activities; Recombinant DNA.... lactis certified host-vector 1 system. In addition, the Office of Biotechnology Activities is updating...: Background documentation and additional information can be obtained from the Office of...

  7. Plant molecular biology and biotechnology research in the post-recombinant DNA era.

    Science.gov (United States)

    Tyagi, Akhilesh K; Khurana, Jitendra P

    2003-01-01

    After the beginning of the recombinant DNA era in the mid-1970s, researchers in India started to make use of the new technology to understand the structure of plant genes and regulation of their expression. The outcome started to appear in print in early the 1980s and genes for histones, tubulin, photosynthetic membrane proteins, phototransduction components, organelles and those regulated differentially by developmental and extrinsic signals were sequenced and characterized. Some genes of biotechnological importance like those encoding an interesting seed protein and the enzyme glyoxalase were also isolated. While work on the characterization of genome structure and organization was started quite early, it remained largely focused on the identification of DNA markers and genetic variability. In this context, the work on mustard, rice and wheat is worth mentioning. In the year 2000, India became a member of the international consortium to sequence entire rice genome. Several laboratories have also given attention to regulated expression of plastid and nuclear genes as well as to isolate target-specific promoters or design promoters with improved potential. Simultaneously, transgenic systems for crops like mustard, rice, wheat, cotton, legumes and several vegetables have been established. More recently, genes of agronomic importance like those for insect resistance, abiotic stress tolerance, nutritional improvement and male sterility, isolated in India or abroad, have been utilized for raising transgenics for crop improvement. Some of these transgenics have already shown their potential in containment facility or limited field trials conducted under the stipulated guidelines. Plant molecular biology and biotechnology are thus clearly poised to make an impact on research in basic biology and agriculture in the near future.

  8. Expression of Recombinant Human Amelogenin in Iranian Lizard Leishmania and Its Biological Function Assay

    Directory of Open Access Journals (Sweden)

    Zahra YADEGARI

    2015-10-01

    Full Text Available Background: Amelogenins are the major components of enamel matrix proteins. Enamel matrix derivatives (EMD can be used in periodontal diseases to regenerate periodontal tissues. The main aim of this study was to evaluate ex-pression of full-length functional recombinant human amelogenin (rhAm in Iranian lizard Leishmania (I.L.L. as an alternative eukaryotic expression system.Methods: Human cDNA encoding a 175-amino acid amelogenin expression cassette was sub cloned into a pLEXSY vector. The construct was transferred into Leishmania cells by electroporation. The protein production was surveyed in the transcription and the translation levels. The expressed protein was purified and some of its biological properties were investigated in comparison to EMD and negative control.Results: Expression of rhAm was confirmed by RT-PCR and western blot test in Leishmania cells. Purified rhAm sig-nificantly inhibited the formation of tartrate-resistant acid phosphatase positive (TRAP+ multinuclear cells in calcitriol stimulated mouse marrow cultures. Moreover, it significantly promoted proliferation and DNA synthesis in L929 mouse fibroblast cells.Conclusion: Functional rhAm was successfully expressed in I.L.L. Easy handling and post translation modification were the main advantages of this expression system. It is suggested to investigate molecular properties of this rhAm in the future.

  9. Pharmaceutical aspects of the recombinant human serum albumin dimer: structural characteristics, biological properties, and medical applications.

    Science.gov (United States)

    Taguchi, Kazuaki; Chuang, Victor Tuan Giam; Maruyama, Toru; Otagiri, Masaki

    2012-09-01

    Human serum albumin is the most abundant protein in the blood. It is clinically used in the treatment of severe hypoalbuminemia and as a plasma expander. The use of albumins as a carrier for drugs is currently being developed, and some are now in the preclinical and clinical trial stages. The main technologies for utilizing an albumin as a drug carrier are protein fusion, polymerization and surface modification, and so on. Among these technologies, albumin dimerization has wide clinical applications as a plasma expander as well as a drug carrier. Despite the fact that many reports have appeared on drugs using an albumin dimer as a carrier, our knowledge of the characteristics of the albumin dimer itself is incomplete. In this review, we summarize the structural characteristics of recombinant albumin dimers produced by two methods, namely, chemical linkage with 1,6-bis(maleimido)hexane and genetically linked with an amino acid linker, and the physicochemical characteristics and biological properties of these preparations. Finally, the potential for pharmaceutical applications of albumin dimers in clinical situations is discussed.

  10. Biological activities of substituted trichostatic acid derivatives

    Indian Academy of Sciences (India)

    Cédric Charrier; Joëlle Roche; Jean-Pierre Gesson; Philippe Bertrand

    2009-07-01

    New substituted trichostatic acid derivatives have been synthesized and evaluated for their biological activities towards the H661 non-small lung cancer cell line. These syntheses were achieved by alkylation of propiophenones to introduce the side chain with a terminal precursor of hydroxamic acid and aminobenzamide derivatives. The first fluorinated derivatives of trichostatic acid are described, such as 6-fluoro trichostatin A, with antiproliferative activities in the micromolar range and with histone deacetylase inhibitory activity.

  11. Optimisation of recombinant production of active human cardiac SERCA2a ATPase.

    Science.gov (United States)

    Antaloae, Ana V; Montigny, Cédric; le Maire, Marc; Watson, Kimberly A; Sørensen, Thomas L-M

    2013-01-01

    Methods for recombinant production of eukaryotic membrane proteins, yielding sufficient quantity and quality of protein for structural biology, remain a challenge. We describe here, expression and purification optimisation of the human SERCA2a cardiac isoform of Ca(2+) translocating ATPase, using Saccharomyces cerevisiae as the heterologous expression system of choice. Two different expression vectors were utilised, allowing expression of C-terminal fusion proteins with a biotinylation domain or a GFP- His8 tag. Solubilised membrane fractions containing the protein of interest were purified onto Streptavidin-Sepharose, Ni-NTA or Talon resin, depending on the fusion tag present. Biotinylated protein was detected using specific antibody directed against SERCA2 and, advantageously, GFP-His8 fusion protein was easily traced during the purification steps using in-gel fluorescence. Importantly, talon resin affinity purification proved more specific than Ni-NTA resin for the GFP-His8 tagged protein, providing better separation of oligomers present, during size exclusion chromatography. The optimised method for expression and purification of human cardiac SERCA2a reported herein, yields purified protein (> 90%) that displays a calcium-dependent thapsigargin-sensitive activity and is suitable for further biophysical, structural and physiological studies. This work provides support for the use of Saccharomyces cerevisiae as a suitable expression system for recombinant production of multi-domain eukaryotic membrane proteins.

  12. The autolytic activity of the recombinant amidase of Staphylococcus saprophyticus is inhibited by its own recombinant GW repeats.

    Science.gov (United States)

    Hell, Wolfgang; Reichl, Sylvia; Anders, Agnes; Gatermann, Sören

    2003-10-10

    The Aas (autolysin/adhesin of Staphylococcus saprophyticus) is a multifunctional surface protein containing two enzymatic domains an N-acetyl-muramyl-L-alanine amidase, an endo-beta-N-acetyl-D-glucosaminidase, and two different regions of repetitive sequences, an N-terminal and a C-terminal repetitive domain. The C-terminal repetitive domain is built up by the repeats R1, R2 and R3, which interconnect the putative active centers of the amidase and glucosaminidase. To investigate the influence of the C-terminal repeats and the N-terminal repeats on the amidase activity, the repetitive domains and fragments of them were cloned and expressed in Escherichia coli. The influence of the different fragments on the activity of the recombinant amidase of the Aas, consisting of the active center of the enzyme and repeat R1, was investigated in a turbidimetric microassay. The different fragments derived from the C-terminal repeats inhibited the amidase activity, while the N-terminal repeats did not influence the activity of the enzyme. The inhibiting activity increased with the number of GW repeats the recombinant fragment contained. Thus we conclude, that the C-terminal GW repeats and not the N-terminal repeats are necessary for the cell wall targeting and the autolytic function of the amidase.

  13. Efficient expression and purification of biologically active human cystatin proteins.

    Science.gov (United States)

    Chauhan, Sakshi; Tomar, Raghuvir S

    2016-02-01

    Cystatins are reversible cysteine protease inhibitor proteins. They are known to play important roles in controlling cathepsins, neurodegenerative disease, and in immune system regulation. Production of recombinant cystatin proteins is important for biochemical and function characterization. In this study, we cloned and expressed human stefin A, stefin B and cystatin C in Escherichia coli. Human stefin A, stefin B and cystatin C were purified from soluble fraction. For cystatin C, we used various chaperone plasmids to make cystatin C soluble, as it is reported to localize in inclusion bodies. Trigger factor, GroES-GroEL, DnaK-DnaJ-GrpE chaperones lead to the presence of cystatin C in the soluble fraction. Immobilized metal affinity chromatography, glutathione sepharose and anion exchange chromatography techniques were employed for efficient purification of these proteins. Their biological activities were tested by inhibition assays against cathepsin L and H3 protease.

  14. Biologically Active Substances from Higher Fungi

    Institute of Scientific and Technical Information of China (English)

    Liu Jikai

    2007-01-01

    As a part of our search for naturally occurring bioactive metabolites of mushrooms, we have investigated the chemical constituents of more than 100 Basidiomycetes and Ascomycetes fungi, and isolated over 300 including 150 new terpenoids,phenolics and nitrogen-containing compounds. The isolation, structural elucidation and biologically activity of the natural products from the higher fungi are briefly discussed.

  15. Biological activities of Curcuma longa L.

    OpenAIRE

    Araújo CAC; LL Leon

    2001-01-01

    There are several data in the literature indicating a great variety of pharmacological activities of Curcuma longa L. (Zingiberaceae), which exhibit anti-inflammatory, anti-human immunodeficiency virus, anti-bacteria, antioxidant effects and nematocidal activities. Curcumin is a major component in Curcuma longa L., being responsible for its biological actions. Other extracts of this plant has been showing potency too. In vitro, curcumin exhibits anti-parasitic, antispasmodic, anti-inflammator...

  16. Prokaryotic Expression and Biological Activity Analysis of Human Ar-resten Gene

    Institute of Scientific and Technical Information of China (English)

    SONG Zifang; ZHENG Qichang; LI Wei; XIONG Jun; SHANG Dan; SHU Xiaogang

    2005-01-01

    To express recombinant arresten in Escherichia coli (E. Coli) and investigate its biological activity, prokaryotic expression vector of human arresten gene was constructed by gene engineering. Human arresten gene was amplified from recombinant plasmid pGEMArr by polymerase chain reaction (PCR), and inserted into prokaryotic expression vector pRSET containing T7 promoter. Restriction analysis and DNA sequencing verified that the arresten gene was correctly cloned into the expression vector. The recombinant plasmid pRSETAt was subsequently transformed into E. Coli BL21 (DE3), and the target gene was expressed under induction of IPTG. SDS-PAGE analysis revealed that the recombinant protein with a molecular weight of 29 kD (1 kD=0. 992 1 ku) amounted to 29 % of the total bacterial proteins. After purification and renaturation, the recombinant protein could significantly suppress the proliferation of human umbilical vein endothelial cells (HUVECs). These results suggested that the expression of a biologically active form of human arresten in the pRSET expression system laid a foundation for further study on the mechanistic insight into arresten action on angiogenesis and the development of powerful anti-cancer drugs.

  17. Loranthus micranthus Linn.: Biological Activities and Phytochemistry

    Directory of Open Access Journals (Sweden)

    Soheil Zorofchian Moghadamtousi

    2013-01-01

    Full Text Available Loranthus micranthus Linn. is a medicinal plant from the Loranthaceae family commonly known as an eastern Nigeria species of the African mistletoe and is widely used in folkloric medicine to cure various ailments and diseases. It is semiparasitic plant because of growing on various host trees and shrubs and absorbing mineral nutrition and water from respective host. Hence, the phytochemicals and biological activities of L. micranthus demonstrated strong host and harvesting period dependency. The leaves have been proved to possess immunomodulatory, antidiabetic, antimicrobial, antihypertensive, antioxidant, antidiarrhoeal, and hypolipidemic activities. This review summarizes the information and findings concerning the current knowledge on the biological activities, pharmacological properties, toxicity, and chemical constituents of Loranthus micranthus.

  18. Biologically active substances of stone crop fruits

    Directory of Open Access Journals (Sweden)

    Margarita Makarkina

    2014-04-01

    Full Text Available Stone fruit crops are successfully cultivated in many regions of Russia. Their fruits contain a great diversity of biologically active and mineral substances. The stone fruit varietal collection of the All Russia Research Institute of Fruit Crop Breeding has been estimated on the content of biologically active substances (ascorbic acid and phenolic compounds in fruits: 80 sour cherry varieties, 28 sweet cherry varieties, 29 plum varieties and 24 apricot varieties. High cultivar variability of the content of ascorbic acid and P-active sub-stances in fruits has been determined in each crop. The best genotypes have been singled out according to each biochemical component and a complex of characters.

  19. The biologic properties of recombinant human thrombopoietin in the proliferation and megakaryocytic differentiation of acute myeloblastic leukemia cells.

    Science.gov (United States)

    Matsumura, I; Kanakura, Y; Kato, T; Ikeda, H; Horikawa, Y; Ishikawa, J; Kitayama, H; Nishiura, T; Tomiyama, Y; Miyazaki, H; Matsuzawa, Y

    1996-10-15

    Thrombopoietin (TPO) is implicated as a primary regulator of megakaryopoiesis and thrombopoiesis. However, the biologic effects of TPO on human acute myeloblastic leukemia (AML) cells are largely unknown. To determine if recombinant human (rh) TPO has proliferation-supporting and differentiation-inducing activities in AML cells, 15 cases of AML cells that were exclusively composed of undifferentiated leukemia cells and showed growth response to rhTPO in a short-term culture (72 hours) were subjected to long-term suspension culture with or without rhTPO. Of 15 cases, rhTPO supported proliferation of AML cells for 2 to 4 weeks in 4 cases whose French-American-British subtypes were M0, M2, M4, and M7, respectively. In addition to the proliferation-supporting activity, rhTPO was found to induce AML cells to progress to some degree of megakaryocytic differentiation at both morphologic and surface-phenotypic level in 2 AML cases with M0 and M7 subtypes. The treatment of AML cells with rhTPO resulted in rapid tyrosine phosphorylation of the TPO-receptor, c-mpl, and STAT3 in all of cases tested. By contrast, the expression of erythroid/megakaryocyte-specific transcription factors (GATA-1, GATA-2, and NF-E2) was markedly induced or enhanced in only 2 AML cases that showed megakaryocytic differentiation in response to rhTPO. These results suggested that, at least in a fraction of AML cases, TPO could not only support the proliferation of AML cells irrespective of AML subtypes, but could also induce megakaryocytic differentiation, possibly through activation of GATA-1, GATA-2, and NF-E2.

  20. Glycosides from Marine Sponges (Porifera, Demospongiae: Structures, Taxonomical Distribution, Biological Activities and Biological Roles

    Directory of Open Access Journals (Sweden)

    Valentin A. Stonik

    2012-08-01

    Full Text Available Literature data about glycosides from sponges (Porifera, Demospongiae are reviewed. Structural diversity, biological activities, taxonomic distribution and biological functions of these natural products are discussed.

  1. Glycosides from marine sponges (Porifera, Demospongiae): structures, taxonomical distribution, biological activities and biological roles.

    Science.gov (United States)

    Kalinin, Vladimir I; Ivanchina, Natalia V; Krasokhin, Vladimir B; Makarieva, Tatyana N; Stonik, Valentin A

    2012-08-01

    Literature data about glycosides from sponges (Porifera, Demospongiae) are reviewed. Structural diversity, biological activities, taxonomic distribution and biological functions of these natural products are discussed.

  2. Estrogenic flavonoids: structural requirements for biological activity.

    Science.gov (United States)

    Miksicek, R J

    1995-01-01

    A systematic survey of polycyclic phenols has been performed to identify members of this chemical group with estrogenic activity. Twelve compounds were found to be able to stimulate the transcriptional activity of the human estrogen receptor expressed in cultured cells by transient transfection. These natural estrogens belong to several distinct, but chemically related classes including chalcones, flavanones, flavones, flavonols, and isoflavones. Selected examples of estrogenic flavonoids were further analyzed to determine their biological potencies and their relative affinities for binding to the estrogen receptor. These data are interpreted with respect to the molecular structure of polycyclic phenols required for hormonal activity as nonsteroidal estrogens.

  3. Efficient genome engineering by targeted homologous recombination in mouse embryos using transcription activator-like effector nucleases.

    Science.gov (United States)

    Sommer, Daniel; Peters, Annika; Wirtz, Tristan; Mai, Maren; Ackermann, Justus; Thabet, Yasser; Schmidt, Jürgen; Weighardt, Heike; Wunderlich, F Thomas; Degen, Joachim; Schultze, Joachim L; Beyer, Marc

    2014-01-01

    Generation of mouse models by introducing transgenes using homologous recombination is critical for understanding fundamental biology and pathology of human diseases. Here we investigate whether artificial transcription activator-like effector nucleases (TALENs)-powerful tools that induce DNA double-strand breaks at specific genomic locations-can be combined with a targeting vector to induce homologous recombination for the introduction of a transgene in embryonic stem cells and fertilized murine oocytes. We describe the generation of a conditional mouse model using TALENs, which introduce double-strand breaks at the genomic locus of the special AT-rich sequence-binding protein-1 in combination with a large 14.4 kb targeting template vector. We report successful germline transmission of this allele and demonstrate its recombination in primary cells in the presence of Cre-recombinase. These results suggest that TALEN-assisted induction of DNA double-strand breaks can facilitate homologous recombination of complex targeting constructs directly in oocytes.

  4. Isoflavones: estrogenic activity, biological effect and bioavailability.

    Science.gov (United States)

    Vitale, Daniela Cristina; Piazza, Cateno; Melilli, Barbara; Drago, Filippo; Salomone, Salvatore

    2013-03-01

    Isoflavones are phytoestrogens with potent estrogenic activity; genistein, daidzein and glycitein are the most active isoflavones found in soy beans. Phytoestrogens have similarity in structure with the human female hormone 17-β-estradiol, which can bind to both alpha and beta estrogen receptors, and mimic the action of estrogens on target organs, thereby exerting many health benefits when used in some hormone-dependent diseases. Numerous clinical studies claim benefits of genistein and daidzein in chemoprevention of breast and prostate cancer, cardiovascular disease and osteoporosis as well as in relieving postmenopausal symptoms. The ability of isoflavones to prevent cancer and other chronic diseases largely depends on pharmacokinetic properties of these compounds, in particular absorption and distribution to the target tissue. The chemical form in which isoflavones occur is important because it influences their bioavailability and, therefore, their biological activity. Glucose-conjugated isoflavones are highly polar, water-soluble compounds. They are hardly absorbed by the intestinal epithelium and have weaker biological activities than the corresponding aglycone. Different microbial families of colon can transform glycosylated isoflavones into aglycones. Clinical studies show important differences between the aglycone and conjugated forms of genistein and daidzein. The evaluation of isoflavone metabolism and bioavailability is crucial to understanding their biological effects. Lipid-based formulations such as drug incorporation into oils, emulsions and self-microemulsifying formulations have been introduced to increase bioavailability. Complexation with cyclodextrin also represent a valid method to improve the physicochemical characteristics of these substances in order to be absorbed and distributed to target tissues. We review and discuss pharmacokinetic issues that critically influence the biological activity of isoflavones.

  5. Collaborative Learning in Biology: Debating the Ethics of Recombinant DNA Technology.

    Science.gov (United States)

    Anderson, Rodney P.

    1998-01-01

    Discusses applications of recombinant DNA technology and the controversies surrounding that technique. Provides a cooperative learning project idea that involves teams of students investigating and debating these issues. (DDR)

  6. Spilanthol: occurrence, extraction, chemistry and biological activities

    Directory of Open Access Journals (Sweden)

    Alan F. Barbosa

    2016-02-01

    Full Text Available Abstract Spilanthol (C14H23NO, 221.339 g/mol is a bioactive compound that is found in many different plants that are used as traditional remedies throughout the world. It is present in Heliopsis longipes and several species in the genus Acmella, including A. oleracea L., also known as paracress and jambu. Its leaves and flowers have sensory properties (pungency, tingling, numbing, mouth-watering that make it a popular spice and ingredient in several Brazilian dishes. Spilanthol can exert a variety of biological and pharmacological effects including analgesic, neuroprotective, antioxidant, antimutagenic, anti-cancer, anti-inflammatory, antimicrobial, antilarvicidal and insecticidal activities. So, the aim of this review is to present a literature review on the spilanthol that describes its occurrence, chemistry, extraction and biological activities.

  7. Monitoring Biological Activity at Geothermal Power Plants

    Energy Technology Data Exchange (ETDEWEB)

    Peter Pryfogle

    2005-09-01

    The economic impact of microbial growth in geothermal power plants has been estimated to be as high as $500,000 annually for a 100 MWe plant. Many methods are available to monitor biological activity at these facilities; however, very few plants have any on-line monitoring program in place. Metal coupon, selective culturing (MPN), total organic carbon (TOC), adenosine triphosphate (ATP), respirometry, phospholipid fatty acid (PLFA), and denaturing gradient gel electrophoresis (DGGE) characterizations have been conducted using water samples collected from geothermal plants located in California and Utah. In addition, the on-line performance of a commercial electrochemical monitor, the BIoGEORGE?, has been evaluated during extended deployments at geothermal facilities. This report provides a review of these techniques, presents data on their application from laboratory and field studies, and discusses their value in characterizing and monitoring biological activities at geothermal power plants.

  8. Biological Activities of Polyphenols from Grapes

    OpenAIRE

    Hua-Bin Li; Ya-Jun Guo; Gui-Fang Deng; En-Qin Xia

    2010-01-01

    The dietary consumption of grape and its products is associated with a lower incidence of degenerative diseases such as cardiovascular disease and certain types of cancers. Most recent interest has focused on the bioactive phenolic compounds in grape. Anthocyanins, flavanols, flavonols and resveratrol are the most important grape polyphenols because they possess many biological activities, such as antioxidant, cardioprotective, anticancer, anti-inflammation, antiaging and antimicrobial proper...

  9. ATPase activity tightly regulates RecA nucleofilaments to promote homologous recombination

    Science.gov (United States)

    Zhao, Bailin; Zhang, Dapeng; Li, Chengmin; Yuan, Zheng; Yu, Fangzhi; Zhong, Shangwei; Jiang, Guibin; Yang, Yun-Gui; Le, X Chris; Weinfeld, Michael; Zhu, Ping; Wang, Hailin

    2017-01-01

    Homologous recombination (HR), catalyzed in an evolutionarily conserved manner by active RecA/Rad51 nucleofilaments, maintains genomic integrity and promotes biological evolution and diversity. The structures of RecA/Rad51 nucleofilaments provide information critical for the entire HR process. By exploiting a unique capillary electrophoresis-laser-induced fluorescence polarization assay, we have discovered an active form of RecA nucleofilament, stimulated by ATP hydrolysis, that contains mainly unbound nucleotide sites. This finding was confirmed by a nuclease protection assay and electron microscopy (EM) imaging. We further found that these RecA-unsaturated filaments promote strand exchange in vitro and HR in vivo. RecA mutants (P67D and P67E), which only form RecA-unsaturated nucleofilaments, were able to mediate HR in vitro and in vivo, but mutants favoring the formation of the saturated nucleofilaments failed to support HR. We thus present a new model for RecA-mediated HR in which RecA utilizes its intrinsic DNA binding-dependent ATPase activity to remodel the nucleofilaments to a less saturated form and thereby promote HR. PMID:28101376

  10. Novel biological activity of ameloblastin in enamel matrix derivative

    Directory of Open Access Journals (Sweden)

    Sachiko KURAMITSU-FUJIMOTO

    2015-02-01

    Full Text Available Objective Enamel matrix derivative (EMD is used clinically to promote periodontal tissue regeneration. However, the effects of EMD on gingival epithelial cells during regeneration of periodontal tissues are unclear. In this in vitro study, we purified ameloblastin from EMD and investigated its biological effects on epithelial cells. Material and Methods Bioactive fractions were purified from EMD by reversed-phase high-performance liquid chromatography using hydrophobic support with a C18 column. The mouse gingival epithelial cell line GE-1 and human oral squamous cell carcinoma line SCC-25 were treated with purified EMD fraction, and cell survival was assessed with a WST-1 assay. To identify the proteins in bioactive fractions of EMD, we used proteome analysis with two-dimensional gel electrophoresis followed by identification with liquid chromatography-tandem mass spectrometry (LC-MS/MS analysis. Results Purified fractions from EMD suppressed proliferation of GE-1 and SCC-25. LC-MS/MS revealed that ameloblastin in EMD is the component responsible for inhibiting epithelial cell proliferation. The inhibitory effect of ameloblastin on the proliferation of GE-1 and SCC-25 was confirmed using recombinant protein. Conclusion The inhibitory effects of EMD on epithelial cell proliferation are caused by the biological activities of ameloblastin, which suggests that ameloblastin is involved in regulating epithelial downgrowth in periodontal tissues.

  11. Biologic activity of digoxin-specific antisera.

    Science.gov (United States)

    Watson, J F; Butler, V P

    1972-03-01

    Digoxin-specific antibodies are capable of removing essentially all intracellular digoxin from rat renal cortical slices or from human erythrocytes. In removing digoxin from erythrocytes, these antibodies are capable of reversing an effect of the drug on cellular potassium transport. This study provides direct evidence that antibodies are capable of removing, and thereby reversing the biological effect of, physiologically active low molecular weight substances after they have been taken up by mammalian cells. This biologic property of digoxin-specific antibodies suggests that autidigoxin sera may prove useful in the reversal of digoxin toxicity. Rapid and essentially quantitative removal of digoxin from red cells by antibody is not accompanied by an immediate restoration of patassium influx to normal levels. Identification of the mechanism of this phenomenon may provide useful information concerning the mode of action not only of digoxin, but also of the cation transport system of human erythrocytes.

  12. Biological Activities of Royal Jelly - Review

    Directory of Open Access Journals (Sweden)

    Crenguţa I. Pavel

    2011-10-01

    Full Text Available Royal jelly is a secretion product of the cephalic glands of nurse bees that has been used for centuries for itsextraordinary properties and health effects. This bibliographic study aims to review many of the scientific findingsand research that prove many of the remarkable various actions, effects and some uses of royal jelly. There are takeninto consideration numerous biological properties and effects of royal jelly: antioxidant, neurotrophic, hipoglicemiant, hipocholesterolemiant and hepatoprotective, hypotensive and blood pressure regulatory, antitumor, antibiotic, anti-inflammatory, immunomodulatory and anti-allergic, general tonic and antiaging. Royal jelly is one ofthe most studied bee products, but there still remains much to reveal about its biochemistry and biological activity infuture research for our health and life benefit.

  13. Molecular mechanism of recombinant liver fatty acid binding protein's antioxidant activity

    OpenAIRE

    YAN, JING; Gong, Yuewen; She, Yi-Min; Wang, Guqi; Roberts, Michael S; Burczynski, Frank J.

    2009-01-01

    Hepatocytes expressing liver fatty acid binding protein (L-FABP) are known to be more resistant to oxidative stress than those devoid of this protein. The mechanism for the observed antioxidant activity is not known. We examined the antioxidant mechanism of a recombinant rat L-FABP in the presence of a hydrophilic (AAPH) or lipophilic (AMVN) free radical generator. Recombinant L-FABP amino acid sequence and its amino acid oxidative products following oxidation were identified by MALDI quadrup...

  14. 78 FR 12074 - Office of Biotechnology Activities; Recombinant DNA Research: Actions Under the NIH Guidelines...

    Science.gov (United States)

    2013-02-21

    ... HUMAN SERVICES National Institutes of Health Office of Biotechnology Activities; Recombinant DNA... recommendations of the RAC, the NIH Office of Biotechnology Activities (OBA) concluded that more specific guidance... address or by fax at 301-496-9839 or by mail to the Office of Biotechnology Activities,...

  15. 75 FR 21008 - Office of Biotechnology Activities; Recombinant DNA Research: Proposed Actions Under the NIH...

    Science.gov (United States)

    2010-04-22

    ... HUMAN SERVICES National Institutes of Health Office of Biotechnology Activities; Recombinant DNA... the NIH Guidelines. SUMMARY: In March 2009, the NIH Office of Biotechnology Activities (OBA) published... e-mail address or by fax to 301-496-9839 or mail to the Office of Biotechnology Activities,...

  16. 75 FR 69687 - Office of Biotechnology Activities Recombinant DNA Research: Proposed Actions Under the NIH...

    Science.gov (United States)

    2010-11-15

    ... of Biotechnology Activities Recombinant DNA Research: Proposed Actions Under the NIH Guidelines for... system has been submitted to the NIH Office of Biotechnology Activities (OBA). The data to be considered... Biotechnology Activities, National Institutes of Health, 6705 Rockledge Drive, Suite 750, MSC 7985,...

  17. 76 FR 62816 - Office of Biotechnology Activities; Recombinant DNA Research: Action Under the NIH Guidelines for...

    Science.gov (United States)

    2011-10-11

    ... HUMAN SERVICES National Institutes of Health Office of Biotechnology Activities; Recombinant DNA... Biotechnology Activities (OBA) is updating Appendix B of the NIH Guidelines to specify the risk group (RG...: October 3, 2011. Jacqueline Corrigan-Curay, Acting Director, Office of Biotechnology Activities,...

  18. Biological activities of Curcuma longa L.

    Directory of Open Access Journals (Sweden)

    CAC Araújo

    2001-07-01

    Full Text Available There are several data in the literature indicating a great variety of pharmacological activities of Curcuma longa L. (Zingiberaceae, which exhibit anti-inflammatory, anti-human immunodeficiency virus, anti-bacteria, antioxidant effects and nematocidal activities. Curcumin is a major component in Curcuma longa L., being responsible for its biological actions. Other extracts of this plant has been showing potency too. In vitro, curcumin exhibits anti-parasitic, antispasmodic, anti-inflammatory and gastrointestinal effects; and also inhibits carcinogenesis and cancer growth. In vivo, there are experiments showing the anti-parasitic, anti-inflammatory potency of curcumin and extracts of C. longa L. by parenteral and oral application in animal models. In this present work we make an overview of the pharmacological activities of C. longa L., showing its importance.

  19. Biological Activities of Polyphenols from Grapes

    Directory of Open Access Journals (Sweden)

    Hua-Bin Li

    2010-02-01

    Full Text Available The dietary consumption of grape and its products is associated with a lower incidence of degenerative diseases such as cardiovascular disease and certain types of cancers. Most recent interest has focused on the bioactive phenolic compounds in grape. Anthocyanins, flavanols, flavonols and resveratrol are the most important grape polyphenols because they possess many biological activities, such as antioxidant, cardioprotective, anticancer, anti-inflammation, antiaging and antimicrobial properties. This review summarizes current knowledge on the bioactivities of grape phenolics. The extraction, isolation and identification methods of polyphenols from grape as well as their bioavailability and potential toxicity also are included.

  20. Reconstructing Causal Biological Networks through Active Learning.

    Directory of Open Access Journals (Sweden)

    Hyunghoon Cho

    Full Text Available Reverse-engineering of biological networks is a central problem in systems biology. The use of intervention data, such as gene knockouts or knockdowns, is typically used for teasing apart causal relationships among genes. Under time or resource constraints, one needs to carefully choose which intervention experiments to carry out. Previous approaches for selecting most informative interventions have largely been focused on discrete Bayesian networks. However, continuous Bayesian networks are of great practical interest, especially in the study of complex biological systems and their quantitative properties. In this work, we present an efficient, information-theoretic active learning algorithm for Gaussian Bayesian networks (GBNs, which serve as important models for gene regulatory networks. In addition to providing linear-algebraic insights unique to GBNs, leading to significant runtime improvements, we demonstrate the effectiveness of our method on data simulated with GBNs and the DREAM4 network inference challenge data sets. Our method generally leads to faster recovery of underlying network structure and faster convergence to final distribution of confidence scores over candidate graph structures using the full data, in comparison to random selection of intervention experiments.

  1. Classification of Recombinant Biologics in the EU: Divergence between National Pharmacovigilance Centers

    NARCIS (Netherlands)

    Klein, Kevin; De Bruin, Marie L.; Broekmans, Andre W.; Stolk, Pieter

    2015-01-01

    Background and Objective: Biological medicinal products (biologics) are subject to specific pharmacovigilance requirements to ensure that biologics are identifiable by brand name and batch number in adverse drug reaction (ADR) reports. Since Member States collect ADR data at the national level befor

  2. Recombinant dioscorins of the yam storage protein expressed in Escherichia coli exhibit antioxidant and immunomodulatory activities.

    Science.gov (United States)

    Jheng, Yi-Jyun; Tsai, Wei-Yi; Chen, Kuo-Hsuan; Lin, Kuo-Wei; Chyan, Chia-Lin; Yang, Ching-Chi; Lin, Kuo-Chih

    2012-09-01

    Dioscorins, the major storage proteins in yam tubers, exhibit biochemical and immunomodulatroy activities. To investigate the potential application of dioscorins in biomedical research, we expressed the dioscorin genes Dj-dioA3 and Dp-dioA2 from Dioscorea japonica and Dioscorea pseudojaponica, respectively, in E. coli and routinely obtained approximately 15 mg proteins per liter Escherichia coli culture (mg/L) to 30 mg/L of rDj-dioscorinA3 and 4 to 8 mg/L of rDp-dioscorinA2. Western blot analyses revealed that both recombinant dioscorins contained epitopes with similar antigenicities to those of the native dioscorins. Results from dithiothreitol (DTT) treatment followed by monobromobimane (mBBr) staining showed that both recombinant dioscorins, like the native dioscorins, contain an intramolecular disulfide bond between Cys(28) and Cys(187) residues. Circular dichroism spectroscopy findings indicated that the secondary structural contents of the recombinant dioscorins showed high similarity to those of their corresponding native dioscorins. Both recombinant dioscorins, like the native dioscorins, exhibited 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging and Toll-like receptor 4 signaling activities, and stimulated the phagocytosis of E. coli by macrophage. Overall, our results indicated that substantial amounts of recombinant dioscorins can be purified easily from E. coli and that these recombinant dioscorins are appropriate for application in future investigations of the biomedical functions of dioscorins.

  3. Biologically Active Metabolites Synthesized by Microalgae

    Directory of Open Access Journals (Sweden)

    Michele Greque de Morais

    2015-01-01

    Full Text Available Microalgae are microorganisms that have different morphological, physiological, and genetic traits that confer the ability to produce different biologically active metabolites. Microalgal biotechnology has become a subject of study for various fields, due to the varied bioproducts that can be obtained from these microorganisms. When microalgal cultivation processes are better understood, microalgae can become an environmentally friendly and economically viable source of compounds of interest, because production can be optimized in a controlled culture. The bioactive compounds derived from microalgae have anti-inflammatory, antimicrobial, and antioxidant activities, among others. Furthermore, these microorganisms have the ability to promote health and reduce the risk of the development of degenerative diseases. In this context, the aim of this review is to discuss bioactive metabolites produced by microalgae for possible applications in the life sciences.

  4. High cytokine production and effective antitumor activity of a recombinant vaccinia virus encoding murine interleukin 12.

    Science.gov (United States)

    Meko, J B; Yim, J H; Tsung, K; Norton, J A

    1995-11-01

    We have constructed a recombinant vaccinia virus (recVV), vKT0334 mIL-12, containing the genes encoding the p35 and p40 subunits of murine interleukin-12 (mIL-12). In vitro experiments demonstrated that vKT0334 mIL-12 efficiently infected a variety of murine and human tumor cell lines and produced very high amounts (1.5 micrograms/10(6) cells/24 h) of biologically active mIL-12. Mice injected s.c. with 10(6) MCA 105 sarcoma cells, followed by injection at the same site with saline or a control recVV, vKT033, containing no mIL-12 genes, all developed progressively growing tumor, whereas 60% of animals injected with vKT0334 mIL-12 remained tumor free (P < 0.0005). Furthermore, tumor growth was significantly reduced in the remaining mice treated with vKT0334 mIL-12 that did develop tumor compared with mice treated with vKT033 (P < 0.03) or saline (P < 0.0001). We conclude that recVV expressing high levels of mIL-12 offers an effective in vivo method of cytokine gene delivery and expression in tumors with subsequent antitumor effect.

  5. 75 FR 42114 - Office of Biotechnology Activities; Recombinant DNA Research: Proposed Action Under the NIH...

    Science.gov (United States)

    2010-07-20

    ... HUMAN SERVICES National Institutes of Health Office of Biotechnology Activities; Recombinant DNA... transgenic rodent and a non-transgenic rodent). The NIH Office of Biotechnology Activities (OBA) received a... to the same email address or by fax to 301-496-9839 or mail to the Office of Biotechnology...

  6. 76 FR 3150 - Office of Biotechnology Activities; Recombinant DNA Research: Action Under the NIH Guidelines for...

    Science.gov (United States)

    2011-01-19

    ... HUMAN SERVICES National Institutes of Health Office of Biotechnology Activities; Recombinant DNA...). On July 20, 2010 the NIH Office of Biotechnology Activities (OBA) published a proposed action (75 FR... contact OBA by e- mail at oba@od.nih.gov , telephone, 301-496-9838 or mail to the Office of...

  7. Litchi Flavonoids: Isolation, Identification and Biological Activity

    Directory of Open Access Journals (Sweden)

    Yueming Jiang

    2007-04-01

    Full Text Available The current status of the isolation, identification, biological activity, utilization and development prospects of flavonoids found in litchi fruit pericarp (LFP tissues is reviewed. LFP tissues account for approximately 15% by weight of the whole fresh fruit and are comprised of significant amount of flavonoids. The major flavonoids in ripe LFP include flavonols and anthocyanins. The major flavanols in the LFP are reported to be procyanidin B4, procyanidin B2 and epicatechin, while cyanindin-3-rutinside, cyanidin-3-glucoside, quercetin-3-rutinosde and quercetin-3-glucoside are identified as the important anthocyanins. Litchi flavanols and anthocyanins exhibit good potential antioxidant activity. The hydroxyl radical and superoxide anion scavenging activities of procyanidin B2 are greater than those of procyanidin B4 and epicatechin, while epicatechin has the highest α,α-diphenyl-β-picrylhydrazyl radical (DPPH· scavenging activity. In addition to the antioxidant activity, LFP extract displays a dose- and time-dependent inhibitory effect on human breast cancer, which could be attributed, in part, to its inhibition of proliferation and induction of apoptosis in cancer cells through upregulation and down-regulation of multiple genes. Furthermore, various anticancer activities are observed for epicatechin, procyanidin B2, procyanidin B4 and the ethyl acetate fraction of LFP tissue extracts. Procyanidin B4 and the ethyl acetate fraction show a stronger inhibitory effect on HELF than MCF-7 proliferation, while epicatechin and procyanidin B2 have lower cytotoxicities towards MCF-7 and HELF than paclitaxel. It is therefore suggested that flavonoids from LFP might be potentially useful components for functional foods and/or anti-breast cancer drugs.

  8. ACTIVE AND PARTICIPATORY METHODS IN BIOLOGY: MODELING

    Directory of Open Access Journals (Sweden)

    Brînduşa-Antonela SBÎRCEA

    2011-01-01

    Full Text Available By using active and participatory methods it is hoped that pupils will not only come to a deeper understanding of the issues involved, but also that their motivation will be heightened. Pupil involvement in their learning is essential. Moreover, by using a variety of teaching techniques, we can help students make sense of the world in different ways, increasing the likelihood that they will develop a conceptual understanding. The teacher must be a good facilitator, monitoring and supporting group dynamics. Modeling is an instructional strategy in which the teacher demonstrates a new concept or approach to learning and pupils learn by observing. In the teaching of biology the didactic materials are fundamental tools in the teaching-learning process. Reading about scientific concepts or having a teacher explain them is not enough. Research has shown that modeling can be used across disciplines and in all grade and ability level classrooms. Using this type of instruction, teachers encourage learning.

  9. Biologically Active and Antimicrobial Peptides from Plants

    Directory of Open Access Journals (Sweden)

    Carlos E. Salas

    2015-01-01

    Full Text Available Bioactive peptides are part of an innate response elicited by most living forms. In plants, they are produced ubiquitously in roots, seeds, flowers, stems, and leaves, highlighting their physiological importance. While most of the bioactive peptides produced in plants possess microbicide properties, there is evidence that they are also involved in cellular signaling. Structurally, there is an overall similarity when comparing them with those derived from animal or insect sources. The biological action of bioactive peptides initiates with the binding to the target membrane followed in most cases by membrane permeabilization and rupture. Here we present an overview of what is currently known about bioactive peptides from plants, focusing on their antimicrobial activity and their role in the plant signaling network and offering perspectives on their potential application.

  10. Characterization of ATPase Activity of Recombinant Human Pif1

    Institute of Scientific and Technical Information of China (English)

    Yu HUANG; Deng-Hong ZHANG; Jin-Qiu ZHOU

    2006-01-01

    Saccharomyces cerevisiae Pif1p helicase is the founding member of the Pif1 subfamily that is conserved from yeast to human. The potential human homolog of the yeast PIF1 gene has been cloned from the cDNA library of the Hek293 cell line. Here, we described a purification procedure of glutathione Stransferase (GST)-fused N terminal truncated human Pif1 protein (hPif1△N) from yeast and characterized the enzymatic kinetics of its ATP hydrolysis activity. The ATPase activity of human Pif1 is dependent on divalent cation, such as Mg2+, Ca2+ and single-stranded DNA. Km for ATP for the ATPase activity is approximately 200 μM. As the ATPase activity is essential for hPif1's helicase activity, these results will facilitate the further investigation on hPif1.

  11. Biological active compounds from Georgian Galanthus shaoricus.

    Science.gov (United States)

    Jokhadze, M; Kuchukhidze, J; Chincharadze, D; Murtazashvili, T

    2011-10-01

    Amaryllidaceae alkaloids exhibit antitumour, antiviral and anticholinergic activities. Some of them have been used in the treatment of myasthenia gravis, myopathy and diseases of the nervous system. In this study, the characterization of these compounds from Amaryllidaceae plants along with some biological activities and some regulations to conserve the native flora will be reviewed. Plants materials: Galanthus shaoricus Kem.-Nath., were collected in 2007-2008 during the flowering period in Georgia. The preparation of extracts and fractions were obtained using methanolic maceration. Crude alkaloidal extracts were typically purified by liquid-liquid partitioning of their basic forms in chloroform. Lycorine, galantamine and tazettine has been found as one of the major alkaloid from Amaryllidaceae plants. Galanthus shaoricus have shown good antimalarial and cytotoxic activity in a dose-dependent manner. Methanolic extracts from bulbs demonstrated significant growth inhibition on human Hela and HCT-116 cells lines with IC50 (μg/mL) 16.3±1.8; 22.1±2.9 (aerial parts) and 12.8±1.7; 16.5±1.9 (Bulbs), respectively. Concerning the Amaryllidaceae alkaloids, lycorine with IC50 (μM) 0.8±0.5 and 2.6±0.2, haemantaimene (IC50=1.1±0.7 and 2.7±0.8 μM), hamaine (IC50=3.4±1.0 and 6.2 ±1.4 μM), homolycorine (IC50=1.4±0.9 and 3.3±1.0 μM), hipeastrine (IC50=2.8±1.0 and 7.5±1.8 μM) were found to be responsible for the cytotoxic activity on HCT-116 and Hela cell lines, respectively.

  12. Sulfation and biological activities of konjac glucomannan.

    Science.gov (United States)

    Bo, Surina; Muschin, Tegshi; Kanamoto, Taisei; Nakashima, Hideki; Yoshida, Takashi

    2013-05-15

    The sulfation of konjac glucomannan and its anti-HIV and blood anticoagulant activities were investigated. Konjac glucomannan is a polysaccharide occurring naturally in konjac plant tubers and has high molecular weights. Solubility in water is very low, and the aqueous solutions at low concentrations have high viscosity. Before sulfation, hydrolysis by diluted sulfuric acid was carried out to decrease the molecular weights of M¯n=19.2 × 10(4)-0.2 × 10(4). Sulfation with piperidine-N-sulfonic acid or SO3-pyridine complex gave sulfated konjac glucomannans with molecular weights of M¯n=1.0 × 10(4)-0.4 × 10(4) and degrees of sulfation (DS) of 1.3-1.4. It was found that the sulfated konjac glucomannans had potent anti-HIV activity at a 50% effective concentration, (EC50) of 1.2-1.3 μg/ml, which was almost as high as that of an AIDS drug, ddC, whose EC50=3.2 μg/ml, and moderate blood anticoagulant activity, AA=0.8-22.7 units/mg, compared to those of standard sulfated polysaccharides, curdlan (10 units/mg) and dextran (22.7 units/mg) sulfates. Structural analysis of sulfated konjac glucomannans with negatively charged sulfated groups was performed by high resolution NMR, and the interaction between poly-l-lysine with positively charged amino groups as a model compound of proteins and peptides was measured by surface plasmon resonance measurement, suggesting that the sulfated konjac glucomannans had a high binding stability on immobilized poly-l-lysine. The binding of sulfated konjac glucomannan was concentration-dependent, and the biological activity of the sulfated konjac glucomannans may be due to electrostatic interaction between the sulfate and amino groups.

  13. Expression of a functional recombinant Phoneutria nigriventer toxin active on K+ channels.

    Science.gov (United States)

    Carneiro, A M D; Kushmerick, C; Koenen, J; Arndt, M H L; Cordeiro, M N; Chavez-Olortegui, C; Diniz, C R; Gomez, M V; Kalapothakis, E; Prado, M A M; Prado, V F

    2003-03-01

    PnTx3-1 is a peptide isolated from the venom of the spider Phoneutria nigriventer that specifically inhibits A-type K(+) currents (I(A)) in GH(3) cells. Here we used a bacterial expression system to produce an NH(2)-extended mutant of PnTx3-1 (ISEF-PnTx3-1) and tested whether the toxin is functional. The recombinant toxin was purified from bacterial extracts by a combination of affinity and ion-exchange chromatography. The recombinant toxin blocked A-type K(+) currents in GH(3) cells in a fashion similar to that observed with the wild-type toxin purified from the spider venom. These results suggest that recombinant cDNA methods provide a novel source for the production of functional Phoneutria toxins. The recombinant ISEF-PnTx3-1 should be useful for further understanding of the role of A-type K(+) currents in biological processes.

  14. Patterns of recombination activity on mouse chromosome 11 revealed by high resolution mapping.

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    Timothy Billings

    Full Text Available The success of high resolution genetic mapping of disease predisposition and quantitative trait loci in humans and experimental animals depends on the positions of key crossover events around the gene of interest. In mammals, the majority of recombination occurs at highly delimited 1-2 kb long sites known as recombination hotspots, whose locations and activities are distributed unevenly along the chromosomes and are tightly regulated in a sex specific manner. The factors determining the location of hotspots started to emerge with the finding of PRDM9 as a major hotspot regulator in mammals, however, additional factors modulating hotspot activity and sex specificity are yet to be defined. To address this limitation, we have collected and mapped the locations of 4829 crossover events occurring on mouse chromosome 11 in 5858 meioses of male and female reciprocal F1 hybrids of C57BL/6J and CAST/EiJ mice. This chromosome was chosen for its medium size and high gene density and provided a comparison with our previous analysis of recombination on the longest mouse chromosome 1. Crossovers were mapped to an average resolution of 127 kb, and thirteen hotspots were mapped to <8 kb. Most crossovers occurred in a small number of the most active hotspots. Females had higher recombination rate than males as a consequence of differences in crossover interference and regional variation of sex specific rates along the chromosome. Comparison with chromosome 1 showed that recombination events tend to be positioned in similar fashion along the centromere-telomere axis but independently of the local gene density. It appears that mammalian recombination is regulated on at least three levels, chromosome-wide, regional, and at individual hotspots, and these regulation levels are influenced by sex and genetic background but not by gene content.

  15. Base composition, selection, and phylogenetic significance of indels in the recombination activating gene-1 in vertebrates

    NARCIS (Netherlands)

    Chiari, Y.; Meijden, van der A.; Madsen, O.; Vences, M.; Meyer, A.

    2009-01-01

    Background: The Recombination Activating Proteins, RAG1 and RAG2, play a crucial role in the immune response in vertebrates. Among the nuclear markers currently used for phylogenetic purposes, Rag1 has especially enjoyed enormous popularity, since it successfully contributed to elucidating the relat

  16. Evidence for reduced charge recombination in carbon nanotube/perovskite-based active layers

    Science.gov (United States)

    Bag, Monojit; Renna, Lawrence A.; Jeong, Seung Pyo; Han, Xu; Cutting, Christie L.; Maroudas, Dimitrios; Venkataraman, D.

    2016-10-01

    Using impedance spectroscopy and computation, we show that incorporation of multi-walled carbon nanotubes (MWCNTs) in the bulk of the active layer of perovskite-based solar cells reduces charge recombination and increases the open circuit voltage. An ∼87% reduction in recombination was achieved when MWCNTs were introduced in the planar-heterostructure perovskite solar cell containing mixed counterions. The open circuit voltage (Voc) of perovskite/MWCNTs devices was increased by 70 mV, while the short circuit current density (Jsc) and fill factor (FF) remained unchanged.

  17. High-Level Expression of Recombinant Bovine Lactoferrin in Pichia pastoris with Antimicrobial Activity

    Science.gov (United States)

    Iglesias-Figueroa, Blanca; Valdiviezo-Godina, Norberto; Siqueiros-Cendón, Tania; Sinagawa-García, Sugey; Arévalo-Gallegos, Sigifredo; Rascón-Cruz, Quintín

    2016-01-01

    In this study, bovine lactoferrin (bLf), an iron-binding glycoprotein considered an important nutraceutical protein because of its several properties, was expressed in Pichia pastoris KM71-H under AOX1 promoter control, using pJ902 as the recombinant plasmid. Dot blotting analysis revealed the expression of recombinant bovine lactoferrin (rbLf) in Pichia pastoris. After Bach fermentation and purification by molecular exclusion, we obtained an expression yield of 3.5 g/L of rbLf. rbLf and predominantly pepsin-digested rbLf (rbLfcin) demonstrated antibacterial activity against Escherichia coli (E. coli) BL21DE3, Staphylococcus aureus (S. aureus) FRI137, and, in a smaller percentage, Pseudomonas aeruginosa (Ps. Aeruginosa) ATCC 27833. The successful expression and characterization of functional rbLf expressed in Pichia pastoris opens a prospect for the development of natural antimicrobial agents produced recombinantly. PMID:27294912

  18. High-Level Expression of Recombinant Bovine Lactoferrin in Pichia pastoris with Antimicrobial Activity

    Directory of Open Access Journals (Sweden)

    Blanca Iglesias-Figueroa

    2016-06-01

    Full Text Available In this study, bovine lactoferrin (bLf, an iron-binding glycoprotein considered an important nutraceutical protein because of its several properties, was expressed in Pichia pastoris KM71-H under AOX1 promoter control, using pJ902 as the recombinant plasmid. Dot blotting analysis revealed the expression of recombinant bovine lactoferrin (rbLf in Pichia pastoris. After Bach fermentation and purification by molecular exclusion, we obtained an expression yield of 3.5 g/L of rbLf. rbLf and predominantly pepsin-digested rbLf (rbLfcin demonstrated antibacterial activity against Escherichia coli (E. coli BL21DE3, Staphylococcus aureus (S. aureus FRI137, and, in a smaller percentage, Pseudomonas aeruginosa (Ps. Aeruginosa ATCC 27833. The successful expression and characterization of functional rbLf expressed in Pichia pastoris opens a prospect for the development of natural antimicrobial agents produced recombinantly.

  19. BIOLOGICALLY ACTIVE SUBSTANCES OF SPIRIT PRODUCTION WASTE

    Directory of Open Access Journals (Sweden)

    A. S. Kayshev

    2014-01-01

    Full Text Available A content of biologically active compounds (BAC with signified pharmacological activity in distillers grains was proved. It is prospective for applications of these grains as a raw material resource of pharmaceuticals. A composition of BAC distillers grains received from wheat, corn, barley, millet at different spirit enterprises which use hydro fermentative grain processing. Considering polydispersity of distillers grains they were separated on solid and liquid phases preliminary. Physical and chemical characteristics of distillers grains' liquid base were identified. Elementary composition of distillers grains is signified by active accumulation of biogenic elements (phosphorus, potassium, magnesium, calcium, sodium, iron and low content of heavy metals. The solid phase of distillers grains accumulates carbon, hydrogen and nitrogen in high concentration. The liquid phase of distillers grains contains: proteins and amino acids (20-46%, reducing sugars (5,6%-17,5%, galacturonides (0,8-1,4%, ascorbic acid (6,2-11,4 mg%. The solid base of distillers grains contains: galacturonides (3,4-5,3%, fatty oil (8,4-11,1% with predomination of essential fatty acids, proteins and amino acids (2,1-2,5%, flavonoids (0,4-0,9%, tocopherols (3,4-7,7 mg%. A method of complex processing of distillers grains based on application of membrane filtering of liquid phase and liquid extraction by inorganic and organic solvents of solid phase, which allows almost full extraction of the sum of biologically active compounds (BAC from liquid phase (Biobardin BM and solid phase (Biobardin UL. Biobardin BM comprises the following elements: proteins and amino acids (41-69%, reducing sugars (3,5-15,6%, fatty oil (0,2-0,3%, flavonoids (0,2-0,7%, ascorbic acid (17-37 mg%. Biobardin UL includes: oligouronids (16,4-19,5%, proteins and amino acids (11-21%, fatty oil (3,2-4,9% which includes essential acids; flavonoids (0,6-1,5%, tocopherols (6,6-10,2 mg%, carotinoids (0,13-0,21 mg

  20. Monascus secondary metabolites: production and biological activity.

    Science.gov (United States)

    Patakova, Petra

    2013-02-01

    The genus Monascus, comprising nine species, can reproduce either vegetatively with filaments and conidia or sexually by the formation of ascospores. The most well-known species of genus Monascus, namely, M. purpureus, M. ruber and M. pilosus, are often used for rice fermentation to produce red yeast rice, a special product used either for food coloring or as a food supplement with positive effects on human health. The colored appearance (red, orange or yellow) of Monascus-fermented substrates is produced by a mixture of oligoketide pigments that are synthesized by a combination of polyketide and fatty acid synthases. The major pigments consist of pairs of yellow (ankaflavin and monascin), orange (rubropunctatin and monascorubrin) and red (rubropunctamine and monascorubramine) compounds; however, more than 20 other colored products have recently been isolated from fermented rice or culture media. In addition to pigments, a group of monacolin substances and the mycotoxin citrinin can be produced by Monascus. Various non-specific biological activities (antimicrobial, antitumor, immunomodulative and others) of these pigmented compounds are, at least partly, ascribed to their reaction with amino group-containing compounds, i.e. amino acids, proteins or nucleic acids. Monacolins, in the form of β-hydroxy acids, inhibit hydroxymethylglutaryl-coenzyme A reductase, a key enzyme in cholesterol biosynthesis in animals and humans.

  1. Biological activity of Serratia marcescens cytotoxin

    Directory of Open Access Journals (Sweden)

    G.V. Carbonell

    2003-03-01

    Full Text Available Serratia marcescens cytotoxin was purified to homogeneity by ion-exchange chromatography on a DEAE Sepharose Fast Flow column, followed by gel filtration chromatography on a Sephadex G100 column. The molecular mass of the cytotoxin was estimated to be about 50 kDa. Some biological properties of the cytotoxin were analyzed and compared with well-characterized toxins, such as VT1, VT2 and CNF from Escherichia coli and hemolysin produced by S. marcescens. The sensitivity of the cell lines CHO, HeLa, HEp-2, Vero, BHK-21, MA 104 and J774 to the cytotoxin was determined by the cell viability assay using neutral red. CHO and HEp-2 were highly sensitive, with massive cellular death after 1 h of treatment, followed by BHK-21, HeLa, Vero and J774 cells, while MA 104 was insensitive to the toxin. Cytotoxin induced morphological changes such as cell rounding with cytoplasmic retraction and nuclear compactation which were evident 15 min after the addition of cytotoxin. The cytotoxic assays show that 15 min of treatment with the cytotoxin induced irreversible intoxication of the cells, determined by loss of cell viability. Concentrations of 2 CD50 (0.56 µg/ml of purified cytotoxin did not present any hemolytic activity, showing that the cytotoxin is distinct from S. marcescens hemolysin. Antisera prepared against S. marcescens cytotoxin did not neutralize the cytotoxic activity of VT1, VT2 or CNF toxin, indicating that these toxins do not share antigenic determinants with cytotoxin. Moreover, we did not detect gene sequences for any of these toxins in S. marcescens by PCR assay. These results suggest that S. marcescens cytotoxin is not related to any of these toxins from E. coli.

  2. Biological activity of Serratia marcescens cytotoxin.

    Science.gov (United States)

    Carbonell, G V; Amorim, C R N; Furumura, M T; Darini, A L C; Fonseca, B A L; Yano, T

    2003-03-01

    Serratia marcescens cytotoxin was purified to homogeneity by ion-exchange chromatography on a DEAE Sepharose Fast Flow column, followed by gel filtration chromatography on a Sephadex G100 column. The molecular mass of the cytotoxin was estimated to be about 50 kDa. Some biological properties of the cytotoxin were analyzed and compared with well-characterized toxins, such as VT1, VT2 and CNF from Escherichia coli and hemolysin produced by S. marcescens. The sensitivity of the cell lines CHO, HeLa, HEp-2, Vero, BHK-21, MA 104 and J774 to the cytotoxin was determined by the cell viability assay using neutral red. CHO and HEp-2 were highly sensitive, with massive cellular death after 1 h of treatment, followed by BHK-21, HeLa, Vero and J774 cells, while MA 104 was insensitive to the toxin. Cytotoxin induced morphological changes such as cell rounding with cytoplasmic retraction and nuclear compactation which were evident 15 min after the addition of cytotoxin. The cytotoxic assays show that 15 min of treatment with the cytotoxin induced irreversible intoxication of the cells, determined by loss of cell viability. Concentrations of 2 CD50 (0.56 g/ml) of purified cytotoxin did not present any hemolytic activity, showing that the cytotoxin is distinct from S. marcescens hemolysin. Antisera prepared against S. marcescens cytotoxin did not neutralize the cytotoxic activity of VT1, VT2 or CNF toxin, indicating that these toxins do not share antigenic determinants with cytotoxin. Moreover, we did not detect gene sequences for any of these toxins in S. marcescens by PCR assay. These results suggest that S. marcescens cytotoxin is not related to any of these toxins from E. coli.

  3. Recombinant interferon alfa-2a, an active agent in advanced cutaneous T-cell lymphomas.

    Science.gov (United States)

    Bunn, P A; Ihde, D C; Foon, K A

    1987-01-01

    The cutaneous T-cell lymphomas including mycosis fungoides and the Sézary syndrome, are indolent lymphomas with early systemic dissemination. Like the indolent B-cell lymphomas, they cannot be cured by currently available systemic chemotherapy so new systemic therapies need to be developed. A study of very high-dose recombinant interferon alfa-2a was, therefore, initiated in 20 patients with advanced cutaneous T-cell lymphoma (5 in stage II, 2 in stage III and 13 in stage IV). All patients were refractory to at least 2 standard therapies, including topical nitrogen mustard (18 patients), psoralens and ultraviolet A light (12 patients), total skin electron irradiation (14 patients) and systemic chemotherapy (16 patients). Nine out of 20 patients (45%; 95% confidence interval 25-69%) had either objective partial or complete responses within 3 months of starting treatment. Maximal response, however, often did not occur for at least one year. The median duration of response was 5.5 months and all complete responses lasted more than 2 years. Response frequencies were equal at both cutaneous and extracutaneous sites and in patients with or without prior chemotherapy. Toxicity was exhibited primarily as a flu-like syndrome consisting of fever, malaise, fatigue, anorexia and weight loss which necessitated dose reductions in all patients. Transient elevations in liver function and decreases in renal function and granulocyte counts occurred in some patients. It is concluded that interferon alfa-2a is highly active against advanced cutaneous T-cell lymphomas and that it should be studied in its early stages. It should also be evaluated in combination with other biological agents and with chemotherapy.

  4. Low intensity ultrasound stimulates biological activity of aerobic activated sludge

    Institute of Scientific and Technical Information of China (English)

    LIU Hong; YAN Yixin; WANG Wenyan; YU Yongyong

    2007-01-01

    This work aims to explore a procedure to improve biological wastewater treatment efficiency using low intensity ultrasound.The aerobic activated sludge from a municipal wastewater treatment plant was used as the experimental material.Oxygen uptake rate(OUR)of the activated sludge (AS)was determined to indicate the changes of AS activity stimulated by ultrasound at 35 kHZ for 0-40 min with ultrasonic intensities of 0-1.2 W/cm2.The highest OUR was observed at the ultrasonic intensity of 0.3 W/cm2 and an irradiation period of 10 min;more than 15% increase was achieved immediately after sonication.More significantly,the AS activity stimulated by ultrasound could last 24 h after sonication,and the AS activity achieved its peak value within 8 h after sonication.or nearly 100% higher than the initial level after sonication.Therefore,to improve the wastewater treatment efficiency of bioreactors,ultrasound with an intensity of 0.3 W/cm2 could be employed to irradiate a part of the AS in the bioreactor for 10 min every 8 h.

  5. An Acidic Thermostable Recombinant Aspergillus nidulans Endoglucanase Is Active towards Distinct Agriculture Residues

    Directory of Open Access Journals (Sweden)

    Eveline Queiroz de Pinho Tavares

    2013-01-01

    Full Text Available Aspergillus nidulans is poorly exploited as a source of enzymes for lignocellulosic residues degradation for biotechnological purposes. This work describes the A. nidulans Endoglucanase A heterologous expression in Pichia pastoris, the purification and biochemical characterization of the recombinant enzyme. Active recombinant endoglucanase A (rEG A was efficiently secreted as a 35 kDa protein which was purified through a two-step chromatography procedure. The highest enzyme activity was detected at 50°C/pH 4. rEG A retained 100% of activity when incubated at 45 and 55°C for 72 h. Purified rEG A kinetic parameters towards CMC were determined as Km=27.5±4.33 mg/mL, Vmax=1.185±0.11 mmol/min, and 55.8 IU (international units/mg specific activity. Recombinant P. pastoris supernatant presented hydrolytic activity towards lignocellulosic residues such as banana stalk, sugarcane bagasse, soybean residues, and corn straw. These data indicate that rEG A is suitable for plant biomass conversion into products of commercial importance, such as second-generation fuel ethanol.

  6. The relationship between the connecting peptide of recombined single chain insulin and its biological function

    Institute of Scientific and Technical Information of China (English)

    HUANG; Yiding; (

    2001-01-01

    [1]Straus, D. S., Growth-stimulatory of insulin in vitro and in vivo, Endocr. Rev., 1984, 5(2): 356-369.[2]Svenningsen, A. F., Kanje, M., Insulin and the insulin-like growth factors I and II are mitogenic to cultured rat sciatic nerve segments and stimulate [3H] thuymidine incorporation through their respective receptors, Glia, 1996, 18(1): 68-72.[3]Ogihara, S., Yamada, M., Saito, T. et al., Insulin potentiates mitogenic effect of epidermal growth factor on cultured guinea pig gastric mucous cells, Am. J. Physiol., 1996, 271(1 Pt 1): G104-121.[4]Steiner, D. F., Oyer, P. E., The biosynthesis of insulin and a probable precursor of insulin by a human islet cell adenoma, Proc. Nalt. Acad. Sci. USA, 1967, 57(2): 473-480.[5]King, G. L., Kahn, C. R., The growth-promoting effects of insulin, in Growth and Maturation Factors(ed. Guroff, G.), New York: John Wiley & Sons, 1984, 223-265.[6]Peavy, D. E., Brunner, M. R., Duckworth, W. C. et al., Receptor binding and biological potency of several split forms (conversion intermediates) of human proinsulin, Studies in cultured IM-9 lymphocytes and in vivo and in vitro in rats, J. Biol. Chem., 1985, 260: 13989-13994.[7]Derewenda, U., Derewenda, Z., Dodson, E. J. et al., X-ray analysis of the single chain B29-A1 peptide-linked insulin molecule. A completely inactive analogue, J. Mol. Biol., 1991, 220: 425-433.[8]Hua, Q. X., Shoelson, S. E., Kochoyan, M. et al., Receptor binding redefined by a structural switch in a mutant human insulin, Nature, 1991, 354: 238-241.[9]Hua, Q. X., Gozani, S. N., Chance, R. E. et al., Structure of a protein in a kinetic trap, Nat. Struc. Boil, 1995, 2: 129-138.[10]Kristensen, C., Andersen, A. S., Hach, M., A single-chain insulin-like growth factor I/insulin hybrid binds with high affinity to the insulin receptor, Biochem. J., 1995, 305: 981-986.[11]Humbel, R. E., Insulin-like growth factors I and II, Euro. J. Biochem., 1990, 190: 445-462.[12]Cooke, R. M

  7. RECOMBINANT LENTIVIRUS-MEDIATED SILENCING OF ADAPTOR PROTEIN RUK/CIN85 EXPRESSION INFLUENCES BIOLOGICAL RESPONSES OF TUMOR CELLS

    Directory of Open Access Journals (Sweden)

    A. A. Samoylenko

    2013-08-01

    Full Text Available Ruk/CIN85 is an adaptor protein that plays important roles in the regulation of cellular processes such as cell death, proliferation and motility. It was recently shown that overexpression of Ruk/CIN85 increases the oncogenic potential of human breast adenocarcinoma MCF-7 cells. It was the aim of the present study to investigate whether inhibition of Ruk/CIN85 expression has an effect on the biological properties of the cells. In order to down-regulate Ruk/CIN85 expression of small interfering RNA-based approach was used. For down-regulation of Ruk/CIN85 lentiviral constructs encoding Ruk/CIN85-specific small hairpin RNA sequences were generated. By using the obtained recombinant lentiviruses it was shown that inhibition of Ruk/CIN85 expression influences biological properties (motility, proliferation, ABCG2 expression, and ROS generation of various tumour cell types such as human breast adenocarcinoma MCF-7, human colorectal adenocarcinoma HT-29, and Lewis mouse lung carcinoma cells.

  8. Secretion of biologically active interferon-gamma inducible protein-10 (IP-10 by Lactococcus lactis

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    Saucedo-Cardenas Odila

    2008-07-01

    Full Text Available Abstract Background Chemokines are a large group of chemotactic cytokines that regulate and direct migration of leukocytes, activate inflammatory responses, and are involved in many other functions including regulation of tumor development. Interferon-gamma inducible-protein-10 (IP-10 is a member of the C-X-C subfamily of the chemokine family of cytokines. IP-10 specifically chemoattracts activated T lymphocytes, monocytes, and NK cells. IP-10 has been described also as a modulator of other antitumor cytokines. These properties make IP-10 a novel therapeutic molecule for the treatment of chronic and infectious diseases. Currently there are no suitable live biological systems to produce and secrete IP-10. Lactococcus lactis has been well-characterized over the years as a safe microorganism to produce heterologous proteins and to be used as a safe, live vaccine to deliver antigens and cytokines of interest. Here we report a recombinant strain of L. lactis genetically modified to produce and secrete biologically active IP-10. Results The IP-10 coding region was isolated from human cDNA and cloned into an L. lactis expression plasmid under the regulation of the pNis promoter. By fusion to the usp45 secretion signal, IP-10 was addressed out of the cell. Western blot analysis demonstrated that recombinant strains of L. lactis secrete IP-10 into the culture medium. Neither degradation nor incomplete forms of IP-10 were detected in the cell or supernatant fractions of L. lactis. In addition, we demonstrated that the NICE (nisin-controlled gene expression system was able to express IP-10 "de novo" even two hours after nisin removal. This human IP-10 protein secreted by L. lactis was biological active as demonstrated by Chemotaxis assay over human CD3+T lymphocytes. Conclusion Expression and secretion of mature IP-10 was efficiently achieved by L. lactis forming an effective system to produce IP-10. This recombinant IP-10 is biologically active as

  9. The fed-batch principle for the molecular biology lab: controlled nutrient diets in ready-made media improve production of recombinant proteins in Escherichia coli.

    Science.gov (United States)

    Krause, Mirja; Neubauer, Antje; Neubauer, Peter

    2016-06-17

    While the nutrient limited fed-batch technology is the standard of the cultivation of microorganisms and production of heterologous proteins in industry, despite its advantages in view of metabolic control and high cell density growth, shaken batch cultures are still the standard for protein production and expression screening in molecular biology and biochemistry laboratories. This is due to the difficulty and expenses to apply a controlled continuous glucose feed to shaken cultures. New ready-made growth media, e.g. by biocatalytic release of glucose from a polymer, offer a simple solution for the application of the fed-batch principle in shaken plate and flask cultures. Their wider use has shown that the controlled diet not only provides a solution to obtain significantly higher cell yields, but also in many cases folding of the target protein is improved by the applied lower growth rates; i.e. final volumetric yields for the active protein can be a multiple of what is obtained in complex medium cultures. The combination of the conventional optimization approaches with new and easy applicable growth systems has revolutionized recombinant protein production in Escherichia coli in view of product yield, culture robustness as well as significantly increased cell densities. This technical development establishes the basis for successful miniaturization and parallelization which is now an important tool for synthetic biology and protein engineering approaches. This review provides an overview of the recent developments, results and applications of advanced growth systems which use a controlled glucose release as substrate supply.

  10. Recombination-activating gene 1 and 2 (RAG1 and RAG2) in flounder (Paralichthys olivaceus)

    Indian Academy of Sciences (India)

    Xianlei Wang; Xungang Tan; Pei-Jun Zhang; Yuqing Zhang; Peng Xu

    2014-12-01

    During the development of B and T lymphocytes, Ig and TCR variable region genes are assembled from germline V, D, and J gene segments by a site-specific recombination reaction known as V(D)J recombination. The process of somatic V(D)J recombination, mediated by the recombination-activating gene (RAG) products, is the most significant characteristic of adaptive immunity in jawed vertebrates. Flounder (Paralichthys olivaceus) RAG1 and RAG2 were isolated by Genome Walker and RT-PCR, and their expression patterns were analysed by RT-PCR and in situ hybridization on sections. RAG1 spans over 7.0 kb, containing 4 exons and 3 introns, and the full-length ORF is 3207 bp, encoding a peptide of 1068 amino acids. The first exon lies in the 5′-UTR, which is an alternative exon. RAG2 full-length ORF is 1062 bp, encodes a peptide of 533 amino acids, and lacks introns in the coding region. In 6-month-old flounders, the expression of RAG1 and RAG2 was essentially restricted to the pronephros (head kidney) and mesonephros (truck kidney). Additionally, both of them were mainly expressed in the thymus. These results revealed that the thymus and kidney most likely serve as the primary lymphoid tissues in the flounder.

  11. Insecticidal activity of two proteases against Spodoptera frugiperda larvae infected with recombinant baculoviruses

    Directory of Open Access Journals (Sweden)

    Nagata Tatsuya

    2010-06-01

    Full Text Available Abstract Background Baculovirus comprise the largest group of insect viruses most studied worldwide, mainly because they efficiently kill agricutural insect pests. In this study, two recombinant baculoviruses containing the ScathL gene from Sarcophaga peregrina (vSynScathL, and the Keratinase gene from the fungus Aspergillus fumigatus (vSynKerat, were constructed. and their insecticidal properties analysed against Spodoptera frugiperda larvae. Results Bioassays of third-instar and neonate S. frugiperda larvae with vSynScathL and vSynKerat showed a decrease in the time needed to kill the infected insects when compared to the wild type virus. We have also shown that both recombinants were able to increase phenoloxidase activity in the hemolymph of S. frugiperda larvae. The expression of proteases in infected larvae resulted in destruction of internal tissues late in infection, which could be the reason for the increased viral speed of kill. Conclusions Baculoviruses and their recombinant forms constitute viable alternatives to chemical insecticides. Recombinant baculoviruses containing protease genes can be added to the list of engineered baculoviruses with great potential to be used in integrated pest management programs.

  12. Human IgE is efficiently produced in glycosylated and biologically active form in lepidopteran cells.

    Science.gov (United States)

    Bantleon, Frank; Wolf, Sara; Seismann, Henning; Dam, Svend; Lorentzen, Andrea; Miehe, Michaela; Jabs, Frederic; Jakob, Thilo; Plum, Melanie; Spillner, Edzard

    2016-04-01

    TH2-biased immunity to parasites and allergens is often associated with increased levels of antigen-specific and high affinity IgE. The role in reacting against minute amounts of target structures and to provoke severe anaphylactic reactions renders IgE a mechanistically outstanding isotype. IgE represents the least abundant serum antibody isotype and exhibits a variety of peculiarities including structure, extensive glycosylation and effector functions. Despite large progress in antibody technologies, however, the recombinant access to isotypes beyond IgG such as IgE still is scarce. The capacity of expression systems has to meet the complex structural conformations and the extensive posttranslational modifications that are indispensable for biological activity. In order to provide alternatives to mammalian expression systems with often low yield and a more complex glycosylation pattern we established the recombinant production of the highly complex IgE isotype in insect cells. Recombinant IgE (rIgE) was efficiently assembled and secreted into the supernatant in yields of >30 mg/L. Purification from serum free medium using different downstream processing methods provided large amounts of rIgE. This exhibited a highly specific interaction with its antigen, therapeutic anti-IgE and its high affinity receptor, the FcεRI. Lectins and glyco-proteomic analyses proved the presence of prototypic insect type N-glycans on the epsilon heavy chain. Mediator release assays demonstrated a biological activity of the rIgE comparable to IgE derived from mammalian cells. In summary the expression in insect cells provides rIgE with variant glycosylation pattern, but retained characteristics and biological activity. Therefore our data contribute to the understanding of functional and structural aspects and potential use of the IgE isotype.

  13. The ice nucleation activity of biological aerosols

    Science.gov (United States)

    Grothe, H.; Pummer, B.; Bauer, H.; Bernardi, J.

    2012-04-01

    Primary Biological Aerosol Particles (PBAPs), including bacteria, spores and pollen may be important for several atmospheric processes. Particularly, the ice nucleation caused by PBAPs is a topic of growing interest, since their impact on ice cloud formation and thus on radiative forcing, an important parameter in global climate is not yet fully understood. In laboratory model studies we investigated the ice nucleation activity of selected PBAPs. We studied the immersion mode freezing using water-oil emulsion, which we observed by optical microscopy. We particularly focused on pollen. We show that pollen of different species strongly differ in their ice nucleation behavior. The average freezing temperatures in laboratory experiments range from 240 K to 255 K. As the most efficient nuclei (silver birch, Scots pine and common juniper pollen) have a distribution area up to the Northern timberline, their ice nucleation activity might be a cryoprotective mechanism. For comparison the ice nucleation activity of Snomax, fungal spores, and mushrooms will be discussed as well. In the past, pollen have been rejected as important atmospheric IN, as they are not as abundant in the atmosphere as bacteria or mineral dust and are too heavy to reach higher altitudes. However, in our experiments (Pummer et al. 2011) it turned out that water, which had been in contact with pollen and then been separated from the bodies, nucleates as good as the pollen grains themselves. So the ice nuclei have to be easily-suspendable macromolecules (100-300 kDa) located on the pollen. Once extracted, they can be distributed further through the atmosphere than the heavy pollen grains and so augment the impact of pollen on ice cloud formation even in the upper troposphere. It is widely known, that material from the pollen, like allergens and sugars, can indeed leave the pollen body and be distributed independently. The most probable mechanism is the pollen grain bursting by rain, which releases

  14. Multifunctional and biologically active matrices from multicomponent polymeric solutions

    Science.gov (United States)

    Kiick, Kristi L. (Inventor); Yamaguchi, Nori (Inventor); Rabolt, John (Inventor); Casper, Cheryl (Inventor)

    2012-01-01

    A functionalized electrospun matrix for the controlled-release of biologically active agents, such as growth factors, is presented. The functionalized matrix comprises a matrix polymer, a compatibilizing polymer and a biomolecule or other small functioning molecule. In certain aspects the electrospun polymer fibers comprise at least one biologically active molecule functionalized with low molecular weight heparin.

  15. MOLECULAR CLONING, SEQUENCING, EXPRESSION AND BIOLOGICAL ACTIVITY OF GIANT PANDA (AILUROPODA MELANOLEUCA) INTERFERON-GAMMA.

    Science.gov (United States)

    Zhu, Hui; Wang, Wen-Xiu; Wang, Bao-Qin; Zhu, Xiao-Fu; Wu, Xu-Jin; Ma, Qing-Yi; Chen, De-Kun

    2012-06-29

    The giant panda (Ailuropoda melanoleuca) is an endangered species and indigenous to China. Interferon-gamma (IFN-γ) is the only member of type □ IFN and is vital for the regulation of host adapted immunity and inflammatory response. Little is known aboutthe FN-γ gene and its roles in giant panda.In this study, IFN-γ gene of Qinling giant panda was amplified from total blood RNA by RT-CPR, cloned, sequenced and analysed. The open reading frame (ORF) of Qinling giant panda IFN-γ encodes 152 amino acidsand is highly similar to Sichuan giant panda with an identity of 99.3% in cDNA sequence. The IFN-γ cDNA sequence was ligated to the pET32a vector and transformed into E. coli BL21 competent cells. Expression of recombinant IFN-γ protein of Qinling giant panda in E. coli was confirmed by SDS-PAGE and Western blot analysis. Biological activity assay indicated that the recombinant IFN-γ protein at the concentration of 4-10 µg/ml activated the giant panda peripheral blood lymphocytes,while at 12 µg/mlinhibited. the activation of the lymphocytes.These findings provide insights into the evolution of giant panda IFN-γ and information regarding amino acid residues essential for their biological activity.

  16. Effect of recombinant erythropoietin on functional activity of cultured human cells.

    Science.gov (United States)

    Emel'yanova, E A; Kosykh, A V; Sukhanov, Yu V; Vorotelyak, E A; Vasil'ev, A V

    2012-08-01

    We studied the effect of recombinant human erythropoietin on functional activity of skin cells in vitro. It was found that erythropoietin stimulated proliferation of mesenchymal and epithelial cells and effectively protected epidermal HaCaT cells from apoptosis. Insignificant effect of erythropoietin on contraction of collagen gel by mesenchymal cells was revealed. These findings suggest that erythropoietin can be a promising component of wound-healing preparations.

  17. Targeting Platinum Compounds: synthesis and biological activity

    OpenAIRE

    VAN ZUTPHEN, Steven

    2005-01-01

    Inspired by cisplatin, the inorganic drug discovered by Barnett Rosenberg in 1965, the research described in this thesis uses targeting ligands, or ligands varied in a combinatorial fashion, to find platinum complexes with more specific modes of action. These studies have lead to the development of novel (solid-phase) synthetic methods and to the discovery of several compounds with promising biological properties.

  18. Milk inhibits the biological activity of ricin

    Science.gov (United States)

    Ricin is a highly toxic protein produced by the castor plant Ricinus communis. The toxin is relatively easy to isolate and can be used as a biological weapon. There is great interest in identifying effective inhibitors for ricin. In this study, we demonstrated by three independent assays that compon...

  19. Immunoadjuvant activities of a recombinant chicken IL-12 in chickens vaccinated with Newcastle disease virus recombinant HN protein.

    Science.gov (United States)

    Su, Bor Sheu; Yin, Hsien Sheng; Chiu, Hua Hsien; Hung, Li Hsiang; Huang, Ji Ping; Shien, Jui Hung; Lee, Long Huw

    2011-08-05

    Recombinant fowlpox virus (rFPV/HN) expressing Newcastle disease virus (NDV) HN gene and rFPV/HN/chIL-12 co-expressing chicken IL-12 (chIL-12) and HN (rHN/chIL-12) genes have been characterized. rHN/chIL-12 or rchIL-12, expressed by our previous construct rFPV/chIL-12, co-administered with rHN was assessed for adjuvant activities of chIL-12. Chickens were vaccinated with various amounts of rHN/chIL-12 mixed with mineral oil (MO), intramuscularly. Levels of hemagglutination-inhibition (HI) antibody production depended on the concentration of the injected rHN or rHN/chIL-12. The lower HI antibody titers were obtained in chicken groups rHN/chIL-12/7-rHN/chIL-12/9, receiving 60ng rHN/8ng chIL-12 with MO, 30ng rHN/4ng chIL-12 with MO or 15ng rHN/2ng chIL-12 with MO, respectively, compared to those in chicken groups rHN/7-rHN/9, receiving rHN with MO alone. However, chickens in group rHN/chIL-12/7 or rHN/chIL-12/8 and rHN with MO alone showed the same effective protection. Chicken group rHN/chIL-12/9 was even more protective than that in group rHN/9. When rchIL-12 was co-injected with 15ng rHN plus MO, chickens produced low levels of HI antibody titers; while higher levels of IFN-γ production and an effective protection rate (83%) were obtained. On the other hand, low levels of IFN-γ production and low protection response (50%) were obtained in chickens injected with rHN with MO alone. Taken together, when the concentration of rHN decreased to certain levels, rchIL-12 reduced HI antibody production. The increase in the induction of IFN-γ production might suggest the enhancement of the cell-mediated immunity which conferred the protection from the NDV challenge.

  20. Recombinant production of enzymatically active male contraceptive drug target hTSSK2 - Localization of the TSKS domain phosphorylated by TSSK2.

    Science.gov (United States)

    Shetty, Jagathpala; Sinville, Rondedrick; Shumilin, Igor A; Minor, Wladek; Zhang, Jianhai; Hawkinson, Jon E; Georg, Gunda I; Flickinger, Charles J; Herr, John C

    2016-05-01

    The testis-specific serine/threonine kinase 2 (TSSK2) has been proposed as a candidate male contraceptive target. Development of a selective inhibitor for this kinase first necessitates the production of highly purified, soluble human TSSK2 and its substrate, TSKS, with high yields and retention of biological activity for crystallography and compound screening. Strategies to produce full-length, soluble, biologically active hTSSK2 in baculovirus expression systems were tested and refined. Soluble preparations of TSSK2 were purified by immobilized-metal affinity chromatography (IMAC) followed by gel filtration chromatography. The biological activities of rec.hTSSK2 were verified by in vitro kinase and mobility shift assays using bacterially produced hTSKS (isoform 2), casein, glycogen synthase peptide (GS peptide) and various TSKS peptides as target substrates. Purified recombinant hTSSK2 showed robust kinase activity in the in vitro kinase assay by phosphorylating hTSKS isoform 2 and casein. The ATP Km values were similar for highly and partially purified fractions of hTSSK2 (2.2 and 2.7 μM, respectively). The broad spectrum kinase inhibitor staurosporine was a potent inhibitor of rec.hTSSK2 (IC50 = 20 nM). In vitro phosphorylation experiments carried out with TSKS (isoform 1) fragments revealed particularly strong phosphorylation of a recombinant N-terminal region representing aa 1-150 of TSKS, indicating that the N-terminus of human TSKS is phosphorylated by human TSSK2. Production of full-length enzymatically active recombinant TSSK2 kinase represents the achievement of a key benchmark for future discovery of TSSK inhibitors as male contraceptive agents.

  1. How-To-Do-It: Recombinant DNA Technology in the High School Biology Laboratory.

    Science.gov (United States)

    Myers, Richard

    1988-01-01

    Describes a basic biotechnology investigation that includes restriction and ligation of plasmid DNA, transformation of bacteria and cloning of these bacterial cells. Discusses laboratory procedures and another activity in the identification of unknown plasmids by studying agarose gel electrophoresis photographs. (CW)

  2. Evolution of class switch recombination function in fish activation-induced cytidine deaminase, AID.

    Science.gov (United States)

    Wakae, Koshou; Magor, Brad G; Saunders, Holly; Nagaoka, Hitoshi; Kawamura, Akemi; Kinoshita, Kazuo; Honjo, Tasuku; Muramatsu, Masamichi

    2006-01-01

    Following activation of mammalian B cells, class switch recombination (CSR) and somatic hypermutation (SHM) of the Ig heavy chain (IgH) gene can improve the functions of the expressed antibodies. Activation-induced cytidine deaminase (AID) is the only known B cell-specific protein required for inducing CSR and SHM in mammals. Lower vertebrates have an AID homologue, and there is some evidence of SHM in vivo. However there is no evidence of CSR in the cartilaginous or bony fishes, and this may be due in part to a lack of cis-elements in the IgH gene that are the normal targets of AID-mediated recombination. We have tested whether bony fish (zebrafish and catfish) AID can mediate CSR and SHM in mammalian cells. As expected, ectopic expression of fish AID in mouse fibroblasts resulted in mutations in an introduced SHM reporter gene, indicating that fish AID can mediate SHM. Unexpectedly, expression of fish AID in mouse AID-/- B cells induced surface IgG expression as well as switched transcripts from Ig gene loci, clearly indicating that the fish AID protein can mediate CSR, at least in mouse cells. These results suggest that the AID protein acquired the ability to mediate CSR before the IgH locus evolved the additional exon clusters and switch regions that are the targets of recombination. We discuss how pleiotropic functions of specific domains within the AID protein may have facilitated the early evolution of CSR in lower vertebrates.

  3. [Expression of angiogenin in COS-7 cells and analysis of its biological activity].

    Science.gov (United States)

    Wang, Yuan-Yuan; Zou, Min-Jig; Cai, Xin; Liu, Shen; Wang, Jin-Feng; Xu, Tao; Wang, Jia-Xi; Su, Hang; Xu, Dong-Gan

    2008-06-01

    This study was purposed to investigate the angiogenin (ANG) expression in COS-7 cells and its biological activity. The gene of angiogenin was obtained from mononuclear cells of peripheral blood by using RT-PCR and inserted into eukaryotic expression vector of pcDNA3.1. After being transfected into COS-7 cells, the recombinant ANG was identified by Western blot assay. The function of promoting proliferation of ANG to ECV304 cells was detected by MTT method, and its activity of vascularization was analyzed by chick embryo chorioallantois treated by the culture supernatant after transfection with pcDNA3.1-ang. The result showed that recombinant ANG was expressed in COS-7 cells after transfection for 24 to 36 hours. It could specifically react with monoclonal antibody against ANG. The recombinant ANG could obviously promote the proliferation of ECV304 cells. In contrast with the control group, the culture supernatant of pcDNA3.1-ang transfected group could stimulate the angiogenesis in embryo chorioallantois. It is concluded that the ang transiently expresses in COS-7 cells, and its expression product obviously stimulates the cell proliferation and angiogenesis.

  4. Recombinant Collagenlike Proteins

    Science.gov (United States)

    Fertala, Andzej

    2007-01-01

    A group of collagenlike recombinant proteins containing high densities of biologically active sites has been invented. The method used to express these proteins is similar to a method of expressing recombinant procollagens and collagens described in U. S. Patent 5,593,859, "Synthesis of human procollagens and collagens in recombinant DNA systems." Customized collagenous proteins are needed for biomedical applications. In particular, fibrillar collagens are attractive for production of matrices needed for tissue engineering and drug delivery. Prior to this invention, there was no way of producing customized collagenous proteins for these and other applications. Heretofore, collagenous proteins have been produced by use of such biological systems as yeasts, bacteria, and transgenic animals and plants. These products are normal collagens that can also be extracted from such sources as tendons, bones, and hides. These products cannot be made to consist only of biologically active, specific amino acid sequences that may be needed for specific applications. Prior to this invention, it had been established that fibrillar collagens consist of domains that are responsible for such processes as interaction with cells, binding of growth factors, and interaction with a number of structural proteins present in the extracellular matrix. A normal collagen consists of a sequence of domains that can be represented by a corresponding sequence of labels, e.g., D1D2D3D4. A collagenlike protein of the present invention contains regions of collagen II that contain multiples of a single domain (e.g., D1D1D1D1 or D4D4D4D4) chosen for its specific biological activity. By virtue of the multiplicity of the chosen domain, the density of sites having that specific biological activity is greater than it is in a normal collagen. A collagenlike protein according to this invention can thus be made to have properties that are necessary for tissue engineering.

  5. Specific activities of poetam preparation (superlow-doses of antibodies to erythropoietin) and recombinant erythropoietin.

    Science.gov (United States)

    Dygai, A M; Zhdanov, V V; Udut, E V; Simanina, E V; Gur'yantseva, L A; Khrichkova, T Yu; Epshtein, O I; Sergeeva, S A

    2006-09-01

    We compared the capacity of superlow-dose of antibodies to erythropoietin (Poetam) and recombinant erythropoietin (Recormon) to stimulate the recovery of adriamycin-suppressed erythropoiesis in mice. Both preparations exhibited high erythron activation capacity and considerably increased the content of erythrocytes and reticulocytes in the peripheral blood and content of erythrokaryocytes and erythroid precursors in the hemopoietic tissue of experimental animals. The effect of Recormon manifested immediately after injection, while the effect of Poetam was somewhat delayed, but more lasting (due to activation of host erythropoietin system).

  6. Catalytic activity of metallic nanoisland coatings. The influence of size effects on the recombination properties

    Science.gov (United States)

    Tomilina, O. A.; Berzhansky, V. N.; Tomilin, S. V.; Shaposhnikov, A. N.

    2016-08-01

    The results of investigations of the quantum-size effects influence on selective properties of heterogeneous nanocatalysts are presents. As etalon exothermic reaction was used the reaction of atomic hydrogen recombination. The nanostructured Pd and Pt films on Teflon substrate were used as a samples of heterogeneous nanocatalysts. It was shown that for nanoparticles with various sizes the catalytic activity has the periodic dependence. It has been found that for certain sizes of nanoparticles their catalytic activity is less than that of Teflon substrate.

  7. Highly efficient recombinant production and purification of streptococcal cysteine protease streptopain with increased enzymatic activity.

    Science.gov (United States)

    Lane, Michael D; Seelig, Burckhard

    2016-05-01

    Streptococcus pyogenes produces the cysteine protease streptopain (SpeB) as a critical virulence factor for pathogenesis. Despite having first been described seventy years ago, this protease still holds mysteries which are being investigated today. Streptopain can cleave a wide range of human proteins, including immunoglobulins, the complement activation system, chemokines, and structural proteins. Due to the broad activity of streptopain, it has been challenging to elucidate the functional results of its action and precise mechanisms for its contribution to S. pyogenes pathogenesis. To better study streptopain, several expression and purification schemes have been developed. These methods originally involved isolation from S. pyogenes culture but were more recently expanded to include recombinant Escherichia coli expression systems. While substantially easier to implement, the latter recombinant approach can prove challenging to reproduce, often resulting in mostly insoluble protein and poor purification yields. After extensive optimization of a wide range of expression and purification conditions, we applied the autoinduction method of protein expression and developed a two-step column purification scheme that reliably produces large amounts of purified soluble and highly active streptopain. This method reproducibly yielded 3 mg of streptopain from 50 mL of expression culture at >95% purity, with an activity of 5306 ± 315 U/mg, and no remaining affinity tags or artifacts from recombinant expression. This improved method therefore enables the facile production of the important virulence factor streptopain at higher yields, with no purification scars that might bias functional studies, and with an 8.1-fold increased enzymatic activity compared to previously described procedures.

  8. Matricryptins and matrikines: biologically active fragments of the extracellular matrix.

    Science.gov (United States)

    Ricard-Blum, Sylvie; Salza, Romain

    2014-07-01

    Numerous extracellular proteins and glycosaminoglycans (GAGs) undergo limited enzymatic cleavage resulting in the release of fragments exerting biological activities, which are usually different from those of the full-length molecules. In this review, we define matrikines and matricryptins, which are bioactive fragments released from the extracellular matrix proteins, proteoglycans and GAGs and report their major biological activities. These fragments regulate a number of physiopathological processes including angiogenesis, cancer, fibrosis, inflammation, neurodegenerative diseases and wound healing. The challenges to translate these fragments from molecules biologically active in vitro and in experimental models to potential drugs are discussed in the last part of the review.

  9. Ficus carica L. (Moraceae: Phytochemistry, Traditional Uses and Biological Activities

    Directory of Open Access Journals (Sweden)

    Shukranul Mawa

    2013-01-01

    Full Text Available This paper describes the botanical features of Ficus carica L. (Moraceae, its wide variety of chemical constituents, its use in traditional medicine as remedies for many health problems, and its biological activities. The plant has been used traditionally to treat various ailments such as gastric problems, inflammation, and cancer. Phytochemical studies on the leaves and fruits of the plant have shown that they are rich in phenolics, organic acids, and volatile compounds. However, there is little information on the phytochemicals present in the stem and root. Reports on the biological activities of the plant are mainly on its crude extracts which have been proven to possess many biological activities. Some of the most interesting therapeutic effects include anticancer, hepatoprotective, hypoglycemic, hypolipidemic, and antimicrobial activities. Thus, studies related to identification of the bioactive compounds and correlating them to their biological activities are very useful for further research to explore the potential of F. carica as a source of therapeutic agents.

  10. In vitro replication competence of a hepatitis B genotype D/A recombinant virus: dissimilar biological behaviour regarding its parental genotypes.

    Science.gov (United States)

    Trinks, Julieta; Sugiyama, Masaya; Tanaka, Yasuhito; Kurbanov, Fuat; Benetucci, Jorge; Giménez, Edgardo; Weissenbacher, Mercedes C; Mizokami, Masashi; Oubiña, José R

    2013-12-01

    Hepatitis B virus (HBV) DNA recombinants contribute to ~30% of the overall full-length sequences already deposited in GenBank. However, their biological behaviour has not been analysed so far. In this study, the in vitro replication kinetics of the first D/A recombinant from the American continent differed from its parental genotypes, exhibiting higher extracellular levels of HBV DNA and hepatitis B e antigen. Southern blots of intracellular core-associated HBV DNA were in agreement with such results. Because this recombinant was obtained from an Argentinian injecting drug user belonging to a vulnerable community, these results are of singular relevance for regional public health. Further in vivo studies are urgently needed to determine the pathogenicity of these replicative competent clones.

  11. Expression of an insect excitatory toxin, BmK IT, from the scorpion, Buthus martensii Karsch, and its biological activity.

    Science.gov (United States)

    Hao, Chan-juan; Xu, Cheng-gang; Wang, Wei; Chai, Bao-feng; Liang, Ai-hua

    2005-12-01

    An insect excitatory toxin from Buthus martensii Karsch (BmK IT) was cloned into the expression vector, pTWIN1, and expressed into Escherichia coli BL21 (DE3) host cells. The soluble fusion expression of CBD-intein-BmK IT was obtained. The recombinant BmK IT was purified by two anion-exchange chromatography columns and one gel chromatography column. Bioassays were carried out to verify the toxicity of this recombinant toxin. At the end of a 96 h experimental period, 83% of cotton bollworm larvae were killed with an LT(50) value of 58-62 h. Furthermore, the average weight of larvae fed on BmK IT-containing media was approx 4% of that of the control groups. The results indicate that the expressed and purified recombinant BmK IT has biological activity.

  12. 重组人胰岛素样生长因子1的原核可溶性表达及活性鉴定%Soluble Expression and Biological Activity Detection of Recombinant Human Insulin Like Growth Factor 1 in Prokaryotic System

    Institute of Scientific and Technical Information of China (English)

    范敏; 徐畅; 李彦; 孙文敬; 温娜

    2011-01-01

    目的:原核可溶性表达人胰岛素样生长因子1(hlGF-1)并进行生物活性测定.方法:PCR扩增hlGF-1核酸序列,克隆至融合表达载体pET-DsbC中,转入大肠杆菌BL21(DE3)进行诱导、表达和纯化,对融合蛋白DsbC -hlGF-1分别进行酶切、Western印迹和生物活性测定.结果:融合蛋白DsbC-hlGF-1在原核系统内经IPTG诱导,主要以可溶性形式表达,其表达量占菌体总蛋白量的30%~35%,Western印迹和MTT法测定结果证明重组DsbC-hlGF-1蛋白具有较强的抗原活性和明显的促NIH/3T3细胞增殖作用.结论:融合蛋白DsbC-hlGF在大肠杆菌中以可溶性表达形式存在并具有类似天然hIGF-1的生物活性,这对进一步研究hIGF-1的结构和功能,开发相应的基因工程产品具有重要意义.%Objective: To obtain human insulin like growth factor 1 (hIGF-1) expressed in solubility in prokary-otic system and detect its biological activity. Methods: hIGF-1 gene was obtained by PCR and was cloned into pET-DsbC fusion vector. After induction, expression and purification, the antigenicity and biological activity of the product were examined by Western blot and MTT assay. Results: DsbC-hlGF-1 was expressed in solubility in Es-cherichia coli and its expression quantity was about 30%~35% to bacterial total protein with IPTG induction, which also showed a strong antigenic ability and stimulate proliferation of NIH/3T3 cells evidently. Conclusion: The results suggest that the DsbC- hIGF-1 protein has the similar biological activity to wide type hIGF-1 and could express in solubility in E.coli It provides a futher investigation of the IGF-1 structure and function and build the foundation for the large scale production of IGF-1.

  13. Construction of eukaryotic expression plasmid of Fasciola hepatica CatL gene and biological activity analysis of recombinant protein%肝片吸虫组织蛋白酶L真核表达载体构建及重组蛋白活性分析

    Institute of Scientific and Technical Information of China (English)

    闻晓波; 冉旭华; 王春仁; 宋佰芬; 魏晓曼; 李晓娟; 王密; 苗艳

    2013-01-01

    In this research we constructed eukaryotic expression plasmid expressing Fasciola hepalica cathepsin L-like proteases (CatL) and analyzed the immunogenicity of recombinant protein. CatL gene was amplified by PCR with the template of recombinant pET30a-FhCatL plasmid and cloned into pEGFP-Nl vector to construct recombinant plasmids pEGFP-Nl-CatL. The recombinant plasmid pEGFP-Nl-CatL was transfected into HeLa cells and fluorescent signal was detected by fluorescence microscope. The immunogenicity of recombinant protein identified by western blotting demonstrated that the recombinant protein specifically reacted with serum from goat infected by Fasciola hepalica and showed robust immunoreactivity. The eukaryotic expression plasmid may be a potential gene vaccine or diagnose preparations in further study.%目的 构建肝片吸虫组织蛋白酶L(CatL)的真核表达载体,研究重组蛋白的生物学活性.方法 以构建好的重组质粒pET30a-FhCatL为模板,利用PCR技术扩增肝片吸虫组织蛋白酶L基因(catL),连接真核表达载体pEGFP-N1,构建重组质粒pEGFP-N1-CatL,转染HeLa细胞,荧光显微镜下观察绿色荧光,Western blotting检测重组蛋白表达情况.结果 重组质粒pEGFP-N1-CatL在HeLa细胞中获得了表达,Western blotting结果表明真核表达质粒表达的重组蛋白能与自然感染肝片吸虫的山羊阳性血清发生特异性反应.结论 肝片吸虫CatL真核表达载体构建成功,真核表达产物可与自然感染的山羊阳性血清发生特异性反应,具有免疫反应性,可做为分子疫苗的候选和诊断抗原进行进一步的研究.

  14. Biological activities of Juglans regia flowers

    Directory of Open Access Journals (Sweden)

    Seyed Fazel Nabavi

    2011-06-01

    Full Text Available Antidepressant, anti-inflammatory, antihypoxic and antioxidant activities of methanol extract of Juglans regia L., Juglandaceae, flower were investigated. Antidepressant activity was examined by forced swimming test and tail suspension test in mice. Antihypoxic activity was investigated in haemic and circulatory models. The effects were pronounced in both models. It produced statistically significant anti-inflammatory activity in carrageenan induced edema at nearly all doses, compared to control groups. IC50 for DPPH radical-scavenging activity was 674±27.6 µg mL-1. Extract showed good Fe2+ chelating ability (IC50 43±1.5 µg mL-1. It exhibited low antioxidant activity in linoleic acid peroxidation test. Its pharmacological effects may be attributed, in part, to the presence of phenols and ISSN 0102-695X flavonoids in the extract.

  15. Construction of an oral recombinant DNA vaccine from H pylori neutrophil activating protein and its immunogenicity

    Institute of Scientific and Technical Information of China (English)

    Bo Sun; Zhao-Shen Li; Zhen-Xing Tu; Guo-Ming Xu; Yi-Qi Du

    2006-01-01

    AIM: To construct a live attenuated Salmonella typhimurium (S.typhimurium) strain harboring the H pylori neutrophil activating protein (HP-NAP) gene as an oral recombinant DNA vaccine, and to evaluate its immunogenicity.METHODS: By genetic engineering methods, the genomic DNA of H pylori was extracted as a template. The total length of the HP-NAP gene was amplified by polymerase chain reaction (PCR) and cloned into pBT vector for sequencing and BLAST analysis, then subcloned into a eukaryotic expression vector pIRES followed by PCR identification and restriction enzyme digestion. The identified recombinant plasmid pIRES-NAP was transfected into COS-7 cells for target fusion protein expression, and its antigenicity was detected by Western blotting. Then the recombinant plasmid was transformed into a live attenuated S. typhimurium strain SL7207 as an oral vaccine strain, and its immunogenicity was evaluated with animal experiments.RESULTS: A 435 bp product was cloned using high homology with HP-NAP gene in GenBank (more than 98%). With identification by PCR and restriction enzyme digestion, a recompinant eukaryotic expression plasmid pIRES-NAP containing the HP-NAP gene of H pylori was successfully constructed. The expressed target protein had a specific reaction with H pylor(i) whole cell antibody and showed a single strip result detected by Western blotting. Oral immunization of mice with recombinant DNA vaccine strain SL7207 (pIRES-NAP) also induced a specific immune response.CONCLUSION: The successful construction of HP-NAP oral DNA vaccine with good immunogenicity may help to further investigate its immunoprotection effects and develop vaccine against H pylori infection.

  16. Biological activities of Croton palanostigma Klotzsch

    Directory of Open Access Journals (Sweden)

    Eduardo Ferreira Mota

    2015-01-01

    Full Text Available Background: Different species of Croton are used in traditional Amazonian medicine. Among the popular uses are treatment of bacterial diseases, poorly healing wounds and fevers. Objective: This study evaluated the antileishmanial, antiplasmodial and antimicrobial activities of the extracts and diterpenes of Croton palanostigma Klotzsch (Euphorbiaceae. Materials and Methods: Leaves and bark were extracted with dichloromethane and methanol. The bark dichloromethane extract (BDE was chromatographed on a column, obtaining cordatin and aparisthman. The extracts and diterpenes were assayed thought agar disk diffusion method and their bactericidal or fungicidal effects were evaluated by minimum bactericidal or fungicidal concentration. The antiplasmodial activity was evaluated after 24 and 72 h of exposition. The antileishmanial activity was performed on promastigotes forms of Leishmania amazonensis. Results: The bark methanol extract (BME and cordatin were not active against any microbial strains tested; BDE and leaves methanol extract (LME were positive for Pseudomonas aeruginosa and aparisthman was positive for Candida albicans. In the determination of the minimum bactericidal concentration, neither of them were active in the highest concentration tested. The extracts and diterpenes were inactive in Plasmodium falciparum, except the LME in 72 h. Any extract was shown to be active in promastigote forms of L. amazonensis. Conclusion: These results indicate that the BDE and LME did not inhibit the bacterial growth, then they probably had bacteriostatic effect. LME presented activity in P. falciparum.

  17. High Resolution Screening of biologically active compounds and metabolites

    NARCIS (Netherlands)

    Kool, J.

    2007-01-01

    High Resolution Screening of biologically active compounds and metabolites Jeroen Kool Biotransformation enzymes play a crucial role in the metabolism of both endogenous compounds and xenobiotics. Usually, the detoxication of these compounds by biotransformation enzymes results in harmless metab

  18. Synthesis and Biological Activity of Philanthotoxin Analogs

    Institute of Scientific and Technical Information of China (English)

    Yong An ZHANG; Ke Zhong LIU; Deng Yuan WANG; Yu Zhu WANG; Liang Jian QU; Chang Jin ZHU

    2006-01-01

    The synthesis of four analogs of philanthotoxin is described. The preliminary bioassay showed that these compounds all had good insecticidal activities, and the compound 6a had the best killing effect.

  19. Total synthesis and biological activity of natural product Urukthapelstatin A.

    Science.gov (United States)

    Lin, Chun-Chieh; Tantisantisom, Worawan; McAlpine, Shelli R

    2013-07-19

    Herein we report the first total synthesis of the natural product Urkuthaplestatin A (Ustat A) utilizing a convergent synthetic strategy. The characterization and biological activity match those of the previously published natural product. Interestingly, several intermediates, including the linear and serine cyclized precursors, show a 100-fold decrease in cytotoxicity, with IC50's in the low micromolar range. These data indicate that the rigidity and the consecutive aromatic heterocyclic system are responsible for the biological activity.

  20. Phytochemicals and Their Biological Activities of Plants in Tagetes L.

    Institute of Scientific and Technical Information of China (English)

    XU Li-wei; CHEN Juan; QI Huan-yang; SHI Yan-ping

    2012-01-01

    Tagetes L.,the genus in the family Asteraceae,consists of about 30 species spread in South and Middle America as well as Mexico.More than one hundred secondary metabolites have been obtained in phytochemical investigation on the species,some of which have potent biological activities.The advances in phytochemical studies and biological activities of the plants in Tagetes L.from 1925 to 2011 are summarized in this paper.

  1. TiO2 nanotubes functionalized with recombinant human bone morphogenetic protein-2 enhance biological activity in vitro%二氧化钛纳米管阵列加载重组人骨形成蛋白2的体外生物活性研究

    Institute of Scientific and Technical Information of China (English)

    孙子环; 夏荣; 孙磊; 胡小晔; 闵曦; 徐基亮

    2015-01-01

    目的 探讨二氧化钛纳米管阵列加载重组人骨形成蛋白2 (recombinant human bone morphogenetic protein-2,rhBMP-2)对小鼠骨髓间充质干细胞(bone mesenchymal stem cell,BMSC)早期活性的影响,为钛种植体表面生物化学改性提供实验依据.方法 利用阳极氧化技术在纯钛片表面制备双层二氧化钛纳米管阵列,化学接枝rhBMP-2(实验组),以机械抛光的纯钛为空白对照组,阴性对照A组为二氧化钛纳米管组、阴性对照B组为二氧化钛纳米管+羰基二咪唑组.用场发射扫描电镜观察各组形貌并用X射线光电子能谱仪检测各组元素.各组试件与BMSC共培养,检测第1天各组细胞黏附铺展情况(每组样本量为3),第1、3、5天各组细胞增殖A值及第5、7、11天各组碱性磷酸酶活性(每组每个时间点样本量为3).结果 场发射扫描电镜示实验组表面可见粟粒状颗粒物,X射线光电子能谱仪示实验组氮峰明显增高.场发射扫描电镜示第1天实验组细胞黏附铺展良好,细胞间连接广泛而紧密,均优于其他3组.第1天各组细胞增殖效果不明显;第3、5天实验组A值(3.295±0.153、3.823±0.059)均显著高于空白对照组(2.479±0.064、3.131±0.096)、阴性对照A组(2.715±0.075、3.371±0.047)及阴性对照B组(2.756±0.132、3.637±0.047) (P<0.05);第5、7、11天实验组碱性磷酸酶活性(0.0477±0.0287、0.0615±0.0016、0.0667±0.0018)均显著高于其他3组(P<0.05).结论 二氧化钛纳米管阵列可通过生物化学法加载rhBMP-2并具备良好的生物相容性.%Objective To investigate the effect of TiO2 nanotube arrays covalently modified by recombinant human bone morphogenetic protein-2(rhBMP-2) on the early bioactivity of mesenchymal stem cells(MSC) in vitro and to provide experimental evidence for the biochemical modification of titanium implants.Methods In the experiment group,double titanium nanotube arrays were prepared by anodization,and were chemically

  2. Biological activity of a genetically modified BMP-2 variant with inhibitory activity

    Directory of Open Access Journals (Sweden)

    Kübler Alexander C

    2009-02-01

    Full Text Available Abstract Background Alterations of the binding epitopes of bone morphogenetic protein-2 (BMP-2 lead to a modified interaction with the ectodomains of BMP receptors. In the present study the biological effect of a BMP-2 double mutant with antagonistic activity was evaluated in vivo. Methods Equine-derived collagenous carriers were loaded with recombinant human BMP-2 (rhBMP-2 in a well-known dose to provide an osteoinductive stimulus. The study was performed in a split animal design: carriers only coupled with rhBMP-2 (control were implanted into prepared cavities of lower limb muscle of rats, specimens coupled with rhBMP-2 as well as BMP-2 double mutant were placed into the opposite limb in the same way. After 28 days the carriers were explanted, measured radiographically and characterized histologically. Results As expected, the BMP-2 loaded implants showed a typical heterotopic bone formation. The specimens coupled with both proteins showed a significant decreased bone formation in a dose dependent manner. Conclusion The antagonistic effect of a specific BMP-2 double mutant could be demonstrated in vivo. The dose dependent influence on heterotopic bone formation by preventing rhBMP-2 induced osteoinduction suggests a competitive receptor antagonism.

  3. Does intravenous administration of recombinant tissue plasminogen activator for ischemic stroke can cause inferior myocardial infarction?

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    Mostafa Almasi

    2016-06-01

    Full Text Available Recombinant tissue plasminogen activator (rTPA is one of the main portions of acute ischemic stroke management, but unfortunately has some complications. Myocardial infarction (MI is a hazardous complication of administration of intravenous rTPA that has been reported recently. A 78-year-old lady was admitted for elective coronary artery bypass graft surgery. On the second day of admission, she developed acute left hemiparesis and intravenous rTPA was administered within 120 minutes. Three hours later, she has had chest pain. Rescue percutaneous coronary intervention was performed on right coronary artery due to diagnosis of inferior MI, and the symptoms were resolved.

  4. Biological Activities of a Thai Luminescent Mushroom

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    Jiraporn BURAKORN

    2015-06-01

    Full Text Available Wild fruit bodies of luminescent mushrooms were collected from wood stumps over a period covering August to October 2011 in the Kosumpisai forest, Mahasarakham province, in the Northeast of Thailand. A study of the morphological and genetic characteristics of the luminescent mushroom suggested that it was Neonothopanus nimbi KS. The fruiting bodies and mycelium of Neonothopanus nimbi KS were assayed for their antimicrobial activities, antifungal activity, inhibitory activity against avian influenza H5N1 neuraminidase (NA, and anticancer activity, using organic solvent extracts. The results showed that only the methanol extract of mycelia was effective against Bacillus sphaericus, with the widest inhibition zone of 11.66±2.71 mm, but this was not effective against the other 3 bacteria (Pseudomonas aeruginosa, Serratia marcescens, and Escherichia coli. On the other hand, all of the fruit body extracts were inactive against all four bacteria. The ethylacetate extract of mycelia inhibited the NCI-H187 small lung cancer cell line, KB oral cavity cancer cell line, and the MCF7 breast cancer cell line, including Magnaporthe grisea and Curvularia lunata. The methanol extract of mycelia inhibited the KB oral cavity cell cancer cell line, Magnaporthe grisea, and Curvularia lunata at 96.66, 95.32 and 95.41 %, respectively. The results imply that polar extracts of mycelia are a resource of bioactive compounds, whereas extracts of fruit bodies have less inhibitory activity against cancer, phytopathogenic-fungi and H5N1 neuraminidase.

  5. Biologically active secondary metabolites from Asphodelus microcarpus.

    Science.gov (United States)

    Ghoneim, Mohammed M; Ma, Guoyi; El-Hela, Atef A; Mohammad, Abd-Elsalam I; Kottob, Saeid; El-Ghaly, Sayed; Cutler, Stephen J; Ross, Samir A

    2013-08-01

    Bioassay guided fractionation of the ethanolic extract of Asphodelus microcarpus Salzm.et Vivi (Asphodelaceae) resulted in the isolation of one new metabolite, 1,6-dimethoxy-3-methyl-2-naphthoic acid (1) as well as nine known compounds: asphodelin (2), chrysophanol (3), 8-methoxychrysophanol (4), emodin (5), 2-acetyl-1,8-dimethoxy-3-methylnaphthalene (6), 10-(chrysophanol-7'-yl)-10-hydroxychrysophanol-9-anthrone (7), aloesaponol-III-8-methyl ether (8), ramosin (9) and aestivin (10). The compounds were identified by 1D and 2D NMR and HRESIMS. Compounds 3, 6 and 10 were isolated for the first time from this species. Compounds 3 and 4 showed moderate to weak antileishmanial activity with IC50 values of 14.3 and 35.1 microg/mL, respectively. Compound 4 exhibited moderate antifungal activity against Cryptococcus neoformans with an IC50 value of 15.0 microg/mL, while compounds 5, 7 and 10 showed good to potent activity against methicillin resistant Staphylococcus aureus (MRSA) with IC50 values of 6.6, 9.4 microg/mL and 1.4 microg/mL respectively. Compounds 5, 8 and 9 displayed good activity against S. aureus with IC50 values of 3.2, 7.3 and 8.5 microg/mL, respectively. Compounds 7 and 9 exhibited a potent cytotoxic activity against leukemia LH60 and K562 cell lines. Compound 10 showed potent antimalarial activities against both chloroquine-sensitive and chloroquine-resistant strains of Plasmodium falciparum with IC50 values in the range of 0.8-0.7 microg/mL without showing any cytotoxicity to mammalian cells.

  6. Expression, Purification and Activity Assay of Two New Recombinant Antagonists of Fibrinogen Receptor

    Directory of Open Access Journals (Sweden)

    Jianbo Yang

    2005-01-01

    Full Text Available The gene sequence of Decorsin which is extracted from a kind of North American leeches was synthesized. Two recombinant proteins, Annexin V plus Decorsin (AnnV-D39 and Annexin V plus the carboxyl terminal 27 amino acid residues of Decorsin(AnnV-D27, were constructed. And a 10 amino acids linker peptide of GGGGSGGGGS was inserted between Annexin V and Decorsin in AnnV-D39. Using pET-28(a+ as an expressing vector, both two recombinant proteins were expressed in E. Coli BL21(DE3 with high efficiency as inclusion bodies. The expression products were purified by DEAE-Cellulose 52 and Sepharose CL-4B chromatography under denaturing condition. Platelet Aggregation Assay (PAA shows that AnnV-D39 has good anti-platelet aggregation activity. However, AnnV-D27 shows no such activities in any PAA test. AnnV-D39 shows good anti-platelet aggregation activity as a new antagonist of fibrinogen receptor, while Annv-D27 needs re-modification

  7. A Butter Aroma Recombinate Activates Human Class-I Odorant Receptors.

    Science.gov (United States)

    Geithe, Christiane; Andersen, Gaby; Malki, Agne; Krautwurst, Dietmar

    2015-11-01

    With ∼400 olfactory G protein-coupled receptors (GPCR), humans sensitively perceive ∼230 key aroma compounds as best natural agonists of ∼10000 food volatiles. An understanding of odorant coding, thus, critically depends on the knowledge about interactions of key food aroma chemicals and their mixtures with their cognate receptors. Genetically designed test cell systems enable the screening, deorphaning, and characterization of single odorant receptors (OR). This study shows for the food aroma-specific and quantitative butter aroma recombinate, and its single components, specific in vitro class-I OR activity patterns, as well as the activation of selected OR in a concentration-dependent manner. Recently, chemosensory receptors, especially class-I OR, were demonstrated to be expressed on blood leukocytes, which may encounter foodborne aroma compounds postprandially. This study shows that butter aroma recombinate induced chemotaxis of isolated human neutrophils in a defined gradient, and in a concentration-dependent and pertussis toxin-sensitive manner, suggesting at least a GPCR-mediated activation of blood leukocytes by key food odorants.

  8. Refolded Recombinant Human Paraoxonase 1 Variant Exhibits Prophylactic Activity Against Organophosphate Poisoning.

    Science.gov (United States)

    Bajaj, Priyanka; Tripathy, Rajan K; Aggarwal, Geetika; Datusalia, Ashok K; Sharma, Shyam S; Pande, Abhay H

    2016-09-01

    Organophosphate (OP) compounds are neurotoxic chemicals, and current treatments available for OP-poisoning are considered as unsatisfactory and inadequate. There is an urgent need for the development of more effective treatment(s) for OP-poisoning. Human paraoxonase 1 (h-PON1) is known to hydrolyze a variety of OP-compounds and is a leading candidate for the development of prophylactic and therapeutic agent against OP-poisoning in humans. Non-availability of effective system(s) for the production of recombinant h-PON1 (rh-PON1) makes it hard to produce improved variant(s) of this enzyme and analyze their in vivo efficacy in animal models. Production of recombinant h-PON1 (rh-PON1) using an Escherichia coli expression system is a key to develop variant(s) of h-PON1. Recently, we have developed a procedure to produce active rh-PON1 enzymes by using E. coli expression system. In this study, we have characterized the OP-hydrolyzing properties of refolded rh-PON1(wt) and rh-PON1(H115W;R192K) variant. Our results show that refolded rh-PON1(H115W;R192K) variant exhibit enhanced OP-hydrolyzing activity in in vitro and ex vivo assays and exhibited prophylactic activity in mouse model of OP-poisoning, suggesting that refolded rh-PON1 can be developed as a therapeutic candidate.

  9. Transport and biological activities of bile acids.

    Science.gov (United States)

    Zwicker, Brittnee L; Agellon, Luis B

    2013-07-01

    Bile acids have emerged as important biological molecules that support the solubilization of various lipids and lipid-soluble compounds in the gut, and the regulation of gene expression and cellular function. Bile acids are synthesized from cholesterol in the liver and eventually released into the small intestine. The majority of bile acids are recovered in the distal end of the small intestine and then returned to the liver for reuse. The components of the mechanism responsible for the recycling of bile acids within the enterohepatic circulation have been identified whereas the mechanism for intracellular transport is less understood. Recently, the ileal lipid binding protein (ILBP; human gene symbol FABP6) was shown to be needed for the efficient transport of bile acids from the apical side to the basolateral side of enterocytes in the distal intestine. This review presents an overview of the transport of bile acids between the liver and the gut as well as within hepatocytes and enterocytes. A variety of pathologies is associated with the malfunction of the bile acid transport system.

  10. The Meiotic Recombination Activator PRDM9 Trimethylates Both H3K36 and H3K4 at Recombination Hotspots In Vivo.

    Science.gov (United States)

    Powers, Natalie R; Parvanov, Emil D; Baker, Christopher L; Walker, Michael; Petkov, Petko M; Paigen, Kenneth

    2016-06-01

    In many mammals, including humans and mice, the zinc finger histone methyltransferase PRDM9 performs the first step in meiotic recombination by specifying the locations of hotspots, the sites of genetic recombination. PRDM9 binds to DNA at hotspots through its zinc finger domain and activates recombination by trimethylating histone H3K4 on adjacent nucleosomes through its PR/SET domain. Recently, the isolated PR/SET domain of PRDM9 was shown capable of also trimethylating H3K36 in vitro, raising the question of whether this reaction occurs in vivo during meiosis, and if so, what its function might be. Here, we show that full-length PRDM9 does trimethylate H3K36 in vivo in mouse spermatocytes. Levels of H3K4me3 and H3K36me3 are highly correlated at hotspots, but mutually exclusive elsewhere. In vitro, we find that although PRDM9 trimethylates H3K36 much more slowly than it does H3K4, PRDM9 is capable of placing both marks on the same histone molecules. In accord with these results, we also show that PRDM9 can trimethylate both K4 and K36 on the same nucleosomes in vivo, but the ratio of K4me3/K36me3 is much higher for the pair of nucleosomes adjacent to the PRDM9 binding site compared to the next pair further away. Importantly, H3K4me3/H3K36me3-double-positive nucleosomes occur only in regions of recombination: hotspots and the pseudoautosomal (PAR) region of the sex chromosomes. These double-positive nucleosomes are dramatically reduced when PRDM9 is absent, showing that this signature is PRDM9-dependent at hotspots; the residual double-positive nucleosomes most likely come from the PRDM9-independent PAR. These results, together with the fact that PRDM9 is the only known mammalian histone methyltransferase with both H3K4 and H3K36 trimethylation activity, suggest that trimethylation of H3K36 plays an important role in the recombination process. Given the known requirement of H3K36me3 for double strand break repair by homologous recombination in somatic cells, we

  11. Expression and purification of recombinant human neuritin from Pichia pastoris and a partial analysis of its neurobiological activity in vitro.

    Science.gov (United States)

    Zhang, Yunhua; Zhang, Shujun; Xian, Lingling; Tang, Juan; Zhu, Jingling; Cui, Lijuan; Li, Shanman; Yang, Lei; Huang, Jin

    2015-10-01

    Neuritin (also known as candidate plasticity gene 15 (cpg15)) is a neurotrophic factor that was recently discovered in a screen aimed at identifying genes involved in activity-dependent synaptic plasticity. Neuritin plays multiple roles in both neural development (Chen et al. Zhonghua Yan Ke Za Zhi 46:978-983 2010; Corriveau et al. J Neurosci 19:7999-8008 1999; Lee and Nedivi J Neurosci 22:1807-1815 2002) and synaptic plasticity (Fujino et al. Gene Dev 25:2674-2685 2011; Leslie and Nedivi Prog 14 Neurobiol 94:223-237 2011; Loebrich and Nedivi Physiol Rev 89:1079 2009). In this study, to produce bioactive, soluble recombinant human neuritin protein, a portion of NRN1 was cloned into the Pichia pastoris expression vector pPIC9K. The recombinant vector was then transformed into the methylotrophic yeast strain P. pastoris GS115, and a shaking flask method and His-tag purification strategy were utilized to express and purify neuritin protein. The resulting protein had a molecular mass of approximately 11 kDa, and subsequent functional analyses indicated that the purified neuritin promoted neurite outgrowth from embryonic chicken dorsal root ganglions, while also prolonging the survival of these ganglions, and from PC12 cells. These findings suggest that neuritin was expressed effectively in vitro and that this protein may play a role in stimulating neurite outgrowth of both dorsal root ganglions and PC12 cells. This study provides a novel strategy for the large-scale production of bioactive neuritin, which will enable further study of the biological function of this protein.

  12. Physical aspects of biological activity and cancer

    Science.gov (United States)

    Pokorný, Jiří

    2012-03-01

    Mitochondria are organelles at the boundary between chemical-genetic and physical processes in living cells. Mitochondria supply energy and provide conditions for physical mechanisms. Protons transferred across the inner mitochondrial membrane diffuse into cytosol and form a zone of a strong static electric field changing water into quasi-elastic medium that loses viscosity damping properties. Mitochondria and microtubules form a unique cooperating system in the cell. Microtubules are electrical polar structures that make possible non-linear transformation of random excitations into coherent oscillations and generation of coherent electrodynamic field. Mitochondria supply energy, may condition non-linear properties and low damping of oscillations. Electrodynamic activity might have essential significance for material transport, organization, intra- and inter-cellular interactions, and information transfer. Physical processes in cancer cell are disturbed due to suppression of oxidative metabolism in mitochodria (Warburg effect). Water ordering level in the cell is decreased, excitation of microtubule electric polar oscilations diminished, damping increased, and non-linear energy transformation shifted towards the linear region. Power and coherence of the generated electrodynamic field are reduced. Electromagnetic activity of healthy and cancer cells may display essential differences. Local invasion and metastastatic growth may strongly depend on disturbed electrodynamic activity. Nanotechnological measurements may disclose yet unknown properties and parameters of electrodynamic oscillations and other physical processes in healthy and cancer cells.

  13. Albinism-causing mutations in recombinant human tyrosinase alter intrinsic enzymatic activity.

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    Monika B Dolinska

    Full Text Available BACKGROUND: Tyrosinase (TYR catalyzes the rate-limiting, first step in melanin production and its gene (TYR is mutated in many cases of oculocutaneous albinism (OCA1, an autosomal recessive cause of childhood blindness. Patients with reduced TYR activity are classified as OCA1B; some OCA1B mutations are temperature-sensitive. Therapeutic research for OCA1 has been hampered, in part, by the absence of purified, active, recombinant wild-type and mutant human enzymes. METHODOLOGY/PRINCIPAL FINDINGS: The intra-melanosomal domain of human tyrosinase (residues 19-469 and two OCA1B related temperature-sensitive mutants, R422Q and R422W were expressed in insect cells and produced in T. ni larvae. The short trans-membrane fragment was deleted to avoid potential protein insolubility, while preserving all other functional features of the enzymes. Purified tyrosinase was obtained with a yield of >1 mg per 10 g of larval biomass. The protein was a monomeric glycoenzyme with maximum enzyme activity at 37°C and neutral pH. The two purified mutants when compared to the wild-type protein were less active and temperature sensitive. These differences are associated with conformational perturbations in secondary structure. CONCLUSIONS/SIGNIFICANCE: The intramelanosomal domains of recombinant wild-type and mutant human tyrosinases are soluble monomeric glycoproteins with activities which mirror their in vivo function. This advance allows for the structure - function analyses of different mutant TYR proteins and correlation with their corresponding human phenotypes; it also provides an important tool to discover drugs that may improve tyrosinase activity and treat OCA1.

  14. Current trends of HIV recombination worldwide

    Directory of Open Access Journals (Sweden)

    Katherine A. Lau

    2013-06-01

    Full Text Available One of the major characteristics of HIV-1 is its high genetic variability and extensive heterogeneity. This characteristic is due to its molecular traits, which in turn allows it to vary, recombine, and diversify at a high frequency. As such, it generates complex molecular forms, termed recombinants, which evade the human immune system and so survive. There is no sequence constraint to the recombination pattern as it appears to occur at inter-group (between groups M and O, as well as inter- and intra-subtype within group M. Rapid emergence and active global transmission of HIV-1 recombinants, known as circulating recombinant forms (CRFs and unique recombinant forms (URFs, requires urgent attention. To date, 55 CRFs have been reported around the world. The first CRF01_AE originated from Central Africa but spread widely in Asia. The most recent CRF; CRF55_01B is a recombinant form of CRF01_AE and subtype B, although its origin is yet to be publicly disclosed. HIV-1 recombination is an ongoing event and plays an indispensable role in HIV epidemics in different regions. Africa, Asia and South America are identified as recombination hot-spots. They are affected by continual emergence and co-circulation of newly emerging CRFs and URFs, which are now responsible for almost 20% of HIV-1 infections worldwide. Better understanding of recombinants is necessary to determine their biological and molecular attributes.

  15. Rhodiola plants: Chemistry and biological activity

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    Hsiu-Mei Chiang

    2015-09-01

    Full Text Available Rhodiola is a genus of medicinal plants that originated in Asia and Europe and are used traditionally as adaptogens, antidepressants, and anti-inflammatory remedies. Rhodiola plants are rich in polyphenols, and salidroside and tyrosol are the primary bioactive marker compounds in the standardized extracts of Rhodiola rosea. This review article summarizes the bioactivities, including adaptogenic, antifatigue, antidepressant, antioxidant, anti-inflammatory, antinoception, and anticancer activities, and the modulation of immune function of Rhodiola plants and its two constituents, as well as their potential to prevent cardiovascular, neuronal, liver, and skin disorders.

  16. Expression and purification of an active cecropin-like recombinant protein against multidrug resistance Escherichia coli.

    Science.gov (United States)

    Téllez, Germán Alberto; Castaño-Osorio, Jhon Carlos

    2014-08-01

    Lucilin is a 36 residue cecropin antimicrobial peptide identified as a partial genetic sequence in Lucilia sericata maggots. The antimicrobial spectrum and toxicity profile of Lucilin is unknown. We first report the expression of Lucilin as an active recombinant fusion protein with a cysteine protease domain (CPD) tag. The fusion protein, GWLK-Lucilin-CPD-His8, showed maximum overexpression in Escherichia coli BL21 cells after 12h induction with 0.5mM IPTG (isopropyl beta-d-thiogalactoside) and growth conditions were 37 °C and 150 rpm shaking. The fusion protein was expressed as a soluble form and was purified by Ni-IMAC. The purified protein was active against E. coli ATCC 35218 with a MIC of 0.68 μM, and a clinical isolate of E. coli with extended spectrum beta-lactamase (ESBL) with a MIC of 0.8 μM. The recombinant GWLK-Lucilin-CPD-His8 was not toxic against human erythrocytes or Vero cells with a therapeutic index >63. The results suggest that GWLK-Lucilin-CPD-His8 represents a potential candidate for therapy against multidrug resistant Gram-negative bacteria.

  17. Expression, Purification and Activity Assay of the Recombinant Protein of Catechol-O-Methyltransferase from Chinese White Shrimp (Fenneropenaeus chinensis

    Directory of Open Access Journals (Sweden)

    Dian-Xiang Li

    2010-01-01

    Full Text Available Problem statement: We have previously cloned a gene of Chinese white shrimp Catechol O-Methyltransferase (designated Fc-COMT and characterized the gene expression pattern. In this study, expression and purification as well as activity assay of the recombinant Fc-COMT was further conducted. Approach: Using pET-30a (+ as a prokaryotic expression vector, the recombinant Fc- COMT was expressed in the supernatant of Escherichia coli lysate and easily purified by His-Bind resin chromatography. SDS-PAGE analysis showed that the molecular mass of recombinant Fc-COMT was approximately 30,000 Da, in good agreement with the software-predicted molecular weight. The enzymatic activity of recombinant Fc-COMT was tested using Dihydroxybenzoic Acid (DHBAc as a substrate. Results: The methyl products of DHBAc, Vanillic Acid (VA and Isovanillic Acid (IVA, were detected in the enzymatic reaction mixture with recombinant Fc-COMT by High Performance Liquid Chromatography-Mass Spectrometry (HPLC-MS. Conclusion: The recombinant Fc-COMT has catalytic activity of transferring methyl group from S-Adenosyl-L-Methionine (SAM to the 3’ hydroxyl or 4’ hydroxyl group of benzyl ring of DHBAc.

  18. Saponins from Swartzia langsdorffii: biological activities

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    Magalhães Aderbal Farias

    2003-01-01

    Full Text Available The presence of saponins and the molluscicidal activity of the roots, leaves, seeds and fruits of Swartzia langsdorffii Raddi (Leguminosae against Biomphalaria glabrata adults and eggs were investigated. The roots, seeds and fruits were macerated in 95% ethanol. These extracts exerted a significant molluscicidal activity against B. glabrata, up to a dilution of 100 mg/l. Four mixtures (A2, B2, C and D of triterpenoid oleanane type saponins were chromatographically isolated from the seed and fruit extracts. Two known saponins (1 and 2 were identified as beta-D-glucopyranosyl-[alpha-L-rhamnopyranosyl-(1->3- beta-D-glucuronopyranosyl-(1->3]-3beta-hydroxyolean-12-ene-28 -oate, and beta-D-glucopyranosyl-(1->3-beta-D-glucuronopyranosyl-(1 ->3]-3beta-hydroxyolean-12-ene-28-oate, respectively. These two saponins were present in all the mixtures, together with other triterpenoid oleane type saponins, which were shown to be less polar, by reversed-phase HPLC. The saponin identifications were based on spectral evidence, including ¹H-¹H two-dimensional correlation spectroscopy, nuclear Overhauser and exchange spectroscopy, heteronuclear multiple quantum coherence, and heteronuclear multiple-bond connectivity experiments. The toxicity of S. langsdorffii saponins to non-target organisms was prescreened by the brine shrimp lethality test.

  19. Hydrolysis and transglycosylation activity of a thermostable recombinant beta-glycosidase from Sulfolobus acidocaldarius.

    Science.gov (United States)

    Park, Ah-Reum; Kim, Hye-Jung; Lee, Jung-Kul; Oh, Deok-Kun

    2010-04-01

    We expressed a putative beta-galactosidase from Sulfolobus acidocaldarius in Escherichia coli and purified the recombinant enzyme using heat treatment and Hi-Trap ion-exchange chromatography. The resultant protein gave a single 57-kDa band by SDS-PAGE and had a specific activity of 58 U/mg. The native enzyme existed as a dimer with a molecular mass of 114 kDa by gel filtration. The maximum activity of this enzyme was observed at pH 5.5 and 90 degrees C. The half-lives of the enzyme at 70, 80, and 90 degrees C were 494, 60, and 0.2 h, respectively. The hydrolytic activity with p-nitrophenyl(pNP) substrates followed the order p-nitrophenyl-beta-D-fucopyranoside > pNP-beta-D-glucopyranoside > pNP-beta-D-galactopyranoside > pNP-beta-D-mannopyranoside > pNP-beta-D-xylopyranoside, but not toward aryl-alpha-glycosides or pNP-beta-L-arabinofuranoside. Thus, the enzyme was actually a beta-glycosidase. The beta-glycosidase exhibited transglycosylation activity with pNP-beta-D-galactopyranoside, pNP-beta-D-glucopyranoside, and pNP-beta-D-fucopyranoside in decreasing order of activity, in the reverse order of its hydrolytic activity. The hydrolytic activity was higher toward cellobiose than toward lactose, but the transglycosylation activity was lower with cellobiose than with lactose.

  20. Biologically active extracts with kidney affections applications

    Science.gov (United States)

    Pascu (Neagu), Mihaela; Pascu, Daniela-Elena; Cozea, Andreea; Bunaciu, Andrei A.; Miron, Alexandra Raluca; Nechifor, Cristina Aurelia

    2015-12-01

    This paper is aimed to select plant materials rich in bioflavonoid compounds, made from herbs known for their application performances in the prevention and therapy of renal diseases, namely kidney stones and urinary infections (renal lithiasis, nephritis, urethritis, cystitis, etc.). This paper presents a comparative study of the medicinal plant extracts composition belonging to Ericaceae-Cranberry (fruit and leaves) - Vaccinium vitis-idaea L. and Bilberry (fruit) - Vaccinium myrtillus L. Concentrated extracts obtained from medicinal plants used in this work were analyzed from structural, morphological and compositional points of view using different techniques: chromatographic methods (HPLC), scanning electronic microscopy, infrared, and UV spectrophotometry, also by using kinetic model. Liquid chromatography was able to identify the specific compounds of the Ericaceae family, present in all three extracts, arbutosid, as well as specific components of each species, mostly from the class of polyphenols. The identification and quantitative determination of the active ingredients from these extracts can give information related to their therapeutic effects.

  1. SYNTHESIS, CHARACTERIZATION AND BIOLOGICAL ACTIVITY OF POLYKETONES

    Institute of Scientific and Technical Information of China (English)

    Ismail A.Alkskas; Altaher M.Alhubge; Faizul Azam

    2013-01-01

    Polyketone resins have been prepared by the Friedel-Crafts polymerization of dithiophenylidenecyclopentanone (Ⅰ),dithiophenylidenecyclohexanone (Ⅱ) and dithiophenylideneacetone (Ⅲ) with adipoyl,sebacoyl and terephthaloyl dichlorides using boron trifluoride as catalyst and carbon disulphide as solvent.Polymers were characterized with IR,1H-NMR,and the results showed the presence of carbonyl of ketonic groups in the main chain.The polyketones have inherent viscosities of 0.40-0.70 dL/g.All the polymers are semicrystalline and most of them are partially soluble in most common organic solvents but freely soluble in aprotic solvents.The temperatures of 50% weight loss are as high as 185℃ to 280℃ in air,indicating that these aromatic polyketones have excellent thermal stability.All the polyketones were tested for their antimicrobial activity against bacteria and fungi.

  2. Intermolecular DNA ligation activity of eukaryotic toposiomerase II: Potential roles in nucleic acid recombination

    Energy Technology Data Exchange (ETDEWEB)

    Gale, K.C.R.

    1992-01-01

    Single-stranded [phi]X174 (+) strand DNA was used as a model substrate for topoisomerase II to determine whether double-stranded DNA cleavage observed in vitro reflects the in vivo intermediate in the enzyme's catalytic cycle and to investigate potential mechanisms for topoisomerase II-mediated DNA recombination. As found previously for topoisomerase II-mediated cleavage of double-stranded DNA, the enzyme was covalently linked to the 5[prime]-termini of cleaved [phi]X174 molecules. Optimal reaction conditions were similar for the two substrates. In contrast to results with double-stranded molecules, single-stranded DNA cleavage increased with time, was not reversible, and did not require the presence of SDS. Cleavage products generated in the absence of protein denaturant contained free 3[prime]-OH DNA termini. These results strongly suggest that the covalent topoisomerase II-cleaved DNA complex observed in vitro is the active intermediate in the enzyme's catalytic code. Topoisomerase II is capable of joining cleaved [phi]X174 (+) strand DNA to duplex oligonucleotide acceptor molecules by an intermolecular ligation reaction. Intermolecular DNA ligation proceeded in a time and oligonucleotide concentration dependent fashion. The covalent linkage is between the 5[prime]-phosphate of [phi]X174 (+) strand DNA and the 3[prime]-OH of oligonucleotide acceptor molecules. The reaction was dependent on the presence of a divalent cation, was inhibited by salt, and was not affected by the presence of ATP. The enzyme was capable of ligating [phi]X174 (+) strand DNA to double-stranded oligonucleotides that contained 5[prime]-overhang, 3[prime]-overhang, or blunt ends. Single-stranded, nicked, or gapped oligonucleotides could also be used as acceptor molecules. These results demonstrate that the type II enzyme has an intrinsic ability to mediate illegitimate DNA recombination in vitro and suggests possible roles for topoisomerase II in nucleic acid recombination in vivo.

  3. Photo-activated biological processes as quantum measurements

    CERN Document Server

    Imamoglu, Atac

    2014-01-01

    We outline a framework for describing photo-activated biological reactions as generalized quantum measurements of external fields, for which the biological system takes on the role of a quantum meter. By using general arguments regarding the Hamiltonian that describes the measurement interaction, we identify the cases where it is essential for a complex chemical or biological system to exhibit non-equilibrium quantum coherent dynamics in order to achieve the requisite functionality. We illustrate the analysis by considering measurement of the solar radiation field in photosynthesis and measurement of the earth's magnetic field in avian magnetoreception.

  4. Transcriptionally active chromatin recruits homologous recombination at DNA double-strand breaks.

    Science.gov (United States)

    Aymard, François; Bugler, Beatrix; Schmidt, Christine K; Guillou, Emmanuelle; Caron, Pierre; Briois, Sébastien; Iacovoni, Jason S; Daburon, Virginie; Miller, Kyle M; Jackson, Stephen P; Legube, Gaëlle

    2014-04-01

    Although both homologous recombination (HR) and nonhomologous end joining can repair DNA double-strand breaks (DSBs), the mechanisms by which one of these pathways is chosen over the other remain unclear. Here we show that transcriptionally active chromatin is preferentially repaired by HR. Using chromatin immunoprecipitation-sequencing (ChIP-seq) to analyze repair of multiple DSBs induced throughout the human genome, we identify an HR-prone subset of DSBs that recruit the HR protein RAD51, undergo resection and rely on RAD51 for efficient repair. These DSBs are located in actively transcribed genes and are targeted to HR repair via the transcription elongation-associated mark trimethylated histone H3 K36. Concordantly, depletion of SETD2, the main H3 K36 trimethyltransferase, severely impedes HR at such DSBs. Our study thereby demonstrates a primary role in DSB repair of the chromatin context in which a break occurs.

  5. Activity and stability of recombinant human superoxide dismutase in buffer solutions and hypothermic perfusates.

    Directory of Open Access Journals (Sweden)

    Senoo,Yoshimasa

    1988-06-01

    Full Text Available The stability of recombinant human superoxide dismutase (r-hSOD in buffer solutions was studied in solutions at various pH and temperatures. Additionally, we studied the effects of incubation with proteases, serum and two types of hypothermic perfusates. R-hSOD was stable in the pH range of 6-11 and at temperatures up to 80 degrees C for 30 min. R-hSOD activity was not affected by incubation with trypsin, aminopeptidase M or serum for 2 h. R-hSOD activity determined at various temperatures (4-37 degrees C did not vary remarkably. R-hSOD in hypothermic perfusates was stable at 4-37 degrees C for 24 h.

  6. Biological activities of radiation-degraded carrageenan

    Energy Technology Data Exchange (ETDEWEB)

    Relleve, Lorna; Dela Rosa, Alumanda; ABAD, Lucille; Aranilla, Charito; Aliganga, Anne Kathrina [Philippine Nuclear Research Institute, Quezon City (Philippines); Yoshii, Fumio; Kume, Tamikazu; Nagasawa, Naotsugu [Japan Atomic Energy Research Inst., Takasaki, Gunma (Japan). Takasaki Radiation Chemistry Research Establishment

    2001-03-01

    Carrageenans were irradiated in solid state to doses 50-1000 kGy in air at ambient temperature. Changes in their molecular weight and functional properties with respect to their FT-IR and UV spectra were evaluated. Irradiation of carrageenans resulted in a rapid decrease of molecular weight indicating main chain scission in their polymeric structures. Formations of some compounds were evident by new absorption peaks in their UV and FT-IR spectra and quantitative analyses of the FT-IR spectra which, in addition, support that there is a breakdown in the carrageenan structure. Irradiated carrageenans were investigated for their plant growth-promoting activity. Carrageenans were added to the nutrient solutions for rice seedlings under non-circulating hydroponics cultivation. Irradiated carrageenan induced weight gain in treated rice seedlings. Maximum weight gain was obtained with KC irradiated at 100 kGy while treatment with IC at 500 kGy. IC exhibited less growth promoting properties than KC. The growth of fungi on the roots disappeared with treatment of IC and KC irradiated at 500 kGy. Growth promotion of some leafy vegetables was also observed with application of degraded KC. The carrageenan molecule has been broken down to smaller molecule (s) or compound (s) that can be absorbed effectively as nourishment factors and anti-microbial agents by plants. (author)

  7. Recombinant human diamine oxidase activity is not inhibited by ethanol, acetaldehyde, disulfiram, diethyldithiocarbamate or cyanamide.

    Science.gov (United States)

    Bartko, Johann; Gludovacz, Elisabeth; Petroczi, Karin; Borth, Nicole; Jilma, Bernd; Boehm, Thomas

    2016-08-01

    Human diamine oxidase (hDAO, EC 1.4.3.22) is the key enzyme in the degradation of extracellular histamine. Consumption of alcohol is a known trigger of mast cell degranulation in patients with mast cell activation syndrome. Ethanol may also interfere with enzymatic histamine degradation, but reports on the effects on DAO activity are controversial. There are also conflicting reports whether disulfiram, an FDA-approved agent in the treatment of alcohol dependence, inhibits DAO. We therefore investigated the inhibitory potential of ethanol and disulfiram and their metabolites on recombinant human DAO (rhDAO) in three different assay systems. Relevant concentrations of ethanol, acetaldehyde, and acetate did not inhibit rhDAO activity in an in vitro assay system using horseradish peroxidase (HRP) -mediated luminol oxidation. The aldehyde dehydrogenase (ALDH; EC 1.2.1.3) inhibitors cyanamide and its dimer dicyanamide also had no effect on DAO activity. In one assay system, the irreversible ALDH inhibitor disulfiram and its main metabolite diethyldithiocarbamate seemed to inhibit DAO activity. However, the decreased product formation was not due to a direct block of DAO activity but resulted from inhibition of peroxidase employed in the coupled system. Our in vitro data do not support a direct blocking effect of ethanol, disulfiram, and their metabolites on DAO activity in vivo.

  8. Correlation between the glycan variations and defibrinogenating activities of acutobin and its recombinant glycoforms.

    Directory of Open Access Journals (Sweden)

    Ying-Ming Wang

    Full Text Available Acutobin isolated from Deinagkistrodon acutus venom has been used to prevent or treat stroke in patients. This defibrinogenating serine protease is a 39 kDa glycoprotein containing terminal disialyl-capped N-glycans. After sialidase treatment, the enzyme showed similar catalytic activities toward chromogenic substrate, and cleaved the Aα chain of fibrinogen as efficiently as the native acutobin did. However, the level of fibrinogen degradation products in mice after i.p.-injection of desialylated-acutobin was significantly lower than the level after acutobin injection, suggesting that the disialyl moieties may improve or prolong the half-life of acutobin. Two recombinant enzymes with identical protein structures and similar amidolytic activities to those of native acutobin were expressed from HEK293T and SW1353 cells and designated as HKATB and SWATB, respectively. Mass spectrometric profiling showed that their glycans differed from those of acutobin. In contrast to acutobin, HKATB cleaved not only the Aα chain but also the Bβ and γ chains of human fibrinogens, while SWATB showed a reduced α-fibrinogenase activity. Non-denaturing deglycosylation of these proteases by peptide N-glycosidase F significantly reduced their fibrinogenolytic activities and thermal stabilities. The in vivo defibrinogenating effect of HKATB was inferior to that of acutobin in mice. Taken together, our results suggest that the conjugated glycans of acutobin are involved in its interaction with fibrinogen, and that the selection of cells optimally expressing efficient glycoforms and further glycosylation engineering are desirable before a recombinant product can replace the native enzyme for clinical use.

  9. Drosophila brca2 is required for mitotic and meiotic DNA repair and efficient activation of the meiotic recombination checkpoint.

    Directory of Open Access Journals (Sweden)

    Martha Klovstad

    2008-02-01

    Full Text Available Heterozygous mutations in the tumor suppressor BRCA2 confer a high risk of breast and other cancers in humans. BRCA2 maintains genome stability in part through the regulation of Rad51-dependent homologous recombination. Much about its precise function in the DNA damage responses is, however, not yet known. We have made null mutations in the Drosophila homolog of BRCA2 and measured the levels of homologous recombination, non-homologous end-joining, and single-strand annealing in the pre-meiotic germline of Drosophila males. We show that repair by homologous recombination is dramatically decreased in Drosophila brca2 mutants. Instead, large flanking deletions are formed, and repair by the non-conservative single-strand annealing pathway predominates. We further show that during meiosis, Drosophila Brca2 has a dual role in the repair of meiotic double-stranded breaks and the efficient activation of the meiotic recombination checkpoint. The eggshell patterning defects that result from activation of the meiotic recombination checkpoint in other meiotic DNA repair mutants can be strongly suppressed by mutations in brca2. In addition, Brca2 co-immunoprecipitates with the checkpoint protein Rad9, suggesting a direct role for Brca2 in the transduction of the meiotic recombination checkpoint signal.

  10. Perlecan domain 1 recombinant proteoglycan augments BMP-2 activity and osteogenesis

    Directory of Open Access Journals (Sweden)

    DeCarlo Arthur A

    2012-09-01

    Full Text Available Abstract Background Many growth factors, such as bone morphogenetic protein (BMP-2, have been shown to interact with polymers of sulfated disacharrides known as heparan sulfate (HS glycosaminoglycans (GAGs, which are found on matrix and cell-surface proteoglycans throughout the body. HS GAGs, and some more highly sulfated forms of chondroitin sulfate (CS, regulate cell function by serving as co-factors, or co-receptors, in GF interactions with their receptors, and HS or CS GAGs have been shown to be necessary for inducing signaling and GF activity, even in the osteogenic lineage. Unlike recombinant proteins, however, HS and CS GAGs are quite heterogenous due, in large part, to post-translational addition, then removal, of sulfate groups to various positions along the GAG polymer. We have, therefore, investigated whether it would be feasible to deliver a DNA pro-drug to generate a soluble HS/CS proteoglycan in situ that would augment the activity of growth-factors, including BMP-2, in vivo. Results Utilizing a purified recombinant human perlecan domain 1 (rhPln.D1 expressed from HEK 293 cells with HS and CS GAGs, tight binding and dose-enhancement of rhBMP-2 activity was demonstrated in vitro. In vitro, the expressed rhPln.D1 was characterized by modification with sulfated HS and CS GAGs. Dose-enhancement of rhBMP-2 by a pln.D1 expression plasmid delivered together as a lyophilized single-phase on a particulate tricalcium phosphate scaffold for 6 or more weeks generated up to 9 fold more bone volume de novo on the maxillary ridge in a rat model than in control sites without the pln.D1 plasmid. Using a significantly lower BMP-2 dose, this combination provided more than 5 times as much maxillary ridge augmentation and greater density than rhBMP-2 delivered on a collagen sponge (InFuse™. Conclusions A recombinant HS/CS PG interacted strongly and functionally with BMP-2 in binding and cell-based assays, and, in vivo, the pln.247 expression plasmid

  11. Biological activity of diterpenoids isolated from Anatolian Lamiaceae Plants

    Directory of Open Access Journals (Sweden)

    Gülaçtı Topçu

    2007-05-01

    Full Text Available In this study, antibacterial, antifungal, antimycobacterial, cytotoxic, antitumor, cardiovascular, antifeedant, insecticidal, antileishmanial and some other single activities of diterpenoids and norditerpenoids isolated from Turkish Lamiaceae plants, are reviewed. The diterpenoids were isolated from species of Salvia, Sideritis, and Ballota species growing in Anatolia. Fifty abietanes, ten kaurenes, seven pimaranes, six labdanes with their biological activities were reported. While twenty five diterpenoids showed antibacterial activity, eight of which showed activity against fungi. The most cytotoxic one was found to be taxodione (44 isolated from species of Salvia. Antifeedant, insecticidal and insect repellent activity of kaurenes, antimycobacterial activity and cardioactivity of abietanes and norabietanes together with labdanes were also reported.

  12. Residual endotoxin contaminations in recombinant proteins are sufficient to activate human CD1c+ dendritic cells.

    Science.gov (United States)

    Schwarz, Harald; Schmittner, Maria; Duschl, Albert; Horejs-Hoeck, Jutta

    2014-01-01

    Many commercially available recombinant proteins are produced in Escherichia coli, and most suppliers guarantee contamination levels of less than 1 endotoxin unit (EU). When we analysed commercially available proteins for their endotoxin content, we found contamination levels in the same range as generally stated in the data sheets, but also some that were higher. To analyse whether these low levels of contamination have an effect on immune cells, we stimulated the monocytic cell line THP-1, primary human monocytes, in vitro differentiated human monocyte-derived dendritic cells, and primary human CD1c+ dendritic cells (DCs) with very low concentrations of lipopolysaccharide (LPS; ranging from 0.002-2 ng/ml). We show that CD1c+ DCs especially can be activated by minimal amounts of LPS, equivalent to the levels of endotoxin contamination we detected in some commercially available proteins. Notably, the enhanced endotoxin sensitivity of CD1c+ DCs was closely correlated with high CD14 expression levels observed in CD1c+ DCs that had been maintained in cell culture medium for 24 hours. When working with cells that are particularly sensitive to LPS, even low endotoxin contamination may generate erroneous data. We therefore recommend that recombinant proteins be thoroughly screened for endotoxin contamination using the limulus amebocyte lysate test, fluorescence-based assays, or a luciferase based NF-κB reporter assay involving highly LPS-sensitive cells overexpressing TLR4, MD-2 and CD14.

  13. Recombinant Expression and Biological Property Analysis of Goose Avian β-Defensin 7 Protein%重组鹅β-防御素7蛋白的原核表达及其生物学特性分析

    Institute of Scientific and Technical Information of China (English)

    高梦莹; 徐倩倩; 张婷婷; 陈玉秋; 赵文钧; 齐甜铭; 马得莹

    2016-01-01

    The objective of the study was to investigate the biological property of goose avian β-defensin ( AvBD) 7. The gene of AvBD7 was cloned into EcoRⅠ and XhoⅠ sites of pProEX HTa vector to construct recombinant plasmid pProEX-AvBD7. The recombinant plasmid was transformed into Escherichia coli Rosetta and the bacteria was induced with isopropy-β-D-thiogalactoside ( IPTG) . It was analyzed by tricine sodium do-decyl sulfate polyacrylamide gel electrophoresis ( Tricine-SDS-PAGE) that the recombinant protein size was 10 to 15 ku, which was the same as the expected size. Antimicrobial activity of the recombinant fusion protein a-gainst Escherichia coli, Salmonella pullorum, Staphylococcus aureus, Micrococcus tetragenus and Bacillus subtilis was measured in vitro. In addition, the effect of ionic strength on the antibacterial activity and hemolytic activity of the recombinant fusion protein were investigated. The results showed that the recombinant goose AvBD7 protein exhibited significant antimicrobial activity against the five bacteria investigated ( P0.05). In conclusion, recombinant goose AvBD7 protein exhibits extensive antimicrobial activity. High salt concentration significantly decrease its antibacterial activity. In addition, the recombinant goose AvBD7 protein shows little hemolytic activity.%为探讨鹅β-防御素7( AvBD7)的生物学特性,将鹅AvBD7基因亚克隆到大肠杆菌原核表达载体pProEX HTa的EcoRⅠ和XhoⅠ双酶切位点上,构建重组表达质粒pProEX-AvBD7,将重组质粒转化到大肠杆菌Rosetta感受态细胞中,用异丙基-β-D-硫代半乳糖苷( IPTG)对其菌液进行诱导表达。经 N-三(羟甲基)甲基甘氨酸十二烷基硫酸钠聚丙烯酰胺凝胶电泳( Tricine-SDS-PAGE)分析,该重组蛋白大小为10~15 ku,与预期大小结果一致。重组鹅AvBD7蛋白纯化后,通过菌落计数的方法测定其体外抗大肠杆菌、鸡白痢沙门氏菌、金黄色葡萄球菌、四联球菌、枯草芽孢杆

  14. Biological Activity of Curcuminoids Isolated from Curcuma longa

    Directory of Open Access Journals (Sweden)

    Simay Çıkrıkçı

    2008-04-01

    Full Text Available Curcumin is the most important fraction of turmeric which is responsible for its biological activity. In this study, isolation and biological assessment of turmeric and curcumin have been discussed against standard bacterial and mycobacterial strains such as E.coli , S.aureus, E.feacalis, P.aeuroginosa, M.smegmatis, M.simiae, M.kansasii, M. terrae, M.szulgai and the fungi Candida albicans. The antioxidant activity of curcumin and turmeric were also determined by the CUPRAC method.

  15. Biological Activity of Aminophosphonic Acids and Their Short Peptides

    Science.gov (United States)

    Lejczak, Barbara; Kafarski, Pawel

    The biological activity and natural occurrence of the aminophosphonic acids were described half a century ago. Since then the chemistry and biology of this class of compounds have developed into the separate field of phosphorus chemistry. Today it is well acknowledged that these compounds possess a wide variety of promising, and in some cases commercially useful, physiological activities. Thus, they have found applications ranging from agrochemical (with the herbicides glyphosate and bialaphos being the most prominent examples) to medicinal (with the potent antihypertensive fosinopril and antiosteoporetic bisphosphonates being examples).

  16. Structurally unique recombinant Kazal-type proteinase inhibitor retains activity when terminally extended and glycosylated.

    Science.gov (United States)

    Kludkiewicz, Barbara; Kodrík, Dalibor; Grzelak, Krystyna; Nirmala, Xavier; Sehnal, Frantisek

    2005-10-01

    Recombinant derivatives of the Kazal-type serine proteinase inhibitor GmSPI2 (36 amino acid residues), which is a component of insect silk, were prepared in the expression vector Pichia pastoris. The rhSPI2 had a C-terminal hexahistidine tag attached to the GmSPI2 sequence, rtSPI2 was extended with GluAlaAla at the N-terminus, and rfSPI2 included this N-terminal extension and a C-terminal tail of 22 residues (myc epitope and hexahistidine). A portion of the secreted rfSI2 was O-glycosylated with a trimannosyl or hexamannosyl. The native inhibitor was active slightly on trypsin and highly on subtilisin and proteinase K. The extended C-terminus in rhSPI2 and rfSPI2 enhanced activity on the two latter enzymes and rendered rfSPI2 active on elastase and pronase, but abolished the inhibition of trypsin. The glycosylation of rfSPI2 reduced its inhibitory activity to a level comparable with the native inhibitor. The rtSPI2 with tripeptide extension at the N-terminus and no C-terminal modification was clearly less active than the native inhibitor. None of the tested compounds inhibited alpha-chymotrypsin and the non-serine proteinases.

  17. Sustained in vivo activity of recombinant bovine granulocyte colony stimulating factor (rbG-CSF) using HEPES buffer.

    Science.gov (United States)

    Kasraian, K; Kuzniar, A; Earley, D; Kamicker, B J; Wilson, G; Manion, T; Hong, J; Reiber, C; Canning, P

    2001-08-01

    The purpose of this study was to develop a long-acting injectable formulation of bG-CSF for veterinary use. However, in order to achieve sustained in vivo activity it was first necessary to stabilize the protein at the injection site. Preformulation studies, as well as literature, suggest that bG-CSF aggregates at neutral pH ranges (i.e., pH 6-8) and at temperatures of approximately 40 degrees C. Therefore, bG-CSF will not retain its activity for an extended period of time at the injection site. During this study we determined that HEPES buffer has a very significant impact on protein stability as well as on biological performance. Recombinant bovine granulocyte colony stimulating factor (rbG-CSF) was formulated in 1 M HEPES buffer for subcutaneous injection into cows. bG-CSF formulated in 1 M HEPES buffer resulted in sustained in vivo activity of bG-CSF compared to the "control" formulation (control formulation: 5% mannitol, 10 mM acetate buffer, 0.004% tween-80, pH 4). White blood cell (WBC) count was used as a marker to evaluate in vivo activity of the formulation. WBC numbers remained above a threshold value for only 24-30 h for the control formula. However, when bG-CSF was formulated in 1 M HEPES, the WBC remained above threshold for 3 days or 72 h. Formulating bG-CSF in 1 M HEPES at pH 7.5 also resulted in greater solution stability. This was surprising since bG-CSF is intrinsically not stable at neutral pH. The effect of 1 M HEPES on the T(M) (temperature at maximum heat flow on calorimetry scan) of bG-CSF was determined by microcalorimetry. In the absence of 1 M HEPES buffer the T(M) was 48 degrees C (onset approximately 40 degrees C), while bG-CSF formulated in 1 M HEPES buffer has a T(M) of 59 degrees C (onset approximately 50 degrees C). Similar organic buffers, such as MOPS, HEPPS, TES, and tricine, also resulted in improved solution stability as well as in sustained in vivo activity. The dramatic effect of these buffers on stability and biological

  18. Recent progress in biological activities of synthesized phenothiazines.

    Science.gov (United States)

    Pluta, Krystian; Morak-Młodawska, Beata; Jeleń, Małgorzata

    2011-08-01

    This review summarizes recent medicinal chemistry investigations in vitro and in vivo in search for new phenothiazines of promising biological activities. New phenothiazine derivatives (over 50 main structures) contain dialkylaminoalkyl, cycloaminoalkyl and aminoalkyl substituents and their acyl and sulfonyl derivatives, and other substituents with varied the monocyclic (pyrazole, thiazole, oxadiazole, thiadiazole, tetrazole) and bicyclic (quinolizine, pyrazolopyrimidine, thiazolopyridine, azabicyclononane and spiro[chromanpyrimidine] heterocycles linked directly or via the alkyl chain with the thiazine nitrogen atom or with the benzene ring. The modifications of the tricyclic ring system with the bicyclic homoaromatic ring (naphthalene) and monocyclic and bicyclic azine rings (pyridine, pyrimidine, pyrazine and quinoline) led to compounds of significant biological activities. Recently obtained phenothiazines exhibit promising antibacterial, antifungal, anticancer, antiviral, anti-inflammatory, antimalarial, antifilarial, trypanocidal, anticonvulsant, analgesic, immunosuppressive and multidrug resistance reversal properties. These activities were the results of the actions of phenothiazines on biological systems via the interaction of the pharmacophoric substituent (in some cases of strict length), via the interaction of the multicyclic ring system (π-π interaction, intercalation in DNA) and via the lipophilic character allowing the penetration through the biological membranes. The activities were examined by using various biological systems such as cell lines, bacteria, viruses, parasites, laboratory mice, rats and rabbits, and monolayer and bilayer membranes. Some mechanisms of the actions are discussed. This review shows current tendency in the phenothiazine synthesis (without synthetic routes) and reveals the phenothiazine core to be very potent pharmacophoric moiety which can be a rich source of new compounds having desirable biological activities.

  19. Failure of Recombinant Activated Factor VII in Treatment of Diffuse Alveolar Hemorrhage due to Cryoglobulinemic Vasculitis

    Directory of Open Access Journals (Sweden)

    Dania Khoulani

    2014-01-01

    Full Text Available Diffuse alveolar hemorrhage (DAH is a serious complication of the small vessel vasculitis syndromes and carries a high mortality. Recombinant activated factor VII (rFVIIa is used to treat bleeding in patients with hemophilia and antibodies to factor VIII or IX. It is increasingly being used in life-threatening hemorrhage in a variety of other settings in which conventional therapy is unsuccessful. Randomized controlled trials of rFVIIa in DAH are lacking. However, several case reports have described a complete or sustained control of DAH using rFVIIa after patients failed to respond to medical treatment. There are no case reports in the literature describing the use or the failure of rFVIIa in DAH associated with cryoglobulinemic vasculitis. We here report the failure of rFVIIa to control DAH in a patient with CD5+ B-cell non-Hodgkin’s lymphoma and cryoglobulinemic vasculitis.

  20. Tankyrases Promote Homologous Recombination and Check Point Activation in Response to DSBs

    Science.gov (United States)

    Furst, Audrey; Koch, Marc; Fischer, Benoit; Soutoglou, Evi

    2016-01-01

    DNA lesions are sensed by a network of proteins that trigger the DNA damage response (DDR), a signaling cascade that acts to delay cell cycle progression and initiate DNA repair. The Mediator of DNA damage Checkpoint protein 1 (MDC1) is essential for spreading of the DDR signaling on chromatin surrounding Double Strand Breaks (DSBs) by acting as a scaffold for PI3K kinases and for ubiquitin ligases. MDC1 also plays a role both in Non-Homologous End Joining (NHEJ) and Homologous Recombination (HR) repair pathways. Here we identify two novel binding partners of MDC1, the poly (ADP-ribose) Polymerases (PARPs) TNKS1 and 2. We find that TNKSs are recruited to DNA lesions by MDC1 and regulate DNA end resection and BRCA1A complex stabilization at lesions leading to efficient DSB repair by HR and proper checkpoint activation. PMID:26845027

  1. Tankyrases Promote Homologous Recombination and Check Point Activation in Response to DSBs.

    Directory of Open Access Journals (Sweden)

    Zita Nagy

    2016-02-01

    Full Text Available DNA lesions are sensed by a network of proteins that trigger the DNA damage response (DDR, a signaling cascade that acts to delay cell cycle progression and initiate DNA repair. The Mediator of DNA damage Checkpoint protein 1 (MDC1 is essential for spreading of the DDR signaling on chromatin surrounding Double Strand Breaks (DSBs by acting as a scaffold for PI3K kinases and for ubiquitin ligases. MDC1 also plays a role both in Non-Homologous End Joining (NHEJ and Homologous Recombination (HR repair pathways. Here we identify two novel binding partners of MDC1, the poly (ADP-ribose Polymerases (PARPs TNKS1 and 2. We find that TNKSs are recruited to DNA lesions by MDC1 and regulate DNA end resection and BRCA1A complex stabilization at lesions leading to efficient DSB repair by HR and proper checkpoint activation.

  2. Annealing effects on recombinative activity of nickel at direct silicon bonded interface

    Energy Technology Data Exchange (ETDEWEB)

    Kojima, Takuto, E-mail: tkojima@toyota-ti.ac.jp; Ohshita, Yoshio; Yamaguchi, Masafumi [Toyota Technological Institute, 2-12-1 Hisakata, Tempaku-ku, Nagoya, 468-8511 (Japan)

    2015-09-15

    By performing capacitance transient analyses, the recombination activity at a (110)/(100) direct silicon bonded (DSB) interface contaminated with nickel diffused at different temperatures, as a model of grain boundaries in multicrystalline silicon, was studied. The trap level depth from the valence band, trap density of states, and hole capture cross section peaked at an annealing temperature of 300 °C. At temperatures ⩾400 °C, the hole capture cross section increased with temperature, but the density of states remained unchanged. Further, synchrotron-based X-ray analyses, microprobe X-ray fluorescence (μ-XRF), and X-ray absorption near edge structure (XANES) analyses were performed. The analysis results indicated that the chemical phase after the sample was annealed at 200 °C was a mixture of NiO and NiSi{sub 2}.

  3. Gemini ester quat surfactants and their biological activity.

    Science.gov (United States)

    Łuczyński, Jacek; Frąckowiak, Renata; Włoch, Aleksandra; Kleszczyńska, Halina; Witek, Stanisław

    2013-03-01

    Cationic gemini surfactants are an important class of surface-active compounds that exhibit much higher surface activity than their monomeric counterparts. This type of compound architecture lends itself to the compound being easily adsorbed at interfaces and interacting with the cellular membranes of microorganisms. Conventional cationic surfactants have high chemical stability but poor chemical and biological degradability. One of the main approaches to the design of readily biodegradable and environmentally friendly surfactants involves inserting a bond with limited stability into the surfactant molecule to give a cleavable surfactant. The best-known example of such a compound is the family of ester quats, which are cationic surfactants with a labile ester bond inserted into the molecule. As part of this study, a series of gemini ester quat surfactants were synthesized and assayed for their biological activity. Their hemolytic activity and changes in the fluidity and packing order of the lipid polar heads were used as the measures of their biological activity. A clear correlation between the hemolytic activity of the tested compounds and their alkyl chain length was established. It was found that the compounds with a long hydrocarbon chain showed higher activity. Moreover, the compounds with greater spacing between their alkyl chains were more active. This proves that they incorporate more easily into the lipid bilayer of the erythrocyte membrane and affect its properties to a greater extent. A better understanding of the process of cell lysis by surfactants and of their biological activity may assist in developing surfactants with enhanced selectivity and in widening their range of application.

  4. Characterization of the peptidase activity of recombinant porcine pregnancy-associated glycoprotein-2.

    Science.gov (United States)

    Telugu, Bhanu Prakash V L; Green, Jonathan A

    2008-12-01

    The pregnancy-associated glycoproteins (PAGs) belong to the aspartic peptidase family. They are expressed exclusively in trophoblasts of even-toed ungulates such as swine, cattle, sheep, etc. In pigs, two distinct PAG transcripts (and some variants) have been described. One of the transcripts, porcine PAG-1 (poPAG-1) may not be capable of acting as a peptidase. The second transcript, poPAG-2, possesses a conserved catalytic centre and has been predicted, but not shown, to have proteolytic activity. The thrust of this work was to test such a possibility. PoPAG-2 was expressed as a recombinant protein with an amino-terminal 'FLAG-tag' in a Baculoviral expression system. The expressed proteins were affinity purified by using an anti-FLAG antibody. The purified preparations were then analysed for proteolytic activity against a fluorescent substrate. Porcine PAG-2 had optimal proteolytic activity around pH 3.5. Against this substrate, it had a k(cat)/K(m) of 1.2 microM(-1) s(-1) and was inhibited by the aspartic peptidase inhibitor, pepstatin A, with a K(i) of 12.5 nM. Since the proteolytic activity of PAGs in the pig has now been established, the search for putative substrates to gain insight into the physiological role of PAGs will likely be the focus of future investigations.

  5. Bimolecular recombination reactions: K-adiabatic and K-active forms of the bimolecular master equations and analytic solutions

    Science.gov (United States)

    Ghaderi, Nima

    2016-03-01

    Expressions for a K-adiabatic master equation for a bimolecular recombination rate constant krec are derived for a bimolecular reaction forming a complex with a single well or complexes with multiple well, where K is the component of the total angular momentum along the axis of least moment of inertia of the recombination product. The K-active master equation is also considered. The exact analytic solutions, i.e., the K-adiabatic and K-active steady-state population distribution function of reactive complexes, g(EJK) and g(EJ), respectively, are derived for the K-adiabatic and K-active master equation cases using properties of inhomogeneous integral equations (Fredholm type). The solutions accommodate arbitrary intermolecular energy transfer models, e.g., the single exponential, double exponential, Gaussian, step-ladder, and near-singularity models. At the high pressure limit, the krec for both the K-adiabatic and K-active master equations reduce, respectively, to the K-adiabatic and K-active bimolecular Rice-Ramsperger-Kassel-Marcus theory (high pressure limit expressions). Ozone and its formation from O + O2 are known to exhibit an adiabatic K. The ratio of the K-adiabatic to the K-active recombination rate constants for ozone formation at the high pressure limit is calculated to be ˜0.9 at 300 K. Results on the temperature and pressure dependence of the recombination rate constants and populations of O3 will be presented elsewhere.

  6. Bimolecular recombination reactions: K-adiabatic and K-active forms of the bimolecular master equations and analytic solutions.

    Science.gov (United States)

    Ghaderi, Nima

    2016-03-28

    Expressions for a K-adiabatic master equation for a bimolecular recombination rate constant krec are derived for a bimolecular reaction forming a complex with a single well or complexes with multiple well, where K is the component of the total angular momentum along the axis of least moment of inertia of the recombination product. The K-active master equation is also considered. The exact analytic solutions, i.e., the K-adiabatic and K-active steady-state population distribution function of reactive complexes, g(EJK) and g(EJ), respectively, are derived for the K-adiabatic and K-active master equation cases using properties of inhomogeneous integral equations (Fredholm type). The solutions accommodate arbitrary intermolecular energy transfer models, e.g., the single exponential, double exponential, Gaussian, step-ladder, and near-singularity models. At the high pressure limit, the krec for both the K-adiabatic and K-active master equations reduce, respectively, to the K-adiabatic and K-active bimolecular Rice-Ramsperger-Kassel-Marcus theory (high pressure limit expressions). Ozone and its formation from O + O2 are known to exhibit an adiabatic K. The ratio of the K-adiabatic to the K-active recombination rate constants for ozone formation at the high pressure limit is calculated to be ∼0.9 at 300 K. Results on the temperature and pressure dependence of the recombination rate constants and populations of O3 will be presented elsewhere.

  7. TRIAZOLE: A POTENTIAL BIOACTIVE AGENT (SYNTHESIS AND BIOLOGICAL ACTIVITY)

    OpenAIRE

    Pandeya Surendra Nath; Pathak Ashish; Mishra Rupesh

    2011-01-01

    Azoles belong to very important class of Antimicrobial drugs. Triazole is very important Azole which exists in two isomeric forms namely 1, 2, 3-Triazole and 1, 2, 4-Triazole. This Review Article covers the Different approaches to synthesize Triazoles having different substitution and their different biological activity. This Review article can be useful to synthesize new compounds having Triazole nucleolus.

  8. Biologically active substances produced by antarctic cryptoendolithic fungi.

    Science.gov (United States)

    Ocampo-Friedmann, R; Friedmann, E I

    1993-01-01

    Researchers report results of laboratory studies of over 200 microbial strains of fungi, algae, cyanobacteria, and heterotrophic bacteria collected in the Ross Desert region of Antarctica. All of the 35 fungal strains produced substances that inhibited the growth of cyanobacteria and algae. The inhibitory effect of the biologically active substance was evident in crushed cell extract but less in spent broth.

  9. Aspartate and glutamate mimetic structures in biologically active compounds.

    Science.gov (United States)

    Stefanic, Peter; Dolenc, Marija Sollner

    2004-04-01

    Glutamate and aspartate are frequently recognized as key structural elements for the biological activity of natural peptides and synthetic compounds. The acidic side-chain functionality of both the amino acids provides the basis for the ionic interaction and subsequent molecular recognition by specific receptor sites that results in the regulation of physiological or pathophysiological processes in the organism. In the development of new biologically active compounds that possess the ability to modulate these processes, compounds offering the same type of interactions are being designed. Thus, using a peptidomimetic design approach, glutamate and aspartate mimetics are incorporated into the structure of final biologically active compounds. This review covers different bioisosteric replacements of carboxylic acid alone, as well as mimetics of the whole amino acid structure. Amino acid analogs presented include those with different distances between anionic moieties, and analogs with additional functional groups that result in conformational restriction or alternative interaction sites. The article also provides an overview of different cyclic structures, including various cycloalkane, bicyclic and heterocyclic analogs, that lead to conformational restriction. Higher di- and tripeptide mimetics in which carboxylic acid functionality is incorporated into larger molecules are also reviewed. In addition to the mimetic structures presented, emphasis in this article is placed on their steric and electronic properties. These mimetics constitute a useful pool of fragments in the design of new biologically active compounds, particularly in the field of RGD mimetics and excitatory amino acid agonists and antagonists.

  10. Polysaccharies of higher fungi: Biological role, structure and antioxidative activity

    NARCIS (Netherlands)

    Kozarski, M.S.; Klaus, A.; Niksic, M.; Griensven, van L.J.L.D.; Vrvic, M.M.; Jakovljevic, D.M.

    2014-01-01

    The fungal polysaccharides attract a lot of attention due to their multiple challenging bio-logical properties, such as: anti-tumor, anti-viral, anticomplementary, anticoagulant, hypo-lipidemic, immunomodulatory and immune-stimulatory activities, which all together make them suitable for application

  11. Solar Energy Education. Renewable energy activities for biology

    Energy Technology Data Exchange (ETDEWEB)

    1982-01-01

    An instructional aid for teachers is presented that will allow biology students the opportunity to learn about renewable energy sources. Some of the school activities include using leaves as collectors of solar energy, solar energy stored in wood, and a fuel value test for green and dry woods. A study of organic wastes as a source of fuel is included. (BCS)

  12. Occurrence, biological activity and synthesis of drimane sesquiterpenoids

    NARCIS (Netherlands)

    Jansen, B.J.M.; Groot, de Æ.

    2004-01-01

    In this review the names, structures and occurrence of all new drimanes and rearranged drimanes, which have been published between January 1990 and January 2003 have been collected. Subjects that have been treated are biosynthesis, analysis, biological activities, with special attention to cytotoxic

  13. Vaccinia virus recombinants expressing an 11-kilodalton beta-galactosidase fusion protein incorporate active beta-galactosidase in virus particles.

    Science.gov (United States)

    Huang, C; Samsonoff, W A; Grzelecki, A

    1988-10-01

    Recombinant plasmids in which vaccinia virus transcriptional regulatory sequences were fused to the Escherichia coli lacZ gene were constructed for insertion of the lacZ gene into the vaccinia virus genome. beta-Galactosidase (beta-gal) was found in some purified recombinant vaccinia virions. By enzyme activity, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and microscopic techniques, the evidence suggested that beta-gal accounted for 5% of the total protein in the virion. These recombinant viruses were constructed so that a portion of the coding sequences of a late vaccinia virus structural polypeptide was fused to the amino terminus of beta-gal to produce the fusion protein. Removal of the coding sequences resulted in the complete loss of beta-gal activity. This demonstrated that a vaccinia virus DNA segment from a late structural gene is responsible for the incorporation of beta-gal into the virion.

  14. Biologically Active Substances of Plants from Sali x L. Genus

    OpenAIRE

    O. O. Frolova; E. V. Kompantseva; T. M. Dementieva

    2016-01-01

    The review systematizes data about chemical composition of bark, leaves, inflorescences, and sprouts of different species of Salix L. The closest attention is payed to investigations of Salix, which has been recently carried out in our country and abroad. For every group of biologically active substances described in Salix there are data about suppressed types of pharmacological activity.

  15. Biologically Active Substances of Plants from Sali x L. Genus

    Directory of Open Access Journals (Sweden)

    O. O. Frolova

    2016-01-01

    Full Text Available The review systematizes data about chemical composition of bark, leaves, inflorescences, and sprouts of different species of Salix L. The closest attention is payed to investigations of Salix, which has been recently carried out in our country and abroad. For every group of biologically active substances described in Salix there are data about suppressed types of pharmacological activity.

  16. Chemical composition and biological activity of the plum seed extract

    OpenAIRE

    Savić, Ivan M.; Nikolić, Vesna D.; Savić-Gajić, Ivana M.; Kundaković, Tatjana D.; Stanojković, Tatjana P.; Najman, Stevo J.; id_orcid 0000-0002-2411-9802

    2016-01-01

    The aim of this paper was to estimate the biological activity of the plum seed extract and to define the chemical composition by using the ESI-MS method. During the investigation of the antioxidant activity, the extract showed a better ability to inhibit DPPH radicals compared with amygdalin standard. The results of the antimicrobial study indicate that the extract has a greater effect on Gram-negative bacteria compared with amygdalin. Gram-positive bacteria and fungi remained resistant in bo...

  17. Secondary Metabolites from Inula britannica L. and Their Biological Activities

    Directory of Open Access Journals (Sweden)

    Yoon-Ha Kim

    2010-03-01

    Full Text Available Inula britannica L., family Asteraceae, is used in traditional Chinese and Kampo Medicines for various diseases. Flowers or the aerial parts are a rich source of secondary metabolites. These consist mainly of terpenoids (sesquiterpene lactones and dimmers, diterpenes and triterpenoids and flavonoids. The isolated compounds have shown diverse biological activities: anticancer, antioxidant, anti-inflammatory, neuroprotective and hepatoprotective activities. This review provides information on isolated bioactive phytochemicals and pharmacological potentials of I. britannica.

  18. IN VITRO ANALYSIS OF τ PHOSPHORYLATION SITES AND ITS BIOLOGICAL ACTIVITY

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    Objective.To explore the association between the abnormal phosphorylation sites found in Alzheimer disease (AD) τ and the inhibition of its biological activity. Methods.Ultracentrifugation,chromatography,manual Edman degradation and autosequence techniques were used to prepare and phosphorylate human recombinant τ ,isolate and purify 32P τ peptides and determine phosphorylation sites. Results.Phosphorylation of τ by casein kinase 1 (CK 1),cyclic AMP dependent protein kinase (PKA) and glycogen synthetase kinase 3 (GSK 3) separately inhibited its biological activity and the inhibition of this activity by GSK 3 was significantly increased if τ was prephosphorylated by CK 1 or PKA.The most potent inhibition was seen by a combined phosphorylation of τ with PKA and GSK 3.The treatment of τ by PKA and GSK 3 combination induced phosphorylation of τ at Ser 195,Ser 198,Ser 199,Ser 202,Thr 205,Thr 231,Ser 235,Ser 262,Ser 356,Ser 404,whereas Thr 181,Ser 184,Ser 262,Ser 356 and Ser 400 were phosphorylated by GSK 3 alone under the same condition. Conclusion.Phosphorylation of τ by PKA plus GSK 3 at Thr 205 might play a key role in τ pathology in AD.

  19. Synthesis, structure and biological properties of active spirohydantoin derivatives

    Directory of Open Access Journals (Sweden)

    Lazić Anita M.

    2016-01-01

    Full Text Available Spirohidantoins represent an pharmacologically important class of heterocycles since many derivatives have been recognized that display interesting activities against a wide range of biological targets. First synthesis of cycloalkanespiro-5-hydantoins was performed by Bucherer and Lieb 1934 by the reaction of cycloalkanone, potassium cyanide and ammonium-carbonate at reflux in a mixture of ethanol and water. QSAR (Quantitative Structure-Activity Relationship studies showed that a wide range of biological activities of spirohydantoin derivatives strongly depend upon their structure. This paper describes different methods of synthesis of spirohydantoin derivatives, their physico-chemical properties and biological activity. It emphasizes the importance of cycloalkanespiro-5-hydantoins with anticonvulsant, antiproliferative, antipsychotic, antimicrobial and antiinflammatory properties as well as their importance in the treatment of diabetes. Numerous spirohydantoin compounds exhibit physiological activity such as serotonin and fibrinogen antagonist, inhibitors of the glycine binding site of the NMDA receptor also, antagonist of leukocyte cell adhesion, acting as allosteric inhibitors of the protein-protein interactions. Some spirohydantoin derivatives have been identified as antitumor agents. Their activity depends on the substituent presented at position N-3 of the hydantoin ring and increases in order alkene > ester > ether. Besides that, compounds that contain two electron withdrawing groups (e.g. fluorine or chlorine on the third and fourth position of the phenyl ring are better antitumor agents than compounds with a single electron withdrawing group. [Projekat Ministarstva nauke Republike Srbije, br. 172013

  20. A macroporous bioreactor super activated by the recombinant human transforming growth factor-beta 3

    Directory of Open Access Journals (Sweden)

    Ugo eRipamonti

    2012-06-01

    Full Text Available Macroporous single-phase hydroxyapatite (HA and biphasic HA/β-tricalcium phosphate with 33% post-sinter hydroxyapatite (HA/β-TCP were combined with 25 or 125 µg recombinant human transforming growth factor-β3 (hTGF-β3 to engineer a super activated bioreactor implanted in orthotopic calvarial and heterotopic rectus abdominis muscle sites and harvested on day 30 and 90. Coral-derived calcium carbonate fully converted (100% and partially converted to 5% and 13% hydroxyapatite/calcium carbonate (HA/CC preloaded with 125 and 250 µg hTGF-β3, and 1:5 and 5:1 binary applications of hTGF-β3: hOP-1 by weight, were implanted in the rectus abdominis and harvested on day 20 and 30, respectively, to monitor spatial/temporal morphogenesis by high doses of hTGF-β3. Bone formation was assessed on decalcified paraffin-embedded sections by measuring the fractional volume of newly-formed bone. On day 30 and 90, single phase HA implants showed greater amounts of bone when compared to biphasic specimens; 5 % and 13 % HA/CC pre-loaded with 125 and 250 µg hTGF-β3 showed substantial induction of bone formation; 250 µg hTGF-β3 induced as yet unreported massive induction of bone formation as early as 20 days prominently outside the profile of the macroporous constructs. The induction of bone formation is controlled by the implanted ratio of the recombinant morphogens, i.e. the 1:5 hTGF-β3:hOP-1 ratio by weight was greater than the inverse ratio. The unprecedented tissue induction by single doses of 250 µg hTGF-β3 resulting in rapid bone morphogenesis of vast mineralized ossicles with multiple trabeculations surfaced by contiguous secreting osteoblasts is the novel molecular and morphological frontier for the induction of bone formation in clinical contexts.

  1. The Helper Activities of Different Avian Viruses for Propagation of Recombinant Avian Adeno-Associated Virus

    Institute of Scientific and Technical Information of China (English)

    WANG An-ping; SUN Huai-chang; WANG Jian-ye; WANG Yong-juan; YUAN Wei-feng

    2007-01-01

    To compare the helper activities of different avian viruses for propagation of recombinant avian adeno-associated virus (rAAAV), AAV-293 cells were cotransfected with the AAAV vector pAITR-GFP containing green fluorescent protein (GFP) gene, the AAAV helper vector pcDNA-ARC expressing the rep and cap genes, and the adenovirus helper vector pHelper expressing Ad5 E2A, E4, and VA-RNA genes. Chicken embryonic fibroblast (CEF) or chicken embryonic liver (CEL) cells were cotransfected with the AAAV vector and the AAAV helper vector, followed by infection with Marek's disease virus (MDV), avian adenovirus, chicken embryo lethal orphan (CELO) virus or infectious bursal disease virus (IBDV). Infectious rAAAV particles generated by the two strategies were harvested and titrated on CEF and CEL cells. A significantly higher viral titer was obtained with the helper activity provided by the pHelper vector than by MDV or CELO virus. Further experiments showed that rAAAV-mediated green fluorescent protein (gfp) expression was overtly enhanced by MDV or CELO virus super infection or treatment with sodium butyric acid, but not by IBDV super infection. These data demonstrated that MDV and CELO viruses could provide weak helper activity for propagation of rAAAV, and rAAAV-mediated transgene expression could be enhanced by super infection with the helper viruses.

  2. Oscillatory Activities in Regulatory Biological Networks and Hopf Bifurcation

    Institute of Scientific and Technical Information of China (English)

    YAN Shi-Wei; WANG Qi; XIE Bai-Song; ZHANG Feng-Shou

    2007-01-01

    Exploiting the nonlinear dynamics in the negative feedback loop, we propose a statistical signal-response model to describe the different oscillatory behaviour in a biological network motif. By choosing the delay as a bifurcation parameter, we discuss the existence of Hopf bifurcation and the stability of the periodic solutions of model equations with the centre manifold theorem and the normal form theory. It is shown that a periodic solution is born in a Hopf bifurcation beyond a critical time delay, and thus the bifurcation phenomenon may be important to elucidate the mechanism of oscillatory activities in regulatory biological networks.

  3. Tumour necrosis factor production and natural killer cell activity in peripheral blood during treatment with recombinant tumour necrosis factor

    OpenAIRE

    Männel, Daniela N.; Kist, A.; Ho, A D; Räth, U.; Reichardt, P; Wiedenmann, B; Schlick, E.; Kirchner, H.

    1989-01-01

    Tumour necrosis factor (TNF) has been found to be an important immunomodulator. Among other functions TNF activates natural killer (NK) cells and stimulates monocytes/macrophages in an autocrine fashion. TNF production and NK activity in peripheral blood mononuclear cells were determined in a clinical phase I study in which recombinant human (rh) TNF was administered as a continuous infusion weekly for a period of 8 weeks. Even though TNF production and NK activity were significantly reduced ...

  4. Synthesis of Lactosylated Piperazinyl Porphyrins and Their Biological Activity

    Institute of Scientific and Technical Information of China (English)

    LI,He-Ping; CAO,Zhong; XIAO,Hua-Wu

    2008-01-01

    The aim of this work is to synthesize of a new family of lactosylated piperazinly porphyrins, in which the galactoside piperazine moieties are linked to the tetra- and mono-phenyl rings of tetraphenylporphyrin (TPP).5,10,15,20-Tetrakis[4-(4-lactobionylpiperazin-1-yl)phenyl]porphyrin (TLPP) and 5-mono[4-(4-lactobionylpiperazin-1-yl)phenyl]-10,15,20-triphenylporphyrin (MLPP) have been synthesized. A detailed 1H NMR study gave their complete structural elucidation. The UV-Visible, mass spectra and elemental analysis are also presented. The biological activity on cancer cells and the pharmacokinetics have also been evaluated, showing a better biological activity, a very high liver to skin ratio and short retention time in tissues. It was suggested that such novel lactosylated piperazinly porphyrins, as potential hepatocyte-selective targeting drugs, exhibit a promising activity in photodynamic therapy (PDT).

  5. Protein stability and enzyme activity at extreme biological temperatures

    Energy Technology Data Exchange (ETDEWEB)

    Feller, Georges, E-mail: gfeller@ulg.ac.b [Laboratory of Biochemistry, Centre for Protein Engineering, Institute of Chemistry B6a, University of Liege, B-4000 Liege (Belgium)

    2010-08-18

    Psychrophilic microorganisms thrive in permanently cold environments, even at subzero temperatures. To maintain metabolic rates compatible with sustained life, they have improved the dynamics of their protein structures, thereby enabling appropriate molecular motions required for biological activity at low temperatures. As a consequence of this structural flexibility, psychrophilic proteins are unstable and heat-labile. In the upper range of biological temperatures, thermophiles and hyperthermophiles grow at temperatures > 100 {sup 0}C and synthesize ultra-stable proteins. However, thermophilic enzymes are nearly inactive at room temperature as a result of their compactness and rigidity. At the molecular level, both types of extremophilic proteins have adapted the same structural factors, but in opposite directions, to address either activity at low temperatures or stability in hot environments. A model based on folding funnels is proposed accounting for the stability-activity relationships in extremophilic proteins. (topical review)

  6. BIOLOGICAL ACTIVITIES OF OXAZINE AND ITS DERIVATIVES: A REVIEW

    Directory of Open Access Journals (Sweden)

    SINDHU T J

    2014-12-01

    Full Text Available Oxazine derivatives are an important class of heterocycles, which has attracted much synthetic interest due to their wide range of biological activities. Oxazine is a heterocyclic compound can be formally derived from benzene, and its reduction products, by suitable substitution of carbon (and hydrogen atoms by nitrogen and oxygen. In the last few years oxazine derivatives have proved to be valuable synthetic intermediates and also possess important biological activities like sedative, analgesic, antipyretic, anticonvulsant, antitubercular, antitumour, antimalarial and antimicrobial. In these days, development of drug resistance is a major problem and to overcome this situation, it is necessary to synthesize new classes of compounds. The aim of the article is to review the generalization of the collected data about the synthesis of oxazine derivatives and their activities. We hope that this work will be a definite interest for researchers concerned with azines in generally and oxazines in particular.

  7. Polysaccharides of higher fungi: Biological role, structure, and antioxidative activity

    Directory of Open Access Journals (Sweden)

    Kozarski Maja S.

    2014-01-01

    Full Text Available Fungal polysaccharides attract a lot of attention due to their multiple challenging biological properties, such as: anti-tumor, anti-viral, anticomplementary, anticoagulant, hypolipidemic and immunomodulatory and immune-stimulatory activities, which all together make them suitable for application in many quite distinctive areas, such as food industry, biomedicine, cosmetology, agriculture, environmental protection and waste water management. This article presents results with respect to biological properties, structure and procedures related to the isolation and activation of polysaccharides of higher fungi. It is considered and presented along with a review of the critical antioxidative activity and possible influence of the structural composition of polysaccharide extracts (isolated from these higher fungi upon their antioxidative properties.

  8. Pharmacokinetics of human recombinant tissue-type plasminogen activator, administered intra-abdominally, in a rat peritonitis model

    NARCIS (Netherlands)

    van Goor, Harry; Bom, VJJ; van der Meer, J; Sluiter, WJ; Geerards, S; de Graaf, JS; Bleichrodt, RP; van der Schaaf, W

    1996-01-01

    Human recombinant tissue-type plasminogen activator (rtPA), administered intraperitoneally, may promote intraabdominal fibrinolysis in peritonitis, thereby preventing adhesion and abscess formation. The pharmacokinetics of a single intraperitoneal dose of 0.5 or 2.0 mg/ml human rtPA were assessed in

  9. Recombinant goose-type lysozyme in channel catfish: Lysozyme activity and efficacy as plasmid DNA immunostimulant against Aeromonas hydrophila infection

    Science.gov (United States)

    The objectives of this study were: 1) to investigate whether recombinant channel catfish lysozyme g (CC-Lys-g) produced in E. coli expression system possesses any lysozyme activity; and 2) to evaluate whether channel catfish lysozyme g plasmid DNA could be used as an immunostimulant to protect chann...

  10. Active immunotherapy of allergic asthma with a recombinant human interleukin-5 protein as vaccine in a murine model

    Institute of Scientific and Technical Information of China (English)

    TAN Guang-hong; WANG Cai-chun; HUANG Feng-ying; WANG Hua; HUANG Yong-hao; LIN Ying-ying

    2007-01-01

    Background Eosinophils are highly related to allergic asthma inflammation. Interleukin (IL)-5 is the major chemokine of eosinophils, inhibition of the activity of IL-5 thus seems to be a potential approach to asthma therapy. The current study was performed to determine whether a recombinant human IL-5 protein as a xenogeneic vaccine has the capability of inducing anti-asthma activities.Methods Recombinant human IL-5 was used as a protein vaccine. Mouse asthma model was established to observe the anti-asthma activities. Lung histology was observed; eosinophils in blood and bronchoalveolar lavage were stained and counted. Airway hyperresponsiveness was determined by whole body plethysmograph. Antibody characters and cytokines were detected with enzyme linked immunosorbent assay (ELISA) and Western blot assay.Results Vaccination with recombinant human IL-5 protein as vaccine significantly reduced airway inflammation and airway hyperresponsiveness, and shifted the cytokine production from Th2 (IL-4) to Th1 (INF-γ) in mice allergic-asthma model. Immunization with recombinant human IL-5 protein vaccine bypassed the immunological tolerance and induced production of polyclonal antibodies that were cross-reactive with murine IL-5.Conclusions Active immunization with xenogeneic homologous IL-5 may be a possible therapeutic approach to the treatment of asthma and potentially of other eosinophilic disorders.

  11. Prolonged binding of radiolabeled recombinant tissue-type plasminogen activator after angioplasty and enclosed thrombolysis of the femoropopliteal arteries

    DEFF Research Database (Denmark)

    Tønnesen, K H; Vinberg, N; Folkenborg, O

    1992-01-01

    The authors measured the binding of indium-111-labeled recombinant tissue-type plasminogen activator (rt-PA) within the recanalized femoropopliteal segment after percutaneous transluminal angioplasty (PTA) and enclosed thrombolysis. In patients with long occlusions (n = 3), 91 micrograms of rt...

  12. Recombinant tissue plasminogen activator as a novel treatment option for infective endocarditis: a retrospective clinical study in 32 children.

    Science.gov (United States)

    Levitas, Aviva; Krymko, Hanna; Richardson, Justin; Zalzstein, Eli; Ioffe, Viktoriya

    2016-01-01

    Infective endocarditis is a life-threatening infectious syndrome, with high morbidity and mortality. Current treatments for infective endocarditis include intravenous antibiotics, surgery, and involve a lengthy hospital stay. We hypothesised that adjunctive recombinant tissue plasminogen activator treatment for infective endocarditis may facilitate faster resolution of vegetations and clearance of positive blood cultures, and therefore decrease morbidity and mortality. This retrospective study included follow-up of patients, from 1997 through 2014, including clinical presentation, causative organism, length of treatment, morbidity, and mortality. We identified 32 patients, all of whom were diagnosed with endocarditis and were treated by recombinant tissue plasminogen activator. Among all, 27 patients (93%) had positive blood cultures, with the most frequent organisms being Staphylococcus epidermis (nine patients), Staphylococcus aureus (six patients), and Candida (nine patients). Upon treatment, in 31 patients (97%), resolution of vegetations and clearance of blood cultures occurred within hours to few days. Out of 32 patients, one patient (3%) died and three patients (9%) suffered embolic or haemorrhagic events, possibly related to the recombinant tissue plasminogen activator. None of the patients required surgical intervention to assist vegetation resolution. In conclusion, it appears that recombinant tissue plasminogen activator may become an adjunctive treatment for infective endocarditis and may decrease morbidity as compared with current guidelines. Prospective multi-centre studies are required to validate our findings.

  13. Bioconjugation of recombinant tissue plasminogen activator to magnetic nanocarriers for targeted thrombolysis

    Directory of Open Access Journals (Sweden)

    Yang HW

    2012-10-01

    Full Text Available Hung-Wei Yang,1,* Mu-Yi Hua,1,* Kun-Ju Lin,2,* Shiaw-Pyng Wey,3 Rung-Ywan Tsai,4 Siao-Yun Wu,5 Yi-Ching Lu,5 Hao-Li Liu,6 Tony Wu,7 Yunn-Hwa Ma5 1Chang Gung Molecular Medicine Research Center, Department of Chemical and Materials Engineering, 2Molecular Imaging Center, Department of Nuclear Medicine, Chang Gung Memorial Hospital, Kuei-Shan, Tao-Yuan, Taiwan, Republic of China; 3Department of Medical Imaging and Radiological Sciences, 4Electronics and Optoelectronics Research Laboratories, Industrial Technology Research Institute, Hsin-chu, Taiwan, Republic of China; 5Department of Physiology and Pharmacology and Healthy Aging Research Center, 6Department of Electrical Engineering, Chang Gung University, Kuei-Shan, Tao-Yuan, Taiwan, Republic of China; 7Department of Neurology, Chang Gung University College of Medicine and Memorial Hospital, Tao-Yuan, Taiwan, Republic of China*These authors contributed equally to this workAbstract: Low-toxicity magnetic nanocarriers (MNCs composed of a shell of poly [aniline-co-N-(1-one-butyric acid aniline] over a Fe3O4 magnetic nanoparticle core were developed to carry recombinant tissue plasminogen activator (rtPA in MNC-rtPA for targeted thrombolysis. With an average diameter of 14.8 nm, the MNCs exerted superparamagnetic properties. Up to 276 µg of active rtPA was immobilized per mg of MNCs, and the stability of the immobilized rtPA was greatly improved during storage at 4°C and 25°C. In vitro thrombolysis testing with a tubing system demonstrated that magnet-guided MNC-rtPA showed significantly improved thrombolysis compared with free rtPA and reduced the clot lysis time from 39.2 ± 3.2 minutes to 10.8 ± 4.2 minutes. In addition, magnet-guided MNC-rtPA at 20% of the regular rtPA dose restored blood flow within 15–25 minutes of treatment in a rat embolism model without triggering hematological toxicity. In conclusion, this improved system is based on magnetic targeting accelerated thrombolysis and is

  14. Using recombinant CD74 protein to inhibit the activity of macrophage migration inhibitory factor (MIF) in vitro

    Institute of Scientific and Technical Information of China (English)

    Zhi-xinSHAN; Xi-yongYU; Qiu-xiongLIN; Yong-hengFU

    2005-01-01

    AIM Macrophage migration inhibitory factor (MIF) is a pro-inflammatory cytokine involved in the pathogenesis of a variety of autoimmune and inflammatory diseases, including arthritis, glomerulonephritis, Gram-positive and Gram-negative sepsis, and atherogenesis. Recent studies showed that CD74(antigen-associated invariant chain Ⅱ) is a high-affinity membrane-binding protein for MIF. The purpose of the present study was to express the recombinant human CD74 in E. coli and inhibit the activity of MIF by using recombinant CD74 in vitro.

  15. Hydrodynamic collective effects of active proteins in biological membranes

    CERN Document Server

    Koyano, Yuki; Mikhailov, Alexander S

    2016-01-01

    Lipid bilayers forming biological membranes are known to behave as viscous 2D fluids on submicrometer scales; usually they contain a large number of active protein inclusions. Recently, it has been shown [Proc. Nat. Acad. Sci. USA 112, E3639 (2015)] that such active proteins should in- duce non-thermal fluctuating lipid flows leading to diffusion enhancement and chemotaxis-like drift for passive inclusions in biomembranes. Here, a detailed analytical and numerical investigation of such effects is performed. The attention is focused on the situations when proteins are concentrated within lipid rafts. We demonstrate that passive particles tend to become attracted by active rafts and are accumulated inside them.

  16. Restricted expression of recombination activating gene (RAG-1) in mouse lymphoid tissues

    Energy Technology Data Exchange (ETDEWEB)

    Yamamoto, Akihito; Fujinaga, Hiroyuki; Hamatani, Kiyohiro [Radiation Effects Research Foundation, Nagasaki (Japan). Nagasaki Branch; Atsuta, Mitsuru

    1993-03-01

    In an attempt to determine the distribution of recombinase activity in the mouse thymus, spleen, and lymph nodes, we used the in situ hybridization method to examine the expression of the recombination activating genes RAG-1 and RAG-2. Expression of RAG-1 was found in most cortical thymocytes but not in the majority of medullary thymocytes. Although hybridization signals of RAG-2 were not as intense as those of RAG-1, the localization of RAG-2 transcripts was similar to that of RAG-1. In the spleen, expression of RAG-1 was found only in limited cells near the splenic sinus, and the majority of the cells within the follicle were negative for RAG-1 transcript. In nude mice, RAG-1-expressing cells were detected in the same regions, which suggests that in situ hybridization signals of RAG-1 in the spleen are due to the cells of B-cell origin. In the lymph nodes, expression of RAG-1 was found only in the medullary region. Expression of RAG-2 transcript in the spleen and the lymph nodes, if any, was too faint to allow determination of the specific localization. These results suggest that most of the cortical thymocytes and some cells in the spleen are capable of rearranging T-cell receptor genes and immunoglobulin genes, respectively, but the possible involvement of the RAG-1 transcript in RAG-1-positive cells of the spleen and the lymph nodes in functions other than the rearrangement of genes could not be ruled out. (author).

  17. Recombinant goose-type lysozyme in channel catfish: lysozyme activity and efficacy as plasmid DNA immunostimulant against Aeromonas hydrophila infection.

    Science.gov (United States)

    Pridgeon, Julia W; Klesius, Phillip H; Dominowski, Paul J; Yancey, Robert J; Kievit, Michele S

    2013-10-01

    The objectives of this study were: 1) to investigate whether recombinant channel catfish lysozyme-g (CC-Lys-g) produced in Escherichia coli expression system possesses any lysozyme activity; and 2) to evaluate whether channel catfish lysozyme-g plasmid DNA could be used as an immunostimulant to protect channel catfish against Aeromonas hydrophila infection. Recombinant CC-Lys-g produced in E. coli expression system exhibited significant (P recombinant channel catfish lysozyme-g (pcDNA-Lys-g) was transfected in channel catfish gill cells G1B, the over-expression of pcDNA-Lys-g offered significant (P DNA injection. Macrophages of fish injected with pcDNA-Lys-g produced significantly (P DNA injection. Taken together, our results suggest that pcDNA-Lys-g could be used as a novel immunostimulant to offer immediate protection to channel catfish against A. hydrophila infection.

  18. Evaluation of Aryoseven Safety (Recombinant Activated Factor VII) in Patients with Bleeding Disorders (An Observational Post-Marketing Surveillance Study)

    Science.gov (United States)

    Toogeh, Gholamreza; Abolghasemi, Hassan; Eshghi, Peyman; Managhchi, Mohammadreza; Shaverdi-niasari, Mohammadreza; Karimi, Katayoon; Roostaei, Samin; Emran, Neda; Abdollahi, Alireza

    2016-01-01

    Background: Recombinant activated factor VII induces hemostasis in patients with coagulopathy disorders. AryoSeven™ as a safe Iranian Recombinant activated factor VII has been available on our market. This study was performed to establish the safety of AryoSeven on patients with coagulopathy disorder. Methods: This single-center, descriptive, cross sectional study was carried out in Thrombus and Homeostasis Research Center ValiAsr Hospital during 2013-2014. Fifty one patients with bleeding disorders who received at least one dose of Aryoseven were enrolled. Patients’ demographic data and adverse effect of drug and reaction related to Aryoseven or previous usage of Recombinant activated FVII were recorded in questionnaires. Finally data were analyzed to compare side effects of Aryoseven and other Recombinant activated FVII brands. Results: Aryoseven was prescribed for 51 Patients. Of all participants with mean age 57.18+21.38 yr, 31 cases were male and 26 subjects had past history of recombinant activated FVII usage. Glanzman was the most frequent disorder followed by congenital FVII deficiency, hemophilia with inhibitors, factor 5 deficiency, acquired hemophilia, hemophilia A with inhibitor, and hemophilia A or B with inhibitor. The majority of bleeding episodes had occurred in joints. Three patients (5.9%) complained about adverse effects of Aryoseven vs. 11.5 % about adverse effects of other brands. However this difference was not significant, statistically. Conclusion: Based on monitor patients closely for any adverse events, we concluded that Aryoseven administration under careful weighing of benefit versus potential harm may comparable with other counterpart drugs. PMID:27799968

  19. Molecular cloning, recombinant gene expression, and antifungal activity of cystatin from taro (Colocasia esculenta cv. Kaosiung no. 1).

    Science.gov (United States)

    Yang, A H; Yeh, K W

    2005-06-01

    A cDNA clone, designated CeCPI, encoding a novel phytocystatin was isolated from taro corms (Colocasia esculenta) using both degenerated primers/RT-PCR amplification and 5'-/3'-RACE extension. The full-length cDNA gene is 1,008 bp in size, encodes 206 amino acid residues, with a deduced molecular weight of 29 kDa. It contains a conserved reactive site motif Gln-Val-Val-Ser-Gly of cysteine protease inhibitors, and another consensus ARFAV sequence for phytocystatin. Sequence analysis revealed that CeCPI is phylogenetically closely related to Eudicots rather than to Monocots, despite taro belonging to Monocot. Recombinant GST-CeCPI fusion protein was overexpressed in Escherichia coli and its inhibitory activity against papain was identified on gelatin/SDS-PAGE. These results confirmed that recombinant CeCPI protein exhibited strong cysteine protease inhibitory activity. Investigation of its antifungal activity clearly revealed a toxic effect on the mycelium growth of phytopathogenic fungi, such as Sclerotium rolfsii Sacc. etc., at a concentration of 80 microg recombinant CeCPI/ ml. Moreover, mycelium growth was completely inhibited and the sclerotia lysed at a concentration of 150-200 microg/ml. Further studies have demonstrated that recombinant CeCPI is capable of acting against the endogenous cysteine proteinase in the fungal mycelium.

  20. Design and construction of an in-plant activation cassette for transgene expression and recombinant protein production in plants.

    Science.gov (United States)

    Dugdale, Benjamin; Mortimer, Cara L; Kato, Maiko; James, Tess A; Harding, Robert M; Dale, James L

    2014-05-01

    Virus-based transgene expression systems have become particularly valuable for recombinant protein production in plants. The dual-module in-plant activation (INPACT) expression platform consists of a uniquely designed split-gene cassette incorporating the cis replication elements of Tobacco yellow dwarf geminivirus (TYDV) and an ethanol-inducible activation cassette encoding the TYDV Rep and RepA replication-associated proteins. The INPACT system is essentially tailored for recombinant protein production in stably transformed plants and provides both inducible and high-level transient transgene expression with the potential to be adapted to diverse crop species. The construction of a novel split-gene cassette, the inducible nature of the system and the ability to amplify transgene expression via rolling-circle replication differentiates this system from other DNA- and RNA-based virus vector systems used for stable or transient recombinant protein production in plants. Here we provide a detailed protocol describing the design and construction of a split-gene INPACT cassette, and we highlight factors that may influence optimal activation and amplification of gene expression in transgenic plants. By using Nicotiana tabacum, the protocol takes 6-9 months to complete, and recombinant proteins expressed using INPACT can accumulate to up to 10% of the leaf total soluble protein.

  1. Biological activity of selected plants with adaptogenic effect

    Directory of Open Access Journals (Sweden)

    Eva Ivanišová

    2016-05-01

    Full Text Available The aim of this study was to determine biological activity of plants with adaptogenic effect: Panax ginseng Mayer., Withania somnifera L., Eleuterococcus senticosus Rupr. et Maxim., Astragallus membranaceus Fisch. and Codonopsis pilosulae Franch. The antioxidant activity was detected by DPPH and phosphomolybdenum method, total polyphenol content with Folin – Ciocalteu reagent, flavonoids content by aluminium chloride method. The detection of antimicrobial activity was carried out by disc diffusion method against three species of Gram-negative bacteria: Escherichia coli CCM 3988, Salmonella enterica subsp. enterica CCM 3807, Yersinia enterocolitica CCM 5671 and two Gram-positive bacteria: Bacillus thuringiensis CCM 19, Stapylococcus aureus subsp. aureus CCM 2461. Results showed that plants with adaptogenic effect are rich for biologically active substances. The highest antioxidant activity by DPPH method was determined in the sample of Eleuterococcus senticosus (3.15 mg TEAC – Trolox equivalent antioxidant capacity per g of sample and by phosphomolybdenum method in the sample of Codonopsis pilosulae (188.79 mg TEAC per g of sample. In the sample of Panax ginseng was measured the highest content of total polyphenols (8.10 mg GAE – galic acid equivalent per g of sample and flavonoids (3.41 μg QE – quercetin equivalent per g of sample. All samples also showed strong antimicrobial activity with the best results in Panax ginseng and Withania somnifera in particular for species Yersinia enterocolitica CCM 5671 and Salmonella enterica subsp. enterica CCM 3807. The analyzed species of plant with high value of biological activity can be used more in the future, not only in food, but also in cosmetics and pharmaceutical industries.

  2. Marine Sponge Lectins: Actual Status on Properties and Biological Activities

    Directory of Open Access Journals (Sweden)

    Sandro Mascena Gomes Filho

    2014-12-01

    Full Text Available Marine sponges are primitive metazoans that produce a wide variety of molecules that protect them against predators. In studies that search for bioactive molecules, these marine invertebrates stand out as promising sources of new biologically-active molecules, many of which are still unknown or little studied; thus being an unexplored biotechnological resource of high added value. Among these molecules, lectins are proteins that reversibly bind to carbohydrates without modifying them. In this review, various structural features and biological activities of lectins derived from marine sponges so far described in the scientific literature are discussed. From the results found in the literature, it could be concluded that lectins derived from marine sponges are structurally diverse proteins with great potential for application in the production of biopharmaceuticals, especially as antibacterial and antitumor agents.

  3. MILK KEFIR: COMPOSITION, MICROBIAL CULTURES, BIOLOGICAL ACTIVITIES AND RELATED PRODUCTS

    Directory of Open Access Journals (Sweden)

    Maria Rosa Prado

    2015-10-01

    Full Text Available In recent years, there has been a strong focus on beneficial foods with probiotic microorganisms and functional organic substances. In this context, there is an increasing interest in the commercial use of kefir, since it can be marketed as a natural beverage that has health promoting bacteria. There are numerous commercially available kefir based-products. Kefir may act as a matrix in the effective delivery of probiotic microorganisms in different types of products. Also, the presence of kefir’s exopolysaccharides, known as kefiran, which has biological activity, certainly adds value to products. Kefiran can also be used separately in other food products and as a coating film for various food and pharmaceutical products. This article aims to update the information about kefir and its microbiological composition, biological activity of the kefir’s microflora and the importance of kefiran as a beneficial health substance.

  4. Marine Omega-3 Phospholipids: Metabolism and Biological Activities

    Directory of Open Access Journals (Sweden)

    Nils Hoem

    2012-11-01

    Full Text Available The biological activities of omega-3 fatty acids (n-3 FAs have been under extensive study for several decades. However, not much attention has been paid to differences of dietary forms, such as triglycerides (TGs versus ethyl esters or phospholipids (PLs. New innovative marine raw materials, like krill and fish by-products, present n-3 FAs mainly in the PL form. With their increasing availability, new evidence has emerged on n-3 PL biological activities and differences to n-3 TGs. In this review, we describe the recently discovered nutritional properties of n-3 PLs on different parameters of metabolic syndrome and highlight their different metabolic bioavailability in comparison to other dietary forms of n-3 FAs.

  5. Biological activities of secondary metabolites of the order Zoanthids

    Directory of Open Access Journals (Sweden)

    Zahra Aminikhoei

    2015-11-01

    Full Text Available The phylum Cnidaria is a large, diverse and ecologically important group of marine invertebrates, which produce powerful toxins and venoms. The number of marine natural product from cnidarians isolated from class Anthozoa. Among the Anthozoa, the order of zoanthids are sessile, clonal and mostly brightly colored invertebrate which produce high biodiversity of cytolitic, neurotoxic and cardiotoxic compounds. Zoanthids containing palytoxins are reportedly among the most toxic marine organisms known. In addition, a high concentration of zoanthamine alkaloids extracted from this group.The zoanthamine alkaloids were isolated over 20 years ago, exhibit a broad range of biological activities.The best studied and most well-known biological activity of zoanthamine derivative significantly suppressed bone resorption and enhanced bone formation.

  6. Biological activities and medicinal properties of Gokhru (Pedalium murex L.)

    Institute of Scientific and Technical Information of China (English)

    V Rajashekar; E Upender Rao; Srinivas P

    2012-01-01

    Bada Gokhru (Pedalium murex L.) is perhaps the most useful traditional medicinal plant in India. Each part of the neem tree has some medicinal property and is thus commercially exploitable. During the last five decades, apart from the chemistry of the Pedalium murex compounds, considerable progress has been achieved regarding the biological activity and medicinal applications of this plant. It is now considered as a valuable source of unique natural products for development of medicines against various diseases and also for the development of industrial products. This review gives a bird’s eye view mainly on the biological activities of some of this compounds isolated, pharmacological actions of the extracts, clinical studies and plausible medicinal applications of gokharu along with their safety evaluation.

  7. Visual Analysis of Biological Activity Data with Scaffold Hunter.

    Science.gov (United States)

    Klein, Karsten; Koch, Oliver; Kriege, Nils; Mutzel, Petra; Schäfer, Till

    2013-12-01

    The growing interest in chemogenomics approaches over the last years has led to an increasing amount of data regarding chemical and the corresponding biological activity space. The resulting data, collected in either in-house or public databases, need to be analyzed efficiently to speed-up the increasingly difficult task of drug discovery. Unfortunately, the discovery of new chemical entities or new targets for known drugs ('drug repurposing') is not suitable to a fully automated analysis or a simple drill down process. Visual interactive interfaces that allow to explore chemical space in a systematic manner and facilitate analytical reasoning can help to overcome these problems. Scaffold Hunter is a tool for the visual analysis of chemical compound databases that provides integrated visualization and analysis of biological activity data and fosters the interactive exploration of data imported from a variety of sources. We describe the features and illustrate the use by means of an exemplary analysis workflow.

  8. Fucoidans from brown alga Fucus evanescens: structure and biological activity

    Directory of Open Access Journals (Sweden)

    Roza Menshova

    2016-08-01

    Full Text Available Brown alga Fucus evanescens, widespread in the Far Eastern seas of Russia, is valuable source of sulfated polysaccharides – fucoidans with beneficial biological activities. The most homogenous fraction of fucoidan from F. evanescens was shown to be molecule containing linear main chain of alternating 2-sulfated 1,3- and 1,4-linked α-L-fucose residues. Few sulfate groups were found in position 4 of some 1,3-linked fucose residues. Acetyl groups occupied free C-3 of 1,4-linked residues and/or the C-4 of 1,3-linked fucose residues. Enzymatic hydrolysis, mild acid hydrolysis and autohydrolysis of native fucoidan were used for elucidation of the fine structural characteristics of fucoidan from F. evanescens. The aim of this review to summarize published data on biological activities of fucoidan from F. evanescens: antiviral, anticoagulant, thrombolytic, hepatoprotective, immunomodulatory, anticancer, and their practical application.

  9. Biological activities and medicinal properties of Cajanus cajan (L Millsp.

    Directory of Open Access Journals (Sweden)

    Dilipkumar Pal

    2011-01-01

    Full Text Available Cajanus cajan (L Millsp. (Sanskrit: Adhaki, Hindi: Arhar, English: Pigeon pea, Bengali: Tur (family: Fabaceae is the most important grain legume crop of rain-fed agriculture in semi-arid tropics. It is both a food crop and a cover/forage crop with high levels of proteins and important amino acids like methionine, lysine and tryptophan. During the last few decades extensive studies have been carried out regarding the chemistry of C. cajan and considerable progress has been achieved regarding its biological activities and medicinal applications. This review article gives an overview on the biological activities of the compounds isolated, pharmacological actions and clinical studies of C. cajan extracts apart from its general details.

  10. Recombinant-activated factor Ⅶ and neuronal apoptosis in a rat model of intracerebral hemorrhage

    Institute of Scientific and Technical Information of China (English)

    Qiang Li; Wei Li; Suju Ding; Jianping Tang; Jing Fang; Benqiang Deng; Tao Wu

    2009-01-01

    BACKGROUND:Activated clotting factor Ⅶ has been demonstrated to exhibit obvious anti-apoptosis effects.OBJECTIVE:To observe the effect of activated clotting factor Ⅶ on neuronal apoptosis at different time points following rat intracerebral hemorrhage (ICH).DESIGN,TIME AND SETTING:A randomized,controlled,animal experiment was performed at the Neurobiological Laboratory of Second Military Medical University from October 2005 to April 2006.MATERIALS:Recombinant-activated clotting factor Vlla (rFⅧa) was purchased from Danish Novo Nordisk,Denmark.In situ cell death detection kit-POD kit was purchased from Roche,Switzerland.Caspase-3 activity determination kit from Biovision,USA.METHODS:A total of 72 healthy,male,Sprague Dawley rats,aged 5-8 months,were randomly assigned to three groups (n=24):sham-operated,ICH model,and rFⅧa.In the ICH model and rFⅧa groups,80.0 μL autologous non-clotting blood from rat tails was injected into the right caudate putamen to establish the ICH.The empty microinjector was inserted into the caudate putamen in the sham-operated group.The ICH model and rFⅧa groups were subdivided into four subsets separately:6,24,72 hours and 7 clays following ICH.The rats in the rFⅧa group were injected with 160 μg/kg rFⅧa via the dorsal vein of the penis.MAIN OUTCOME MEASURES:Apoptotic cells were detected in the right caudate putamen by TUNEL;caspase-3 activity by spectrophotometry;and rat neurological function was evaluated by neurological functional impairment scales.RESULTS:Rat neurological function was deteriorated at 24,72 hours,and 7 days following ICH.The TUNEL-positive cells and caspase-3 activity in the right caudate putamen was significantly increased in the ICH rats (P<0.05);rFVlla treatment reduced the number of TUNEL-positive cells and caspase-3 activity in the right caudate putamen (P<0.05),and neurological function was significantly improved (P<0.05).CONCLUSION:rFⅧa was applied within 72 hours after ICH,which reduced

  11. Design of a Comprehensive Biochemistry and Molecular Biology Experiment: Phase Variation Caused by Recombinational Regulation of Bacterial Gene Expression

    Science.gov (United States)

    Sheng, Xiumei; Xu, Shungao; Lu, Renyun; Isaac, Dadzie; Zhang, Xueyi; Zhang, Haifang; Wang, Huifang; Qiao, Zheng; Huang, Xinxiang

    2014-01-01

    Scientific experiments are indispensable parts of Biochemistry and Molecular Biology. In this study, a comprehensive Biochemistry and Molecular Biology experiment about "Salmonella enterica" serovar Typhi Flagellar phase variation has been designed. It consisted of three parts, namely, inducement of bacterial Flagellar phase variation,…

  12. Synthesis,Characterization and Biological Activities of Novel Acrylamide Compounds

    Institute of Scientific and Technical Information of China (English)

    XU Liang-zhong; XU Zhong-jie; ZHANG Gong-sheng; ZHOU Kai; ZHAI Zhi-wei

    2008-01-01

    With dimethomorph and flumorph as the leading compounds,four novel acrylamide compounds with two types of structure were designed and synthesized by means of the method of"me too chemistry".The target compounds were characterized by 1H NMR,IR,MS,and elemental analysis.The influences of solvent and raw material on the yield were investigated and optimum processing conditions were determined.The results of preliminary biological tests show that all those compounds exhibit certain antifungal activities.

  13. Protein folding activity and the central dogma of molecular biology

    OpenAIRE

    Pallavi, Ghosh; Dipankar, Chatterji

    2003-01-01

    Biological systems, in general, can function effectively when the products of the system are in proper configuration and harmful effects due to misaggregation are avoided. Folding of proteins and their functional consequences have been a subject of active research since several years now. However it is not clear whether during protein synthesis from genetic message, the same set of rules are employed or participation of new efforts take place. In this review we show that at least in the case ...

  14. Chemistry and Biological Activities of Flavonoids: An Overview

    OpenAIRE

    Shashank Kumar; Pandey, Abhay K

    2013-01-01

    There has been increasing interest in the research on flavonoids from plant sources because of their versatile health benefits reported in various epidemiological studies. Since flavonoids are directly associated with human dietary ingredients and health, there is need to evaluate structure and function relationship. The bioavailability, metabolism, and biological activity of flavonoids depend upon the configuration, total number of hydroxyl groups, and substitution of functional groups about...

  15. Biological Activities of Phenolic Compounds Present in Virgin Olive Oil

    OpenAIRE

    2010-01-01

    The Mediterranean diet is associated with a lower incidence of atherosclerosis, cardiovascular disease, neurodegenerative diseases and certain types of cancer. The apparent health benefits have been partially ascribed to the dietary consumption of virgin olive oil by Mediterranean populations. Much research has focused on the biologically active phenolic compounds naturally present in virgin olive oils to aid in explaining reduced mortality and morbidity experienced by people consuming a trad...

  16. Current status of pyrazole and its biological activities.

    Science.gov (United States)

    Naim, Mohd Javed; Alam, Ozair; Nawaz, Farah; Alam, Md Jahangir; Alam, Perwaiz

    2016-01-01

    Pyrazole are potent medicinal scaffolds and exhibit a full spectrum of biological activities. This review throws light on the detailed synthetic approaches which have been applied for the synthesis of pyrazole. This has been followed by an in depth analysis of the pyrazole with respect to their medical significance. This follow-up may help the medicinal chemists to generate new leads possessing pyrazole nucleus with high efficacy.

  17. Biological Activities and Phytochemicals of Swietenia macrophylla King

    Directory of Open Access Journals (Sweden)

    Habsah Abdul Kadir

    2013-08-01

    Full Text Available Swietenia macrophylla King (Meliaceae is an endangered and medicinally important plant indigenous to tropical and subtropical regions of the World. S. macrophylla has been widely used in folk medicine to treat various diseases. The review reveals that limonoids and its derivatives are the major constituents of S. macrophylla. There are several data in the literature indicating a great variety of pharmacological activities of S. macrophylla, which exhibits antimicrobial, anti-inflammatory, antioxidant effects, antimutagenic, anticancer, antitumor and antidiabetic activities. Various other activities like anti-nociceptive, hypolipidemic, antidiarrhoeal, anti-infective, antiviral, antimalarial, acaricidal, antifeedant and heavy metal phytoremediation activity have also been reported. In view of the immense medicinal importance of S. macrophylla, this review aimed at compiling all currently available information on its ethnomedicinal uses, phytochemistry and biological activities of S. macrophylla, showing its importance.

  18. Embryonic stem cells deficient for Brca2 or Blm exhibit divergent genotoxic profiles that support opposing activities during homologous recombination

    Energy Technology Data Exchange (ETDEWEB)

    Marple, Teresa [Department of Molecular Medicine and Institute of Biotechnology, University of Texas Health Science Center at San Antonio, 15355 Lambda Drive San Antonio, TX 78245-3207 (United States); Kim, Tae Moon [Department of Molecular Medicine and Institute of Biotechnology, University of Texas Health Science Center at San Antonio, 15355 Lambda Drive San Antonio, TX 78245-3207 (United States); Hasty, Paul [Department of Molecular Medicine and Institute of Biotechnology, University of Texas Health Science Center at San Antonio, 15355 Lambda Drive San Antonio, TX 78245-3207 (United States)]. E-mail: hastye@uthscsa.edu

    2006-12-01

    The breast cancer susceptibility protein, Brca2 and the RecQ helicase, Blm (Bloom syndrome mutated) are tumor suppressors that maintain genome integrity, at least in part, through homologous recombination (HR). Brca2 facilitates HR by interacting with Rad51 in multiple regions, the BRC motifs encoded by exon 11 and a single domain encoded by exon 27; however, the exact importance of these regions is not fully understood. Blm also interacts with Rad51 and appears to suppress HR in most circumstances; however, its yeast homologue Sgs1 facilitates HR in response to some genotoxins. To better understand the biological importance of these two proteins, we performed a genotoxic screen on mouse embryonic stem (ES) cells impaired for either Brca2 or Blm to establish their genotoxic profiles (a cellular dose-response to a wide range of agents). This is the first side-by-side comparison of these two proteins in an identical genetic background. We compared cells deleted for Brca2 exon 27 to cells reduced for Blm expression and find that the Brca2- and Blm-impaired cells exhibit genotoxic profiles that reflect opposing activities during HR. Cells deleted for Brca2 exon 27 are hypersensitive to {gamma}-radiation, streptonigrin, mitomycin C and camptothecin and mildly resistant to ICRF-193 which is similar to HR defective cells null for Rad54. By contrast, Blm-impaired cells are hypersensitive to ICRF-193, mildly resistant to camptothecin and mitomycin C and more strongly resistant to hydroxyurea. These divergent profiles support the notion that Brca2 and Blm perform opposing functions during HR in mouse ES cells.

  19. Activity of the Human Rhinovirus 3C Protease Studied in Various Buffers, Additives and Detergents Solutions for Recombinant Protein Production

    OpenAIRE

    2016-01-01

    Proteases are widely used to remove affinity and solubility tags from recombinant proteins to avoid potential interference of these tags with the structure and function of the fusion partner. In recent years, great interest has been seen in use of the human rhinovirus 3C protease owing to its stringent sequence specificity and enhanced activity. Like other proteases, activity of the human rhinovirus 3C protease can be affected in part by the buffer components and additives that are generally ...

  20. Isolation of biologically active nanomaterial (inclusion bodies from bacterial cells

    Directory of Open Access Journals (Sweden)

    Peternel Špela

    2010-09-01

    Full Text Available Abstract Background In recent years bacterial inclusion bodies (IBs were recognised as highly pure deposits of active proteins inside bacterial cells. Such active nanoparticles are very interesting for further downstream protein isolation, as well as for many other applications in nanomedicine, cosmetic, chemical and pharmaceutical industry. To prepare large quantities of a high quality product, the whole bioprocess has to be optimised. This includes not only the cultivation of the bacterial culture, but also the isolation step itself, which can be of critical importance for the production process. To determine the most appropriate method for the isolation of biologically active nanoparticles, three methods for bacterial cell disruption were analyzed. Results In this study, enzymatic lysis and two mechanical methods, high-pressure homogenization and sonication, were compared. During enzymatic lysis the enzyme lysozyme was found to attach to the surface of IBs, and it could not be removed by simple washing. As this represents an additional impurity in the engineered nanoparticles, we concluded that enzymatic lysis is not the most suitable method for IBs isolation. During sonication proteins are released (lost from the surface of IBs and thus the surface of IBs appears more porous when compared to the other two methods. We also found that the acoustic output power needed to isolate the IBs from bacterial cells actually damages proteins structures, thereby causing a reduction in biological activity. High-pressure homogenization also caused some damage to IBs, however the protein loss from the IBs was negligible. Furthermore, homogenization had no side-effects on protein biological activity. Conclusions The study shows that among the three methods tested, homogenization is the most appropriate method for the isolation of active nanoparticles from bacterial cells.

  1. EVALUATION OF BIOLOGICALLY ACTIVE MOLECULES ISOLATED FROM OBLIGATE MARINE FUNGI

    Directory of Open Access Journals (Sweden)

    A. KRISHNA SATYA

    2013-01-01

    Full Text Available The marine environment is a tremendous source of natural products. Marine microorganisms have become an important source of pharmacologically active metabolites Fungi are well known for their vast diversity of secondary metabolites that include many life-saving drugs and highly toxic mycotoxins. In general, fungal cultures producing such metabolites are immune to their toxic effects. However, some are known to produce self-toxic compounds that can pose production optimization challenges if the metabolites are needed in large amounts for chemical modification. Objective: The main objective of the present study was the isolation of new and preferably biologically active secondary metabolites from marine microorganisms, especially marine-derived fungi. Method: Marine fungi had isolated from marine soil by serial dilution method from Rose Bengal medium. Single colony was isolated by microscopic and macroscopic observation. Secondary metabolites are produced by marine fungi. Biological evaluation was performed by microbial studies. TLC is performed to identify the number of sub compounds in the crude extract. Further species level identification and structure elucidation of the compound are to be done. Results: The isolated marine fungi Aspergillus sp, showed maximum activity against the Candida rugosa with a zone diameter of 16mm at a concentration of 200μg and for bacterial strains it showed maximum activity against the E.coli with a diameter of 24mm at a concentration of 200μg. From the thin layer chromatography it has nearly 2-3 compounds to be purified. Conclusion: The selected organism which produces the compounds contains the biological activities which include anti-bacterial and anti fungal activities.

  2. Depletion of arginine by recombinant arginine deiminase induces nNOS-activated neurotoxicity in neuroblastoma cells.

    Science.gov (United States)

    Lin, Shan-Erh; Wu, Fe-Lin Lin; Wei, Ming-Feng; Shen, Li-Jiuan

    2014-01-01

    The abnormal regulation of inducible nitric oxide synthase (iNOS) and neuronal nitric oxide synthase (nNOS) is associated with neurodegenerative disorders. Recombinant arginine deiminase (rADI) is a selective NO modulator of iNOS and eNOS in endothelial cells, and it also exhibits neuroprotective activity in an iNOS-induced neuron-microglia coculture system. However, the effect of rADI on nNOS remains unknown. Addressing this issue is important for evaluating the potential application of rADI in neurodegenerative diseases. SH-SY5Y cells were treated with N-methyl-D-aspartic acid (NMDA) to activate nNOS. NMDA increased NO production by 39.7 ± 3.9% via nNOS under arginine-containing conditions, but there was no significant increase in both arginine-free and rADI pretreated arginine-containing (citrulline) buffer. Subsequently, neither NMDA nor rADI alone caused cytotoxicity, whereas cotreatment with NMDA and rADI resulted in dissipation of the cell mitochondrial membrane potential and decreased cell viability. The mechanism of rADI cytotoxicity in the presence of NMDA is caused by the inhibition of NO production via nNOS mediated by the NMDA receptor, which was abolished when extracellular arginine was absent, even in the presence of citrulline. rADI not only reduced NO production but also caused cellular toxicity in nNOS-activated SH-SY5Y cells, suggesting a dual role for rADI in NOS-mediated neurotoxicity.

  3. Depletion of Arginine by Recombinant Arginine Deiminase Induces nNOS-Activated Neurotoxicity in Neuroblastoma Cells

    Directory of Open Access Journals (Sweden)

    Shan-Erh Lin

    2014-01-01

    Full Text Available The abnormal regulation of inducible nitric oxide synthase (iNOS and neuronal nitric oxide synthase (nNOS is associated with neurodegenerative disorders. Recombinant arginine deiminase (rADI is a selective NO modulator of iNOS and eNOS in endothelial cells, and it also exhibits neuroprotective activity in an iNOS-induced neuron-microglia coculture system. However, the effect of rADI on nNOS remains unknown. Addressing this issue is important for evaluating the potential application of rADI in neurodegenerative diseases. SH-SY5Y cells were treated with N-methyl-D-aspartic acid (NMDA to activate nNOS. NMDA increased NO production by 39.7 ± 3.9% via nNOS under arginine-containing conditions, but there was no significant increase in both arginine-free and rADI pretreated arginine-containing (citrulline buffer. Subsequently, neither NMDA nor rADI alone caused cytotoxicity, whereas cotreatment with NMDA and rADI resulted in dissipation of the cell mitochondrial membrane potential and decreased cell viability. The mechanism of rADI cytotoxicity in the presence of NMDA is caused by the inhibition of NO production via nNOS mediated by the NMDA receptor, which was abolished when extracellular arginine was absent, even in the presence of citrulline. rADI not only reduced NO production but also caused cellular toxicity in nNOS-activated SH-SY5Y cells, suggesting a dual role for rADI in NOS-mediated neurotoxicity.

  4. Synthesis, crystal structure and biological activity of novel diester cyclophanes

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Pengfei; Yang, Bingqin; Fang, Xianwen; Cheng, Zhao; Yang, Meipan, E-mail: yangbq@nwu.edu.cn [Department of Chemistry, Key Laboratory of Synthetic and Natural Functional Molecule Chemistry, Northwest University, Shaanxi (China)

    2012-10-15

    A series of novel diester cyclophanes was synthesized by esterification of 1,2-benzenedicarbonyl chloride with eight different diols under high dilution conditions. The structures of the compounds were verified by elemental analysis, {sup 1}H nuclear magnetic resonance (NMR), IR spectroscopy and high resolution mass spectrometry (HRMS). The crystal structures of two compounds were characterized by single crystal X-ray diffractometry (XRD). All the new cyclophanes were evaluated for biological activities and the results showed that some of these compounds have low antibacterial or antifungal activities (author)

  5. Established and emerging biological activity markers of inflammatory bowel disease

    DEFF Research Database (Denmark)

    Nielsen, O H; Vainer, B; Madsen, S M;

    2000-01-01

    Assessment of disease activity in inflammatory bowel disease (IBD), i.e., ulcerative colitis (UC) and Crohn's disease (CD), is done using clinical parameters and various biological disease markers. Ideally, a disease marker must: be able to identify individuals at risk of a given disorder......, be disease specific, mirror the disease activity and, finally, be easily applicable for routine clinical purposes. However, no such disease markers have yet been identified for IBD. In this article, classical disease markers including erythrocyte sedimentation rate, acute phase proteins (especially...

  6. Recombinant N-Terminal Slit2 Inhibits TGF-β-Induced Fibroblast Activation and Renal Fibrosis.

    Science.gov (United States)

    Yuen, Darren A; Huang, Yi-Wei; Liu, Guang-Ying; Patel, Sajedabanu; Fang, Fei; Zhou, Joyce; Thai, Kerri; Sidiqi, Ahmad; Szeto, Stephen G; Chan, Lauren; Lu, Mingliang; He, Xiaolin; John, Rohan; Gilbert, Richard E; Scholey, James W; Robinson, Lisa A

    2016-09-01

    Fibrosis and inflammation are closely intertwined injury pathways present in nearly all forms of CKD for which few safe and effective therapies exist. Slit glycoproteins signaling through Roundabout (Robo) receptors have been described to have anti-inflammatory effects through regulation of leukocyte cytoskeletal organization. Notably, cytoskeletal reorganization is also required for fibroblast responses to TGF-β Here, we examined whether Slit2 also controls TGF-β-induced renal fibrosis. In cultured renal fibroblasts, which we found to express Slit2 and Robo-1, the bioactive N-terminal fragment of Slit2 inhibited TGF-β-induced collagen synthesis, actin cytoskeletal reorganization, and Smad2/3 transcriptional activity, but the inactive C-terminal fragment of Slit2 did not. In mouse models of postischemic renal fibrosis and obstructive uropathy, treatment with N-terminal Slit2 before or after injury inhibited the development of renal fibrosis and preserved renal function, whereas the C-terminal Slit2 had no effect. Our data suggest that administration of recombinant Slit2 may be a new treatment strategy to arrest chronic injury progression after ischemic and obstructive renal insults by not only attenuating inflammation but also, directly inhibiting renal fibrosis.

  7. Early intracardiac thrombosis in preterm infants and thrombolysis with recombinant tissue type plasminogen activator

    Science.gov (United States)

    Ferrari, F; Vagnarelli, F; Gargano, G; Roversi, M; Biagioni, O; Ranzi, A; Cavazzuti, G

    2001-01-01

    OBJECTIVES—To determine the incidence of catheter related thrombosis and to test the efficacy of recombinant tissue type plasminogen activator (rt-PA) in preterm infants.
STUDY DESIGN—From January 1995 to December 1998, echocardiography was performed in the first few days of life in 76 very low birthweight (⩽ 1500 g) infants out of a total of 147 having an umbilical catheter placed. When intracardiac thrombosis was diagnosed, rt-PA infusion was performed.
RESULTS—Four infants (5%) developed an intracardiac thrombosis during the first few days of life. In three of them, rt-PA at a dose of 0.4-0.5 mg/kg in a 20-30 minute bolus led to dissolution of the clot. One patient received a three hour infusion after the bolus, at a dose of 0.1 mg/kg/h, with resolution of the thrombus. No systemic effects were observed after rt-PA infusion.
CONCLUSIONS—Early thrombosis may occur as a complication of umbilical catheterisation in preterm infants; early echocardiographic detection of this disorder allows complete, safe, and rapid lysis with rt-PA.

 PMID:11420328

  8. Evidence supporting the use of recombinant activated factor VII in congenital bleeding disorders

    Directory of Open Access Journals (Sweden)

    Pär I Johansson

    2010-06-01

    Full Text Available Pär I Johansson, Sisse R OstrowskiCapital Region Blood Bank, Section for Transfusion Medicine, Rigshospitalet, University of Copenhagen, Copenhagen, DenmarkBackground: Recombinant activated factor VII (rFVIIa, NovoSeven® was introduced in 1996 for the treatment of hemophilic patients with antibodies against coagulation factor VIII or IX.Objective: To review the evidence supporting the use of rFVIIa for the treatment of patients with congenital bleeding disorders.Patients and methods: English-language databases were searched in September 2009 for reports of randomized controlled trials (RCTs evaluating the ability of rFVIIa to restore hemostasis in patients with congenital bleeding disorders.Results: Eight RCTs involving 256 hemophilic patients with antibodies against coagulation factors, also known as inhibitors, were identified. The evidence supporting the use of rFVIIa in these patients was weak with regard to dose, clinical setting, mode of administration, efficacy, and adverse events, given the limited sample size of each RCT and the heterogeneity of the studies.Conclusion: The authors suggest that rFVIIa therapy in hemophilic patients with inhibitors should be based on the individual’s ability to generate thrombin and form a clot, and not on the patient’s weight alone. Therefore, assays for thrombin generation, such as whole-blood thromboelastography, have the potential to significantly improve the treatment of these patients.Keywords: hemophilia, inhibitors, coagulation factor VIII, coagulation factor IX, rFVIIa, NovoSeven, FEIBA, hemostasis, RCT

  9. Preparation of thermosensitive magnetic liposome encapsulated recombinant tissue plasminogen activator for targeted thrombolysis

    Science.gov (United States)

    Hsu, Hao-Lung; Chen, Jyh-Ping

    2017-04-01

    Recombinant tissue plasminogen activator (rtPA) was encapsulated in thermosensitive magnetic liposome (TML) prepared from 1,2-dipalmitoyl-sn-glycero-3-phosphocholine, distearolyphosphatidyl ethanolamine-N-poly(ethylene glycol) 2000, cholesterol and Fe3O4 magnetic nanoparticles by solvent evaporation/sonication and freeze-thaw cycles method. Response surface methodology was proved to be a powerful tool to predict the drug encapsulation efficiency and temperature-sensitive drug release. Validation experiments verified the accuracy of the model that provides a simple and effective method for fabricating TML with controllable encapsulation efficiency and predictable temperature-sensitive drug release behavior. The prepared samples were characterized for physico-chemical properties by dynamic light scattering, transmission electron microscopy, X-ray diffraction and differential scanning calorimetry. Temperature-sensitive release of rtPA could be confirmed from in vitro thrombolysis experiments. A thrombolytic drug delivery system using TML could be proposed for magnetic targeted delivery of rtPA to the site of thrombus followed by temperature-triggered controlled drug release in an alternating magnetic field.

  10. Recombinant human leptin attenuates stress axis activity in common carp (Cyprinus carpio L.).

    Science.gov (United States)

    Gorissen, Marnix; Bernier, Nicholas J; Manuel, Remy; de Gelder, Stefan; Metz, Juriaan R; Huising, Mark O; Flik, Gert

    2012-08-01

    Proper functioning of the endocrine stress axis requires communication between the stress axis and other regulatory mechanisms. We here describe an intimate interplay between the stress axis and recombinant human leptin (rhLeptin) in a teleostean fish, the common carp Cyprinus carpio. Restraint stress (by netting up to 96h) increased plasma cortisol but did not affect hepatic leptin expression. Perifusion of pituitary glands or head kidneys with rhLeptin revealed direct effects of rhLeptin on both tissues. RhLeptin suppresses basal and CRF-induced ACTH-secretion in a rapid and concentration-dependent manner. The rhLeptin effect persisted for over an hour after administration had been terminated. RhLeptin decreases basal interrenal cortisol secretion in vitro, and by doing so attenuates ACTH-stimulated cortisol production; rhLeptin does not affect interrenal ACTH-sensitivity. Our findings show that the endocrine stress axis activity and leptin are inseparably linked in a teleostean fish, a notion relevant to further our insights in the evolution of leptin physiology in vertebrates.

  11. Novel recombinant adeno-associated viruses for Cre activated and inactivated transgene expression in neurons

    Directory of Open Access Journals (Sweden)

    Arpiar eSaunders

    2012-07-01

    Full Text Available Understanding the organization of the nervous system requires methods for dissecting the contributions of each component cell type to circuit function. One widely used approach combines genetic targeting of Cre recombinase to specific cell populations with infection of recombinant adeno-associated viruses (rAAVs whose transgene expression is activated by Cre (Cre-On. Distinguishing how the Cre-expressing neurons differ functionally from neighboring Cre-negative neurons requires rAAVs that are inactivated by Cre (Cre-Off and can be used in tandem with Cre-On viruses. Here we introduce two rAAV vectors that are inactivated by Cre and carry different fluorophore and optogenetic constructs. We demonstrate single and dual rAAV systems to achieve Cre-On and Cre-Off expression in spatially-intermingled cell populations of the striatum. Using these systems, we uncovered cryptic genomic interactions that occur between multiple Cre-sensitive rAAVs or between Cre-sensitive rAAVs and somatic Cre-conditional alleles and devised methods to avoid these interactions. Our data highlight both important experimental caveats associated with Cre-dependent rAAV use as well as opportunities for the development of improved rAAVs for gene delivery.

  12. Recombinant expression and in vitro characterisation of active Huwentoxin-IV.

    Directory of Open Access Journals (Sweden)

    Isabelle Sermadiras

    Full Text Available Huwentoxin-IV (HwTx-IV is a 35-residue neurotoxin peptide with potential application as a novel analgesic. It is a member of the inhibitory cystine knot (ICK peptide family, characterised by a compact globular structure maintained by three intramolecular disulfide bonds. Here we describe a novel strategy for producing non-tagged, fully folded ICK-toxin in a bacterial system. HwTx-IV was expressed as a cleavable fusion to small ubiquitin-related modifier (SUMO in the cytoplasm of the SHuffle T7 Express lysY Escherichia coli strain, which allows cytosolic disulfide bond formation. Purification by IMAC with selective elution of monomeric SUMO fusion followed by proteolytic cleavage and polishing chromatographic steps yielded pure homogeneous toxin. Recombinant HwTx-IV is produced with a C-terminal acid, whereas the native peptide is C-terminally amidated. HwTx-IV(acid inhibited Nav1.7 in a dose dependent manner (IC50 = 463-727 nM. In comparison to HwTx-IV(amide (IC50 = 11 ± 3 nM, the carboxylate was ~50 fold less potent on Nav1.7, which highlights the impact of the C-terminus. As the amide bond of an additional amino acid may mimic the carboxamide, we expressed the glycine-extended analogue HwTx-IV(G36(acid in the SUMO/SHuffle system. The peptide was approximately three fold more potent on Nav1.7 in comparison to HwTx-IV(acid (IC50 = 190 nM. In conclusion, we have established a novel system for expression and purification of fully folded and active HwTx-IV(acid in bacteria, which could be applicable to other structurally complex and cysteine rich peptides. Furthermore, we discovered that glycine extension of HwTx-IV(acid restores some of the potency of the native carboxamide. This finding may also apply to other C-terminally amidated peptides produced recombinantly.

  13. The Chemistry and Biological Activities of Mimosine: A Review.

    Science.gov (United States)

    Nguyen, Binh Cao Quan; Tawata, Shinkichi

    2016-08-01

    Mimosine [β-[N-(3-hydroxy-4-oxypyridyl)]-α-aminopropionic acid] is a non-protein amino acid found in the members of Mimosoideae family. There are a considerable number of reports available on the chemistry, methods for estimation, biosynthesis, regulation, and degradation of this secondary metabolite. On the other hand, over the past years of active research, mimosine has been found to have various biological activities such as anti-cancer, antiinflammation, anti-fibrosis, anti-influenza, anti-virus, herbicidal and insecticidal activities, and others. Mimosine is a leading compound of interest for use in the development of RAC/CDC42-activated kinase 1 (PAK1)-specific inhibitors for the treatment of various diseases/disorders, because PAK1 is not essential for the growth of normal cells. Interestingly, the new roles of mimosine in malignant glioma treatment, regenerative dentistry, and phytoremediation are being emerged. These identified properties indicate an exciting future for this amino acid. The present review is focused on the chemistry and recognized biological activities of mimosine in an attempt to draw a link between these two characteristics. Copyright © 2016 John Wiley & Sons, Ltd.

  14. Biological Activities of Phenolic Compounds of Extra Virgin Olive Oil.

    Science.gov (United States)

    Servili, Maurizio; Sordini, Beatrice; Esposto, Sonia; Urbani, Stefania; Veneziani, Gianluca; Di Maio, Ilona; Selvaggini, Roberto; Taticchi, Agnese

    2013-12-20

    Over the last few decades, multiple biological properties, providing antioxidant, anti-inflammatory, chemopreventive and anti-cancer benefits, as well as the characteristic pungent and bitter taste, have been attributed to Extra Virgin Olive Oil (EVOO) phenols. In particular, growing efforts have been devoted to the study of the antioxidants of EVOO, due to their importance from health, biological and sensory points of view. Hydrophilic and lipophilic phenols represent the main antioxidants of EVOO, and they include a large variety of compounds. Among them, the most concentrated phenols are lignans and secoiridoids, with the latter found exclusively in the Oleaceae family, of which the drupe is the only edible fruit. In recent years, therefore, we have tackled the study of the main properties of phenols, including the relationships between their biological activity and the related chemical structure. This review, in fact, focuses on the phenolic compounds of EVOO, and, in particular, on their biological properties, sensory aspects and antioxidant capacity, with a particular emphasis on the extension of the product shelf-life.

  15. Biological Activities of Phenolic Compounds of Extra Virgin Olive Oil

    Directory of Open Access Journals (Sweden)

    Maurizio Servili

    2013-12-01

    Full Text Available Over the last few decades, multiple biological properties, providing antioxidant, anti-inflammatory, chemopreventive and anti-cancer benefits, as well as the characteristic pungent and bitter taste, have been attributed to Extra Virgin Olive Oil (EVOO phenols. In particular, growing efforts have been devoted to the study of the antioxidants of EVOO, due to their importance from health, biological and sensory points of view. Hydrophilic and lipophilic phenols represent the main antioxidants of EVOO, and they include a large variety of compounds. Among them, the most concentrated phenols are lignans and secoiridoids, with the latter found exclusively in the Oleaceae family, of which the drupe is the only edible fruit. In recent years, therefore, we have tackled the study of the main properties of phenols, including the relationships between their biological activity and the related chemical structure. This review, in fact, focuses on the phenolic compounds of EVOO, and, in particular, on their biological properties, sensory aspects and antioxidant capacity, with a particular emphasis on the extension of the product shelf-life.

  16. Biological function of activation-induced cytidine deaminase (AID

    Directory of Open Access Journals (Sweden)

    Ritu Kumar

    2014-10-01

    Full Text Available Activation-induced Cytidine Deaminase (AID is an essential regulator of B cell diversification, but its full range of action has until recently been an enigma. Based on homology, it was originally proposed to be an RNA-editing enzyme, but so far, no RNA substrates are known. Rather, it functions by deaminating cytidine, and in this manner, coupled with base-excision repair or mismatch repair machinery, it is a natural mutator. This allows it to play a central role in adaptive immunity, whereby it initiates the processes of class switch recombination and somatic hypermutation to help generate a diverse and high-affinity repertoire of immunoglobulin isotypes. More recently, it has been appreciated that methylated cytidine, already known as a key epigenetic mark on DNA controlling gene expression, can also be a target for AID modification. Coupled with repair machinery, this can facilitate the active removal of methylated DNA. This activity can impact the process of cellular reprogramming, including transition of a somatic cell to pluripotency, which requires major reshuffling of epigenetic memory. Thus, seemingly disparate roles for AID in controlling immune diversity and epigenetic memory have a common mechanistic basis. However, the very activity that is so useful for B cell diversity and cellular reprogramming is dangerous for the integrity of the genome. Thus, AID expression and activity is tightly regulated, and deregulation is associated with diseases including cancer. Here, we review the range of AID functions with a focus on its mechanisms of action and regulation. Major questions remain to be answered concerning how and when AID is targeted to specific loci and how this impacts development and disease.

  17. Biological function of activation-induced cytidine deaminase (AID).

    Science.gov (United States)

    Kumar, Ritu; DiMenna, Lauren J; Chaudhuri, Jayanta; Evans, Todd

    2014-01-01

    Activation-induced Cytidine Deaminase (AID) is an essential regulator of B cell diversification, but its full range of action has until recently been an enigma. Based on homology, it was originally proposed to be an RNA-editing enzyme, but so far, no RNA substrates are known. Rather, it functions by deaminating cytidine, and in this manner, coupled with base-excision repair or mismatch repair machinery, it is a natural mutator. This allows it to play a central role in adaptive immunity, whereby it initiates the processes of class switch recombination and somatic hypermutation to help generate a diverse and high-affinity repertoire of immunoglobulin isotypes. More recently, it has been appreciated that methylated cytidine, already known as a key epigenetic mark on DNA controlling gene expression, can also be a target for AID modification. Coupled with repair machinery, this can facilitate the active removal of methylated DNA. This activity can impact the process of cellular reprogramming, including transition of a somatic cell to pluripotency, which requires major reshuffling of epigenetic memory. Thus, seemingly disparate roles for AID in controlling immune diversity and epigenetic memory have a common mechanistic basis. However, the very activity that is so useful for B cell diversity and cellular reprogramming is dangerous for the integrity of the genome. Thus, AID expression and activity is tightly regulated, and deregulation is associated with diseases including cancer. Here, we review the range of AID functions with a focus on its mechanisms of action and regulation. Major questions remain to be answered concerning how and when AID is targeted to specific loci and how this impacts development and disease.

  18. The cell biology of T-dependent B cell activation

    DEFF Research Database (Denmark)

    Owens, T; Zeine, R

    1989-01-01

    The requirement that CD4+ helper T cells recognize antigen in association with class II Major Histocompatibility Complex (MHC) encoded molecules constrains T cells to activation through intercellular interaction. The cell biology of the interactions between CD4+ T cells and antigen-presenting cells...... includes multipoint intermolecular interactions that probably involve aggregation of both polymorphic and monomorphic T cell surface molecules. Such aggregations have been shown in vitro to markedly enhance and, in some cases, induce T cell activation. The production of T-derived lymphokines that have been...... implicated in B cell activation is dependent on the T cell receptor for antigen and its associated CD3 signalling complex. T-dependent help for B cell activation is therefore similarly MHC-restricted and involves T-B intercellular interaction. Recent reports that describe antigen-independent B cell...

  19. Biologically active traditional medicinal herbs from Balochistan, Pakistan.

    Science.gov (United States)

    Zaidi, Mudassir A; Crow, Sidney A

    2005-01-01

    The biological activities of the following four important medicinal plants of Balochistan, Pakistan were checked; Grewia erythraea Schwein f. (Tiliaceae), Hymenocrater sessilifolius Fisch. and C.A. Mey (Lamiaceae), Vincetoxicum stocksii Ali and Khatoon (Asclepiadaceae) and Zygophyllum fabago L. (Zygophyllaceae). The methanolic extracts were fractionated into hexane, ethyl acetate, chloroform, butanol and water. The antifungal and antibacterial activities of these plants were determined against 12 fungal and 12 bacterial strains by agar well diffusion and disk diffusion assays. The extract of Zygophyllum fabago was found to be highly effective against Candida albicans and Escherichia coli. The extract of Vincetoxicum stocksii was also found to be significantly active against Candida albicans, Bacillus subtilis and Bacillus cereus. Extracts of Hymenocrater sessilifolius and Grewia erythraea showed good activity only against Pseudomonas aeruginosa.

  20. [Influence of biological activated carbon dosage on landfill leachate treatment].

    Science.gov (United States)

    Cui, Yan-Rui; Guo, Yan; Wu, Qing

    2014-08-01

    Effects of biological activated carbon (BAC) dosage on COD removal in landfill leachate treatment were compared. The COD removal efficiency of reactors with 0, 100 and 300 g activated carbon dosage per litre activated sludge was 12.9%, 19.6% and 27.7%, respectively. The results indicated that BAC improved the refractory organic matter removal efficiency and there was a positive correlation between COD removal efficiency and BAC dosage. The output of carbon dioxide after 8h of aeration in reactors was 109, 193 and 306 mg corresponding to the activated carbon dosages mentioned above, which indicated the amount of biodegradation and BAC dosage also had a positive correlation. The combination of adsorption and bioregeneration of BAC resulted in the positive correlation betweem organic matter removal efficiency and BAC dosage, and bioregeneration was the root cause for the microbial decomposition of refractory organics.

  1. Biological activities of triazine derivatives. Combining DFT and QSAR results

    Directory of Open Access Journals (Sweden)

    Majdouline Larif

    2017-02-01

    Full Text Available In order to investigate the relationship between activities and structures, a 3D-QSAR study is applied to a set of 43 molecules based on triazines. This study was conducted using the principal component analysis (PCA method, the multiple linear regression method (MLR and the artificial neural network (ANN. The predicted values of activities are in good agreement with the experimental results. The artificial neural network (ANN techniques, considering the relevant descriptors obtained from the MLR, showed a correlation coefficient of 0.9 with an 8-3-1 ANN model which is a good result. As a result of quantitative structure–activity relationships, we found that the model proposed in this study is constituted of major descriptors used to describe these molecules. The obtained results suggested that the proposed combination of several calculated parameters could be useful to predict the biological activity of triazine derivatives.

  2. Biological activities of water-soluble fullerene derivatives

    Energy Technology Data Exchange (ETDEWEB)

    Nakamura, S; Mashino, T [Department of Pharmaceutical Sciences, Faculty of Pharmacy, Keio University, 1-5-30 Shiba-koen, Minato-ku, Tokyo 105-8512 (Japan)], E-mail: mashino-td@pha.keio.ac.jp

    2009-04-01

    Three types of water-soluble fullerene derivatives were synthesized and their biological activities were investigated. C{sub 60}-dimalonic acid, an anionic fullerene derivative, showed antioxidant activity such as quenching of superoxide and relief from growth inhibition of E. coli by paraquat. C{sub 60}-bis(7V,7V-dimethylpyrrolidinium iodide), a cationic fullerene derivative, has antibacterial activity and antiproliferative effect on cancer cell lines. The mechanism is suggested to be respiratory chain inhibition by reactive oxygen species produced by the cationic fullerene derivative. Proline-type fullerene derivatives showed strong inhibition activities on HIV-reverse transcriptase. The IC{sub 50} values were remarkably lower than nevirapine, a clinically used anti-HIV drug. Fullerene derivatives have a big potential for a new type of lead compound to be used as medicine.

  3. Production of barley endoprotease B2 in Pichia pastoris and its proteolytic activity against native and recombinant hordeins

    DEFF Research Database (Denmark)

    Rosenkilde, Anne Lind; Dionisio, Giuseppe; Holm, Preben Bach

    2014-01-01

    Barley (Hordeum vulgare L.) cysteine proteases are of fundamental biological importance during germination but may also have a large potential as commercial enzyme. Barley cysteine endoprotease B2 (HvEPB2) was expressed in Pichia pastoris from a pPICZαA based construct encoding a HvEPB2 C......-terminal truncated version (HvEPB2ΔC) and a proteolytic resistant His6 tag. Maximum yield was obtained after 4 days of induction. Recombinant HvEPB2ΔC (r-HvEPB2ΔC) was purified using a single step of Ni2+-affinity chromatography. Purified protein was evaluated by SDS–PAGE, Western blotting and activity assays...

  4. Activation and modulation of recombinantly expressed serotonin receptor type 3A by terpenes and pungent substances.

    Science.gov (United States)

    Ziemba, Paul M; Schreiner, Benjamin S P; Flegel, Caroline; Herbrechter, Robin; Stark, Timo D; Hofmann, Thomas; Hatt, Hanns; Werner, Markus; Gisselmann, Günter

    2015-11-27

    Serotonin receptor type 3 (5-HT3 receptor) is a ligand-gated ion channel that is expressed in the central nervous system (CNS) as well as in the peripheral nervous system (PNS). The receptor plays an important role in regulating peristalsis of the gastrointestinal tract and in functions such as emesis, cognition and anxiety. Therefore, a variety of pharmacologically active substances target the 5-HT3 receptor to treat chemotherapy-induced nausea and vomiting. The 5-HT3 receptors are activated, antagonized, or modulated by a wide range of chemically different substances, such as 2-methyl-serotonin, phenylbiguanide, setrones, or cannabinoids. Whereas the action of all of these substances is well described, less is known about the effect of terpenoids or fragrances on 5-HT3A receptors. In this study, we screened a large number of natural odorous and pungent substances for their pharmacological action on recombinantly expressed human 5-HT3A receptors. The receptors were functionally expressed in Xenopus oocytes and characterized by electrophysiological recordings using the two-electrode voltage-clamp technique. A screening of two odorous mixes containing a total of 200 substances revealed that the monoterpenes, thymol and carvacrol, act as both weak partial agonists and positive modulators on the 5-HT3A receptor. In contrast, the most effective blockers were the terpenes, citronellol and geraniol, as well as the pungent substances gingerol, capsaicin and polygodial. In our study, we identified new modulators of 5-HT3A receptors out of the classes of monoterpenes and vanilloid substances that frequently occur in various plants.

  5. Mutant p53: multiple mechanisms define biologic activity in cancer

    Directory of Open Access Journals (Sweden)

    Michael Paul Kim

    2015-11-01

    Full Text Available The functional importance of p53 as a tumor suppressor gene is evident through its pervasiveness in cancer biology. The p53 gene is the most commonly altered gene in human cancer; however, not all genetic alterations are biologically equivalent. The majority of p53 alterations involve missense mutations that result in the production of mutant p53 proteins. Such mutant p53 proteins lack normal p53 function and may acquire novel functions, often with deleterious effects. Here, we review characterized mechanisms of mutant p53 gain of function in multiple model systems. In addition, we review mutant p53 addiction as emerging evidence suggests that tumors may depend on sustained mutant p53 activity for continued growth. We also discuss the role of p53 in stromal elements and their contribution to tumor initiation and progression. Lastly, current genetic mouse models of mutant p53 are reviewed and their limitations discussed.

  6. Bone-inducing Activity of Biological Piezoelectric Ceramic

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    To simulate the piezoelectric effect of nature bone, two kinds of biological piezoelectric composite ceramics consisted of hydroxyapatite ( HA ) and lithium sodium potassium riobate (LNK) ceramic of which the ratio of HA/ LNK was 1: 10 and 5:5( wt/ wt ) were prepared. Their piezoelectric property and growth of apatite crystal in the ceramics surface were investigated. With the increase of LNK amount, piezoelectric activity increased correspondingly. By immersing the poled piezoelectric ceramics in simulated body fluid (SBF) at 36.5 ℃ for 7,14, and 21 days, apatite crystal was formed on negatively charged surfaces. After 21 days immersion in SBF,the thickest apatite crystal on the negatively charged surfaces increased to 3.337μm. The novel biological piezoelectric ceramics show an excellent piezoelectric property and superior potential bioactivity.

  7. Radiometric microbiologic assay for the biologically active forms of niacin

    Energy Technology Data Exchange (ETDEWEB)

    Kertcher, J.A.; Guilarte, T.R.; Chen, M.F.; Rider, A.A.; McIntyre, P.A.

    1979-05-01

    A radiometric microbiologic assay has been developed for the determination of niacin in biologic fluids. Lactobacillus plantarum produced /sup 14/CO/sub 2/ from L-(U-/sup 14/C) malic acid in quantities proportional to the amount of niacin present. The assay is specific for the biologically active forms of niacin in humans. Thirty normal hemolysates were analyzed and the values ranged from 13.0 to 17.8 ..mu..g niacin/ml RBC (mean = 15.27 +- 1.33 s.d.). Good recovery and reproducibility studies were obtained with this assay. On thirty blood samples, correlation was excellent between the radiometric and the conventional turbidimetric assays.

  8. Preparation and characterization of new biologically active polyurethane foams.

    Science.gov (United States)

    Savelyev, Yuri; Veselov, Vitali; Markovskaya, Ludmila; Savelyeva, Olga; Akhranovich, Elena; Galatenko, Natalya; Robota, Ludmila; Travinskaya, Tamara

    2014-12-01

    Biologically active polyurethane foams are the fast-developed alternative to many applications of biomedical materials. Due to the polyurethane structure features and foam technology it is possible to incorporate into their structure the biologically active compounds of target purpose via structural-chemical modification of macromolecule. A series of new biologically active polyurethane foams (PUFs) was synthesized with polyethers (MM 2500-5000), polyesters MM (500-2200), 2,4(2,6) toluene diisocyanate, water as a foaming agent, catalysts, foam stabilizers and functional compounds. Different functional compounds: 1,4-di-N-oxy-2,3-bis-(oxymethyl)-quinoxaline (DOMQ), partial sodium salt of poly(acrylic acid) and 2,6-dimethyl-N,N-diethyl aminoacetatanilide hydrochloride were incorporated into the polymer structure/composition due to the chemical and/or physical bonding. Structural peculiarities of PUFs were studied by FTIR spectroscopy and X-ray scattering. Self-adhesion properties of PUFs were estimated by measuring of tensile strength at break of adhesive junction. The optical microscopy method was performed for the PUF morphology studies. Toxicological estimation of the PUFs was carried out in vitro and in vivo. The antibacterial action towards the Gram-positive and Gram-negative bacteria (Escherichia coli ATC 25922, E. coli ATC 2150, Klebsiella pneumoniae 6447, Staphylococcus aureus 180, Pseudomonas aeruginosa 8180, Proteus mirabilis F 403, P. mirabilis 6054, and Proteus vulgaris 8718) was studied by the disc method on the solid nutrient. Physic-chemical properties of the PUFs (density, tensile strength and elongation at break, water absorption and vapor permeability) showed that all studied PUFs are within the operational requirements for such materials and represent fine-cellular foams. Spectral studies confirmed the incorporation of DOMQ into the PUF's macrochain. PUFs are characterized by microheterogeneous structure. They are antibacterially active, non

  9. Soil biological activity at European scale - two calculation concepts

    Science.gov (United States)

    Krüger, Janine; Rühlmann, Jörg

    2014-05-01

    The CATCH-C project aims to identify and improve the farm-compatibility of Soil Management Practices including to promote productivity, climate change mitigation and soil quality. The focus of this work concentrates on turnover conditions for soil organic matter (SOM). SOM is fundamental for the maintenance of quality and functions of soils while SOM storage is attributed a great importance in terms of climate change mitigation. The turnover conditions depend on soil biological activity characterized by climate and soil properties. To assess the turnover conditions two model concepts are applied: (I) Biological active time (BAT) regression approach derived from CANDY model (Franko & Oelschlägel 1995) expresses the variation of air temperature, precipitation and soil texture as a timescale and an indicator of biological activity for soil organic matter (SOM) turnover. (II) Re_clim parameter within the Introductory Carbon Balance Model (Andrén & Kätterer 1997) states the soil temperature and soil water to estimate soil biological activity. The modelling includes two strategies to cover the European scale and conditions. BAT was calculated on a 20x20 km grid basis. The European data sets of precipitation and air temperature (time period 1901-2000, monthly resolution), (Mitchell et al. 2004) were used to derive long-term averages. As we focus on agricultural areas we included CORINE data (2006) to extract arable land. The resulting BATs under co-consideration of the main soil textures (clay, silt, sand and loam) were investigated per environmental zone (ENZs, Metzger et al. 2005) that represents similar conditions for precipitation, temperature and relief to identify BAT ranges and hence turnover conditions for each ENZ. Re_clim was quantified by climatic time series of more than 250 weather stations across Europe presented by Klein Tank et al. (2002). Daily temperature, precipitation and potential evapotranspiration (maximal thermal extent) were used to calculate

  10. Biological activity of trisporoids and trisporoid analogues in Mucor mucedo (-).

    Science.gov (United States)

    Schachtschabel, Doreen; Schimek, Christine; Wöstemeyer, Johannes; Boland, Wilhelm

    2005-06-01

    In the course of their sexual interactions, zygomycete fungi communicate via an elaborate series of carotene-derived compounds, namely trisporic acid and its biosynthetic progenitors. A novel building-block strategy allowed the systematic generation of structurally modified trisporoids along with putative early biosynthetic precursors for physiological tests. The impact of discrete structural elements was documented by the ability of individual compounds to induce sexually committed hyphae in Mucor mucedo. The activity screening contributed to establish general structure-function relationships for trisporoid action. Most crucial for activity were the dimension of the longer side chain, the polarity of functional groups at C(4) and C(13), and the number of conjugated double bonds in the side chain. The presence of an oxygen substituent at the cyclohexene ring is not essential for function. The overall biological activity apparently results from the combination of the various structural elements.

  11. Hydrodynamic collective effects of active proteins in biological membranes

    Science.gov (United States)

    Koyano, Yuki; Kitahata, Hiroyuki; Mikhailov, Alexander S.

    2016-08-01

    Lipid bilayers forming biological membranes are known to behave as viscous two-dimensional fluids on submicrometer scales; usually they contain a large number of active protein inclusions. Recently, it was shown [A. S. Mikhailov and R. Kapral, Proc. Natl. Acad. Sci. USA 112, E3639 (2015), 10.1073/pnas.1506825112] that such active proteins should induce nonthermal fluctuating lipid flows leading to diffusion enhancement and chemotaxislike drift for passive inclusions in biomembranes. Here, a detailed analytical and numerical investigation of such effects is performed. The attention is focused on the situations when proteins are concentrated within lipid rafts. We demonstrate that passive particles tend to become attracted by active rafts and are accumulated inside them.

  12. Chemical Analysis and Biological Activity of Jordanian Chamomile Extracts

    Directory of Open Access Journals (Sweden)

    Nawal Hassan Al Bahtiti

    2012-02-01

    Full Text Available The Jordanian chamomile (Matricaria chamomilla has been researched more thoroughly to evaluate its useful properties. It is investigated and found that Jordanian chamomile is rich in phenolic compounds, with beneficial biological activities. By applying the most promising HPLC method, the content of total phenolics in methanolic extract was determined according to the Folin-Clocalteu procedure, and was found (GAE>20 mg/g. The flavonoid types were found as flavones and flavonolos.The minimum inhibitory concentration values for methanolic extracts of Jordanian chamomile were determined for different kinds of bacteria. The extracts have activity against Staphylococcus aurous, candida albicans, Esherichia Coli, Betula pubescens and Pinus sylvestris. The activity has been observed to be due to the tannins and a pigenin present in the extract. To utilize these significant sources of natural compounds, further characterization of phenolic composition is needed.

  13. Established and emerging biological activity markers of inflammatory bowel disease

    DEFF Research Database (Denmark)

    Nielsen, O H; Vainer, B; Madsen, S M;

    2000-01-01

    Assessment of disease activity in inflammatory bowel disease (IBD), i.e., ulcerative colitis (UC) and Crohn's disease (CD), is done using clinical parameters and various biological disease markers. Ideally, a disease marker must: be able to identify individuals at risk of a given disorder...... molecules. It is concluded that none of the pertinent laboratory surrogate markers of disease activity in IBD are specific or sensitive enough to replace basic clinical observation such as the number of daily bowel movements, general well-being, and other parameters in parallel. Further studies are highly...... warranted to identify and assess the clinical importance and applicability of new laboratory markers for the diagnosis or the disease activity of IBD....

  14. Circumventing furin enhances factor VIII biological activity and ameliorates bleeding phenotypes in hemophilia models

    Science.gov (United States)

    Siner, Joshua I.; Samelson-Jones, Benjamin J.; Crudele, Julie M.; French, Robert A.; Lee, Benjamin J.; Zhou, Shanzhen; Merricks, Elizabeth; Raymer, Robin; Camire, Rodney M.; Arruda, Valder R.

    2016-01-01

    Processing by the proprotein convertase furin is believed to be critical for the biological activity of multiple proteins involved in hemostasis, including coagulation factor VIII (FVIII). This belief prompted the retention of the furin recognition motif (amino acids 1645–1648) in the design of B-domain–deleted FVIII (FVIII-BDD) products in current clinical use and in the drug development pipeline, as well as in experimental FVIII gene therapy strategies. Here, we report that processing by furin is in fact deleterious to FVIII-BDD secretion and procoagulant activity. Inhibition of furin increases the secretion and decreases the intracellular retention of FVIII-BDD protein in mammalian cells. Our new variant (FVIII-ΔF), in which this recognition motif is removed, efficiently circumvents furin. FVIII-ΔF demonstrates increased recombinant protein yields, enhanced clotting activity, and higher circulating FVIII levels after adeno-associated viral vector–based liver gene therapy in a murine model of severe hemophilia A (HA) compared with FVIII-BDD. Moreover, we observed an amelioration of the bleeding phenotype in severe HA dogs with sustained therapeutic FVIII levels after FVIII-ΔF gene therapy at a lower vector dose than previously employed in this model. The immunogenicity of FVIII-ΔF did not differ from that of FVIII-BDD as a protein or a gene therapeutic. Thus, contrary to previous suppositions, FVIII variants that can avoid furin processing are likely to have enhanced translational potential for HA therapy. PMID:27734034

  15. Pereskia aculeata Muller (Cactaceae Leaves: Chemical Composition and Biological Activities

    Directory of Open Access Journals (Sweden)

    Lucèia Fàtima Souza

    2016-09-01

    Full Text Available The aims of this work were to study the chemical composition of the essential oil from the leaves of Pereskia aculeata and to evaluate some biological activities of three leaf extracts. The phenolic content, antioxidant activity, and in vitro antimicrobial and antifungal activities were determined. The methanol extract showed antioxidant activity (EC50 7.09 mg/mL and high polyphenols content (15.04 ± 0.31 mg gallic acid equivalents (GAE/g. The petroleum ether extract exhibited potent antibacterial activity against Escherichia coli, whereas the chloroform extract showed inhibitory activity against Bacillus cereus and Staphylococcus aureus. The petroleum ether and methanol extracts were more effective in inhibiting the growth of Aspergillus versicolor. The possible cytotoxicity of extracts on neuroblastoma SH-SY5Y cancer cell line and the influence on adenylate cyclase (ADCY expression was also studied. P. aculeata chloroform extract showed antiproliferative activity with an IC50 value of 262.83 µg/mL. Treatments of SH-SY5Y neuroblastoma cells with 100 µg/mL of methanol extract significantly reduced ADCY1 expression.

  16. Pereskia aculeata Muller (Cactaceae) Leaves: Chemical Composition and Biological Activities.

    Science.gov (United States)

    Souza, Lucèia Fàtima; Caputo, Lucia; Inchausti De Barros, Ingrid Bergman; Fratianni, Florinda; Nazzaro, Filomena; De Feo, Vincenzo

    2016-09-03

    The aims of this work were to study the chemical composition of the essential oil from the leaves of Pereskia aculeata and to evaluate some biological activities of three leaf extracts. The phenolic content, antioxidant activity, and in vitro antimicrobial and antifungal activities were determined. The methanol extract showed antioxidant activity (EC50 7.09 mg/mL) and high polyphenols content (15.04 ± 0.31 mg gallic acid equivalents (GAE)/g). The petroleum ether extract exhibited potent antibacterial activity against Escherichia coli, whereas the chloroform extract showed inhibitory activity against Bacillus cereus and Staphylococcus aureus. The petroleum ether and methanol extracts were more effective in inhibiting the growth of Aspergillus versicolor. The possible cytotoxicity of extracts on neuroblastoma SH-SY5Y cancer cell line and the influence on adenylate cyclase (ADCY) expression was also studied. P. aculeata chloroform extract showed antiproliferative activity with an IC50 value of 262.83 µg/mL. Treatments of SH-SY5Y neuroblastoma cells with 100 µg/mL of methanol extract significantly reduced ADCY1 expression.

  17. Human recombinant truncated RNASET2, devoid of RNase activity; A potential cancer therapeutic agent

    Science.gov (United States)

    Nesiel-Nuttman, Liron; Schwartz, Betty; Shoseyov, Oded

    2014-01-01

    Human RNASET2 has been implicated in antitumorigenic and antiangiogenic activities, independent of its ribonuclease capacities. We constructed a truncated version of human RNASET2, starting at E50 (trT2-50) and devoid of ribonuclease activity. trT2-50 maintained its ability to bind actin and to inhibit angiogenesis and tumorigenesis. trT2-50 binds to cell surface actin and formed a complex with actin in vitro. The antiangiogenic effect of this protein was demonstrated in human umbilical vein endothelial cells (HUVECs) by its ability to arrest tube formation on Matrigel, induced by angiogenic factors. Immunofluorescence staining of HUVECs showed nuclear and cytosolic RNASET2 protein that was no longer detectable inside the cell following trT2-50 treatment. This effect was associated with disruption of the intracellular actin network. trT2-50 co-localized with angiogenin, suggesting that both molecules bind (or compete) for similar cellular epitopes. Moreover, trT2-50 led to a significant inhibition of tumor development. Histological analysis demonstrated abundant necrotic tissue and a substantial loss of endothelial structure in trT2-50-treated tumors. Collectively, the present results indicate that trT2-50, a molecule engineered to be deficient of its catalytic activity, still maintained its actin binding and anticancer-related biological activities. We therefore suggest that trT2-50 may serve as a potential cancer therapeutic agent. PMID:25426551

  18. Activating and inhibiting connections in biological network dynamics

    Directory of Open Access Journals (Sweden)

    Knight Rob

    2008-12-01

    Full Text Available Abstract Background Many studies of biochemical networks have analyzed network topology. Such work has suggested that specific types of network wiring may increase network robustness and therefore confer a selective advantage. However, knowledge of network topology does not allow one to predict network dynamical behavior – for example, whether deleting a protein from a signaling network would maintain the network's dynamical behavior, or induce oscillations or chaos. Results Here we report that the balance between activating and inhibiting connections is important in determining whether network dynamics reach steady state or oscillate. We use a simple dynamical model of a network of interacting genes or proteins. Using the model, we study random networks, networks selected for robust dynamics, and examples of biological network topologies. The fraction of activating connections influences whether the network dynamics reach steady state or oscillate. Conclusion The activating fraction may predispose a network to oscillate or reach steady state, and neutral evolution or selection of this parameter may affect the behavior of biological networks. This principle may unify the dynamics of a wide range of cellular networks. Reviewers Reviewed by Sergei Maslov, Eugene Koonin, and Yu (Brandon Xia (nominated by Mark Gerstein. For the full reviews, please go to the Reviewers' comments section.

  19. Synthesis and biological activities of turkesterone 11?-acyl derivatives

    Directory of Open Access Journals (Sweden)

    Laurence Dinan

    2003-02-01

    Full Text Available Turkesterone is a phytoecdysteroid possessing an 11alpha-hydroxyl group. It is an analogue of the insect steroid hormone 20-hydroxyecdysone. Previous ecdysteroid QSAR and molecular modelling studies predicted that the cavity of the ligand-binding domain of the ecdysteroid receptor would possess space in the vicinity of C-11/C-12 of the ecdysteroid. We report the regioselective synthesis of a series of turkesterone 11alpha-acyl derivatives in order to explore this possibility. The structures of the analogues have been unambiguously determined by spectroscopic means (NMR and low-resolution mass spectrometry. Purity was verified by HPLC. Biological activities have been determined in Drosophila melanogaster BII cell-based bioassay for ecdysteroid agonists and in an in vitro radioligand-displacement assay using bacterially expressed D. melanogaster EcR/USP receptor proteins. The 11alpha-acyl derivatives do retain a significant amount of biological activity relative to the parent ecdysteroid. Further, although activity initially drops with the extension of the acyl chain length (C2 to C4, it then increases (C6 to C10, before decreasing again (C14 and C20. The implications of these findings for the interaction of ecdysteroids with the ecdysteroid receptor and potential applications in the generation of affinity-labelled and fluorescently-tagged ecdysteroids are discussed.

  20. DAGESTAN HAWTHORN - A VALUABLE SOURCE OF BIOLOGICALLY ACTIVE SUBSTANCES

    Directory of Open Access Journals (Sweden)

    Omarieva L. V.

    2016-02-01

    Full Text Available The article deals with hawthorn Dagestan. It is noted that the more than 100 species are found 14 species in Dagestan. The most common of them are studied as a source of biologically active substances and nutrients, and they can be used as food additives for various purposes. The content of biologically active substances and nutrients are defined. The high content of phenolic substances in C. pentagyna Waldst. et Kit, which play an important role in the formation of immunity and possessing P-vitamin activity is defined. Amino acid and fatty acid composition of the fruit of the hawthorn Crataegus pentagyna Waldst. et Kit, as the most valuable for using as food additives is defined by ionexchange and gas-liquid chromatography. Sixteen amino acids were identified and quantified in the protein component of hawthorn fruit. The high content of essential amino acids such as lysine, threonine, valine, methionine, leucine, isoleucine, phenylalanine. The amount of essential amino acids is 296.1 mg / 100 g. Chromatographic separation of fatty acid methyl esters has enabled us to establish the presence of the following fatty acids: palmitic, oleic, stearic, linoleic and linolenic, including established the presence of essential fatty acids - the family of alpha-linolenic acid OMEGA -3 and linoleic acid family of OMEGA -6

  1. Isolation of biologically-active exosomes from human plasma.

    Science.gov (United States)

    Muller, Laurent; Hong, Chang-Sook; Stolz, Donna B; Watkins, Simon C; Whiteside, Theresa L

    2014-09-01

    Effects of exosomes present in human plasma on immune cells have not been examined in detail. Immunological studies with plasma-derived exosomes require their isolation by procedures involving ultracentrifugation. These procedures were largely developed using supernatants of cultured cells. To test biologic activities of plasma-derived exosomes, methods are necessary that ensure adequate recovery of exosome fractions free of contaminating larger vesicles, cell fragments and protein/nucleic acid aggregates. Here, an optimized method for exosome isolation from human plasma/serum specimens of normal controls (NC) or cancer patients and its advantages and pitfalls are described. To remove undesirable plasma-contaminating components, ultrafiltration of differentially-centrifuged plasma/serum followed by size-exclusion chromatography prior to ultracentrifugation facilitated the removal of contaminants. Plasma or serum was equally acceptable as a source of exosomes based on the recovered protein levels (in μg protein/mL plasma) and TEM image quality. Centrifugation on sucrose density gradients led to large exosome losses. Fresh plasma was the best source of morphologically-intact exosomes, while the use of frozen/thawed plasma decreased exosome purity but not their biologic activity. Treatments of frozen plasma with DNAse, RNAse or hyaluronidase did not improve exosome purity and are not recommended. Cancer patients' plasma consistently yielded more isolated exosomes than did NCs' plasma. Cancer patients' exosomes also mediated higher immune suppression as evidenced by decreased CD69 expression on responder CD4+ T effector cells. Thus, the described procedure yields biologically-active, morphologically-intact exosomes that have reasonably good purity without large protein losses and can be used for immunological, biomarker and other studies.

  2. From gene amplification to V(D)J recombination and back: a personal account of my early years in B cell biology.

    Science.gov (United States)

    Alt, Frederick W

    2007-11-01

    I have been invited to write a short historical feature in the context of being a co-recipient with Klaus Rajewsky and Fritz Melchers of the 2007 Novartis Prize in Basic Immunology that was given in the general area of the molecular biology of B cells. In this feature, I cover the main points of the short talk that I presented at the Award Ceremony at the International Immunology Congress in Rio de Janeiro, Brazil. This talk focused primarily on the work and people involved early on in generating the models and ideas that have formed the basis for my ongoing efforts in the areas of V(D)J recombination and B cell development.

  3. Applications of Recombinant DNA Technology in Gastrointestinal Medicine and Hepatology: Basic Paradigms of Molecular Cell Biology. Part A: Eukaryotic Gene Structure and DNA Replication

    Directory of Open Access Journals (Sweden)

    Gary E Wild

    2000-01-01

    Full Text Available Progress in the basic sciences of cell and molecular biology has provided an exciting dimension that has translated into clinically relevant information in every medical subspecialty. Importantly, the application of recombinant DNA technology has played a major role in unravelling the intricacies related to the molecular pathophysiology of disease. This series of review articles constitutes a framework for the integration of the database of new information into the core knowledge base of concepts related to the pathogenesis of gastrointestinal disorders and liver disease. The goal of this series of three articles is to review the basic principles of eukaryotic gene expression. The first article examines the role of DNA in directing the flow of genetic information in eukaryotic cells.

  4. Fusion expression of pedA gone to obtain biologically active pediocin PA-1 in Escherichia coli

    Institute of Scientific and Technical Information of China (English)

    Shan-na LIU; Ye HAN; Zhi-jiang ZHOU

    2011-01-01

    Two heterologous expression systems using thioredoxin (trxA) as a gene fusion part in Eschenchia coli weredeveloped to produce recombinant pediocin PA-1. Pediocin PA-1 structural gene pedA was isolated from Pediococcusacidi/actici PA003 by the method of polymerase chain reaction (PCR), then cloned into vector pET32a(+), and expressedas thioredoxin-PedA fusion protein in the host strain E. coil BL21 (DE3). The fusion protein was in the form of inclusionbody and was refolded before purification by nickel-iminodiacetic acid (Ni-IDA) agarose resin column. Biological activity ofrecombinant pediocin PA-1 was analyzed after cleavage of the fusion protein by enterokinase. Agar diffusion test re-vealed that 512-arbitrary unit (AU) recombinant pediocin PA-1 was obtained from 1 ml culture medium of E. coli(pPA003PED1) using Listeda monocytogenes as the indicator strain. Thioredoxin-PedA fusion gene was further clonedinto pET20b(+). Thioredoxin-PedA fusion protein was detected in both the periplasmic and cytoplasmic spaces. Therecombinant pediocin PA-1 from the soluble fraction attained 384 AU from 1 ml culture medium of E. coli (pPA003PED2).Therefore, biologically active pediocin PA-1 could be obtained by these two hybrid gene expression methods.

  5. Functionalization of hydroxyl terminated polybutadiene with biologically active fluorescent molecule

    Indian Academy of Sciences (India)

    R Murali Sankar; Subhadeep Saha; K Seeni Meera; Tushar Jana

    2009-10-01

    A biologically active molecule, 2-chloro-4,6-bis(dimethylamino)-1,3,5-triazine (CBDT), has been covalently attached at the terminal carbon atoms of the hydroxyl terminated polybutadiene (HTPB) backbone. The modification of HTPB backbone by CBDT molecule does not affect the unique physico-chemical properties such as fluidity, hydroxyl value and microstructure of the parent HTPB. The formation of hydrogen bonding between the terminal hydroxyl groups and the nitrogen atoms of triazine moiety is the driving force for the terminal attachment chemistry. The functionalized HTPB (HTPB–CBDT) shows a strong fluorescence emission at 385 nm.

  6. A Conceptual Framework for Organizing Active Learning Experiences in Biology Instruction

    Science.gov (United States)

    Gardner, Joel; Belland, Brian R.

    2012-08-01

    Introductory biology courses form a cornerstone of undergraduate instruction. However, the predominantly used lecture approach fails to produce higher-order biology learning. Research shows that active learning strategies can increase student learning, yet few biology instructors use all identified active learning strategies. In this paper, we present a framework to design biology instruction that incorporates all active learning strategies. We review active learning research in undergraduate biology courses, present a framework for organizing active learning strategies, and provide clear implications and future research for designing instruction in introductory undergraduate biology courses.

  7. Elucidating GPR Response to Biological Activity: Field and Laboratory Experiments

    Science.gov (United States)

    Tsoflias, G. P.; Schillig, P. C.; McGlashan, M. A.; Roberts, J. A.; Devlin, J. F.

    2010-12-01

    Recent studies of the geophysical signatures of biological processes in earth environments have resulted in the emergent field of “biogeophysics”. The ability to monitor remotely and to quantify active biological processes in the subsurface can have transformative implications to a wide range of investigations, including the bioremediation of contaminated sites. Previous studies have demonstrated that ground-penetrating radar (GPR) can be used to detect the products of microbial activity in the subsurface, such as changes in bulk electrical conductivity, mineral dissolution and precipitation, and the formation of biogenic gas. We present field and laboratory experiments that offer insights to the response of GPR signals to microbial activity. In the field, time-lapse borehole radar tomography was used to monitor biodegradation of a hydrocarbon plume over a period of two years. A dense grid of fourteen borehole pairs monitoring the bioactive region showed radar wave velocity changes of +/-4% and signal attenuation changes of +/-25%. These GPR observations correlated spatially and temporally to independent measurements of groundwater velocity and geochemical variations that occurred in response to microbial activity. The greatest relative changes in radar wave velocity of propagation and attenuation were observed in the region of enhanced bacterial stimulation where biomass growth was the greatest. Radar wave velocity and attenuation decreased during periods of enhanced biostimulation. Two laboratory experiments were conducted to further assess radar response to biomass growth. The first experiment monitored GPR wave transmission through a water-saturated quartz-sand reactor during the course of enhanced biostimulation. Radar wave velocity initially decreased as a result of bacterial activity and subsequently increased rapidly as biogenic gas formed in the pore space. Radar signal attenuation increased during the course of the experiment as a result of an

  8. Influence of Different Types of Recombination Active Defects on the Integral Electrical Properties of Multicrystalline Silicon Solar Cells

    Directory of Open Access Journals (Sweden)

    Dominik Lausch

    2015-01-01

    Full Text Available In this contribution the influence of different types of recombination-active defects on the integral electrical properties of multicrystalline Si solar cells is investigated. Based on a previous classification scheme related to the luminescence behavior of crystal defects, Type-A and Type-B defects are locally distinguished. It is shown that Type-A defects, correlated to iron contaminations, are dominating the efficiency by more than 20% relative through their impact on the short circuit current ISC and open circuit voltage VOC in standard Si material (only limited by recombination active crystal defects. Contrarily, Type-B defects show low influence on the efficiency of 3% relative. The impact of the detrimental Type-A defects on the electrical parameters is studied as a function of the block height. A clear correlation between the area fraction of Type-A defects and both the global Isc and the prebreakdown behavior (reverse current in voltage regime-2 (−11 V is observed. An outlier having an increased full-area recombination activity is traced back to dense inter- and intragrain nucleation of Fe precipitates. Based on these results it is concluded that Type-A defects are the most detrimental defects in Si solar cells (having efficiencies > 15% and have to be prevented by optimized Si material quality and solar cell process conditions.

  9. Highly selective anti-Prelog synthesis of optically active aryl alcohols by recombinant Escherichia coli expressing stereospecific alcohol dehydrogenase.

    Science.gov (United States)

    Li, Ming; Nie, Yao; Mu, Xiao Qing; Zhang, Rongzhen; Xu, Yan

    2016-07-03

    Biocatalytic asymmetric synthesis has been widely used for preparation of optically active chiral alcohols as the important intermediates and precursors of active pharmaceutical ingredients. However, the available whole-cell system involving anti-Prelog specific alcohol dehydrogenase is yet limited. A recombinant Escherichia coli system expressing anti-Prelog stereospecific alcohol dehydrogenase from Candida parapsilosis was established as a whole-cell system for catalyzing asymmetric reduction of aryl ketones to anti-Prelog configured alcohols. Using 2-hydroxyacetophenone as the substrate, reaction factors including pH, cell status, and substrate concentration had obvious impacts on the outcome of whole-cell biocatalysis, and xylose was found to be an available auxiliary substrate for intracellular cofactor regeneration, by which (S)-1-phenyl-1,2-ethanediol was achieved with an optical purity of 97%e.e. and yield of 89% under the substrate concentration of 5 g/L. Additionally, the feasibility of the recombinant cells toward different aryl ketones was investigated, and most of the corresponding chiral alcohol products were obtained with an optical purity over 95%e.e. Therefore, the whole-cell system involving recombinant stereospecific alcohol dehydrogenase was constructed as an efficient biocatalyst for highly enantioselective anti-Prelog synthesis of optically active aryl alcohols and would be promising in the pharmaceutical industry.

  10. The ATPase activity of Fml1 is essential for its roles in homologous recombination and DNA repair

    Science.gov (United States)

    Nandi, Saikat; Whitby, Matthew C.

    2012-01-01

    In fission yeast, the DNA helicase Fml1, which is an orthologue of human FANCM, is a key component of the machinery that drives and governs homologous recombination (HR). During the repair of DNA double-strand breaks by HR, it limits the occurrence of potentially deleterious crossover recombinants, whereas at stalled replication forks, it promotes HR to aid their recovery. Here, we have mutated conserved residues in Fml1’s Walker A (K99R) and Walker B (D196N) motifs to determine whether its activities are dependent on its ability to hydrolyse ATP. Both Fml1K99R and Fml1D196N are proficient for DNA binding but totally deficient in DNA unwinding and ATP hydrolysis. In vivo both mutants exhibit a similar reduction in recombination at blocked replication forks as a fml1Δ mutant indicating that Fml1’s motor activity, fuelled by ATP hydrolysis, is essential for its pro-recombinogenic role. Intriguingly, both fml1K99R and fml1D196N mutants exhibit greater sensitivity to genotoxins and higher levels of crossing over during DSB repair than a fml1Δ strain. These data suggest that without its motor activity, the binding of Fml1 to its DNA substrate can impede alternative mechanisms of repair and crossover avoidance. PMID:22844101

  11. A meta-analysis of controlled trials of recombinant human activated protein C therapy in patients with sepsis

    Directory of Open Access Journals (Sweden)

    Wiedermann Christian J

    2005-10-01

    Full Text Available Abstract Background Meta-analysis of two randomised controlled trials in severe sepsis performed with recombinant human activated protein C may provide further insight as to the therapeutic utility of targeting the clotting cascade in this syndrome. Methods In search for relevant studies published, two randomized clinical trials were found eligible. Results The studies, PROWESS and ADDRESS, enrolled a total of 4329 patients with risk ratio (RR and 95% confidence interval (CI data for effect on 28-day mortality relative to control treatment of 0.92 (0.83–1.02 suggesting that recombinant human activated protein C is not beneficial in severe sepsis. In PROWESS, 873 of 1690 patients presented with low risk, and 2315 of 2639 patients in ADDRESS as defined by APACHE II score Conclusion This meta-analysis, therefore, raises doubts about the clinical usefulness of recombinant activated protein C in patients with severe sepsis and an APACHE II score ≥ 25 which can only be resolved by another properly designed clinical trial.

  12. The OECD Blue Book on Recombinant DNA Safety Considerations: it's influence on ISBR and EFSA activities.

    Science.gov (United States)

    Schiemann, Joachim

    2006-01-01

    Biosafety regulatory frameworks are intended to serve as mechanisms for ensuring the safe use of biotechnology products without imposing unacceptable risk to human health or the environment, or unintended constraints to technology transfer. The OECD Blue Book on "Recombinant DNA Safety Considerations", setting out principles and concepts for handling genetically modified organisms safely outside of contained laboratory conditions, was a milestone in the history of biotechnology. The "Recombinant DNA Safety Considerations" definitively became the major resource for the formulation of national regulatory frameworks and international regulations, including the Cartagena Protocol.

  13. Evaluation of recombinant activated protein C for severe sepsis at a tertiary academic medical center

    Directory of Open Access Journals (Sweden)

    Anger KE

    2013-06-01

    Full Text Available Kevin E Anger,1 Jeremy R DeGrado,1 Bonnie C Greenwood,1 Steven A Cohen,2 Paul M Szumita1 1Department of Pharmacy, Brigham and Women’s Hospital, Boston, MA, USA; 2Department of Family Medicine and Population Health, Division of Epidemiology, Virginia Commonwealth University, Richmond, VA, USA Purpose: Early clinical trials of recombinant human activated protein C (rhAPC for severe sepsis excluded patients at high risk of bleeding. Recent literature suggests bleeding rates are higher in clinical practice and may be associated with worsened outcomes. Our objective was to evaluate baseline demographics; incidence, and risk factors for major bleeding; and mortality of patients receiving rhAPC for severe sepsis at our institution. Methods: A retrospective study was performed for all patients receiving rhAPC for treatment of severe sepsis at a tertiary academic medical center from January 2002 to June 2009. Demographic information, clinical variables, intensive care unit, and hospital outcomes were recorded. Results: Of the 156 patients that received rhAPC, 54 (34.6% did not meet institutional criteria for safe use at baseline due to bleeding precaution or contraindication. Twenty-three (14.7% patients experienced a major bleeding event. Multivariate analysis demonstrated baseline International Normalized Ratio ≥2.5 (odds ratio [OR] 3.68, 95% confidence interval [CI]: 1.28–10.56; P = 0.03 and platelet count ≤100 × 103/mm3 (OR 2.86, 95% CI: 1.07–7.67; P = 0.01 as significant predictors of a major bleed. Overall hospital mortality was 57.7%. Multivariate analysis demonstrated the presence of ≥3 organ dysfunctions (OR 2.46, 95% CI: 1.19–5.09; P < 0.05 and medical intensive care unit admission (OR 1.99, 95% CI: 1.00–3.98; P = 0.05 were independent variables associated with hospital mortality. Conclusion: Patients receiving rhAPC at our institution had higher APACHE II scores, mortality, and major bleeding events than published

  14. Enhanced biological activity of carotenoids stabilized by phenyl groups.

    Science.gov (United States)

    You, Ji Suk; Jeon, Sunhwa; Byun, Youn Jung; Koo, Sangho; Choi, Shin Sik

    2015-06-15

    Carotenoids are lipid soluble food ingredients with multifunction including antioxidant and anticancer activities. However, carotenoids are destructively oxidized upon reaction with radicals resulting in toxic effects on biological systems. Two synthetic carotenoids (BAS and BTS) containing the aromatic phenyl groups with a para-substituent (OMe and Me, respectively) at C-13 and C-13' position were prepared in order to overcome a structural instability of carotenoid. Both BAS and BTS exerted stronger radical scavenging activity than β-carotene in DPPH and ABTS assays. In particular, BTS significantly reduced in vivo ROS (reactive oxygen species) levels and improved body growth and reproduction of Caenorhabditiselegans. BTS has a great potential for the advanced and modified carotenoid material with stability leading to enhanced bioavailability.

  15. Synthesis, characterization and biological activity of Rhein-cyclodextrin conjugate

    Science.gov (United States)

    Liu, Manshuo; Lv, Pin; Liao, Rongqiang; Zhao, Yulin; Yang, Bo

    2017-01-01

    Cyclodextrin conjugate complexation is a useful method to enhance the solubility and absorption of poorly soluble drugs. A series of new Rhein-β-cyclodextrin conjugates (Rh-CD conjugates) have been synthesized and examined. Rhein is covalently linked with the β-CD by amido linkage in a 1:1 molar ratio. The conjugates were characterized by 1H NMR, 13C NMR, HRMS, powder X-ray diffraction (powder XRD) as well as thermogravimetric analysis (TGA). The results reveal that incorporation of β-CD could improve the aqueous solubility of Rhein and the cytotoxicity against hepatocellular carcinoma (HepG2) cell line as well as antibacterial activity against three organisms. The improved biological activity and the satisfactory water solubility of the conjugates will be potentially useful for developing novel drug-cyclodextrin conjugates, such as herbal medicine.

  16. Posttranslational modulation on the biological activities of molecular chaperones

    Institute of Scientific and Technical Information of China (English)

    2009-01-01

    Molecular chaperones are a family of proteins that were first noticed to exist about 45 years ago from their increased transcription under heat shock conditions.As a result,the regulation of their encoding genes has been subject to extensive studies.Recent studies revealed that the biological activities of molecular chaperones can also be effectively modulated at the protein level.The ways of modulation so far elucidated include allosteric effect,covalent modification,protein-protein interaction,and con-formational alteration induced by such macro-environmental conditions as temperature and pH.These latter aspects were reviewed here.Emphasized here is the importance of such immediate structural alterations that lead to an immediate activity increase,providing the immediate protection needed for the cells to survive the stress conditions.

  17. Posttranslational modulation on the biological activities of molecular chaperones

    Institute of Scientific and Technical Information of China (English)

    CHANG ZengYi

    2009-01-01

    Molecular chaperones are a family of proteins that were first noticed to exist about 45 years ago from their increased transcription under heat shock conditions. As a result, the regulation of their encoding genes has been subject to extensive studies. Recent studies revealed that the biological activities of molecular chaperones can also be effectively modulated at the protein level. The ways of modulation so far elucidated include allosteric effect, covalent modification, protein-protein interaction, and con-formational alteration induced by such macro-environmental conditions as temperature and pH. These latter aspects were reviewed here. Emphasized here is the importance of such immediate structural alterations that lead to an immediate activity increase, providing the immediate protection needed for the cells to survive the stress conditions.

  18. Distribution and biological activity of obestatin in the rat.

    Science.gov (United States)

    Dun, Siok L; Brailoiu, G Cristina; Brailoiu, Eugen; Yang, Jun; Chang, Jaw Kang; Dun, Nae J

    2006-11-01

    Obestatin, a 23 amino acid peptide recently isolated from the rat stomach, is encoded by the same gene that encodes ghrelin. With the use of an antiserum directed against the mouse/rat obestatin, obestatin immunoreactivity (irOBS) was detected in cells of the gastric mucosa, myenteric plexus, and in Leydig cells of the testis in Sprague-Dawley rats. Double labeling the myenteric plexus with obestatin antiserum and choline acetyltransferase (ChAT) antiserum revealed that nearly all irOBS neurons were ChAT positive and vice versa. For comparative purposes, myenteric ganglion cells, cells in the gastric mucosa, and Leydig cells of the testis were shown to be immunoreactive to preproghrelin. The biological activity of obestatin on rat central neurons was assessed by the calcium microfluorimetric Fura-2 method. Obestatin (100 nM) administered to dissociated and cultured rat cerebral cortical neurons elevated cytosolic calcium concentrations [Ca2+]i in a population of cortical neurons. The result provides the first immunohistochemical evidence that obestatin is expressed in cells of the gastric mucosa and myenteric ganglion cells, and also in Leydig cells of the testis; the peptide is biologically active on central neurons.

  19. Fruit cuticular waxes as a source of biologically active triterpenoids.

    Science.gov (United States)

    Szakiel, Anna; Pączkowski, Cezary; Pensec, Flora; Bertsch, Christophe

    2012-06-01

    The health benefits associated with a diet rich in fruit and vegetables include reduction of the risk of chronic diseases such as cardiovascular disease, diabetes and cancer, that are becoming prevalent in the aging human population. Triterpenoids, polycyclic compounds derived from the linear hydrocarbon squalene, are widely distributed in edible and medicinal plants and are an integral part of the human diet. As an important group of phytochemicals that exert numerous biological effects and display various pharmacological activities, triterpenoids are being evaluated for use in new functional foods, drugs, cosmetics and healthcare products. Screening plant material in the search for triterpenoid-rich plant tissues has identified fruit peel and especially fruit cuticular waxes as promising and highly available sources. The chemical composition, abundance and biological activities of triterpenoids occurring in cuticular waxes of some economically important fruits, like apple, grape berry, olive, tomato and others, are described in this review. The need for environmentally valuable and potentially profitable technologies for the recovery, recycling and upgrading of residues from fruit processing is also discussed.

  20. Inhalable DNase I microparticles engineered with biologically active excipients.

    Science.gov (United States)

    Osman, Rihab; Al Jamal, Khuloud T; Kan, Pei-Lee; Awad, Gehanne; Mortada, Nahed; El-Shamy, Abd-Elhameed; Alpar, Oya

    2013-12-01

    Highly viscous mucus poses a big challenge for the delivery of particulates carrying therapeutics to patients with cystic fibrosis. In this study, surface modifying DNase I loaded particles using different excipients to achieve better lung deposition, higher enzyme stability or better biological activity had been exploited. For the purpose, controlled release microparticles (MP) were prepared by co-spray drying DNase I with the polymer poly-lactic-co-glycolic acid (PLGA) and the biocompatible lipid surfactant 1,2-dipalmitoyl-Sn-phosphatidyl choline (DPPC) using various hydrophilic excipients. The effect of the included modifiers on the particle morphology, size, zeta potential as well as enzyme encapsulation efficiency, biological activity and release had been evaluated. Powder aerosolisation performance and particle phagocytosis by murine macrophages were also investigated. The results showed that more than 80% of enzyme activity was recovered after MP preparation and that selected surface modifiers greatly increased the enzyme encapsulation efficiency. The particle morphology was greatly modified altering in turn the powders inhalation indices where dextran, ovalbumin and chitosan hydrochloride increased considerably the respirable fraction compared to the normal hydrophilic carriers lactose and PVP. Despite of the improved aerosolisation caused by chitosan hydrochloride, yet retardation of chitosan coated particles in artificial mucus samples discouraged its application. On the other hand, dextran and polyanions enhanced DNase I effect in reducing cystic fibrosis mucus viscosity. DPPC proved good ability to reduce particles phagocytic uptake even in the presence of the selected adjuvants. The prepared MP systems were biocompatible with lung epithelial cells. To conclude, controlled release DNase I loaded PLGA-MP with high inhalation indices and enhanced mucolytic activity on CF sputum could be obtained by surface modifying the particles with PGA or dextran.

  1. Comparison of biological activities of Glycyrrhiza glabra and G. uralensis

    Directory of Open Access Journals (Sweden)

    Park Chun-Geon

    2014-01-01

    Full Text Available The biological activities of Glycyrrhiza (GLs extracts (GL-1, Glycyrrhiza glabra from Eumseong, Korea; GL-2, G. uralensis from Eumseong, Korea; GL-3, G. uralensis from Yeongcheon, Korea; GL-4, G. uralensis from Neimenggu, China: GL-5, G. uralensis purchased from Korea Medicine Herbal Association, Korea were investigated. G. uralensis (GLs-2, -3, -4, and -5 extracts exhibited higher free radical scavenging activity against DPPH and OH radicals than G. glabra (GL-1. In addition, all GLs had antibacterial activity against E. coli, S. aureus, and H. pylori. GL-3 inhibited the growth of E. coli and S. aureus, while GL-1 had antibacterial activity against H. pylori. All GL extracts tested inhibited the lipopolysaccharide- and interferon-γ-induced inflammatory activity of RAW 264.7 cells. G. glabra and G. uralensis reduced NO generation. GL-3 also inhibited the growth of AGS human gastric adenocarcinoma cells. GLs-3 and -4 showed the inhibition of rat lens aldose reductase. GL-4 had a higher total content of glycyrrhizin (1, glycyrrhetinic acid (2, glabridin (3, and isoliquiritigenin (4. G. uralensis (GLs-2, -3, -4, and -5 is thus more effective than G. glabra (GL-1.

  2. Production of recombinant insulin-like androgenic gland hormones from three decapod species: In vitro testicular phosphorylation and activation of a newly identified tyrosine kinase receptor from the Eastern spiny lobster, Sagmariasus verreauxi.

    Science.gov (United States)

    Aizen, Joseph; Chandler, Jennifer C; Fitzgibbon, Quinn P; Sagi, Amir; Battaglene, Stephen C; Elizur, Abigail; Ventura, Tomer

    2016-04-01

    In crustaceans the insulin-like androgenic gland hormone (IAG) is responsible for male sexual differentiation. To date, the biochemical pathways through which IAG exerts its effects are poorly understood and could be elucidated through the production of a functional recombinant IAG (rIAG). We have successfully expressed glycosylated, biologically active IAG using the Pichia pastoris yeast expression system. We co-expressed recombinant single-chain precursor molecules consisting of the B and A chains (the mature hormone) tethered by a flexible linker, producing rIAGs of the following commercially important species: Eastern spiny lobster Sagmariasus verreauxi (Sv), redclaw crayfish Cherax quadricarinatus (Cq) and giant freshwater prawn Macrobrachium rosenbergii (Mr). We then tested the biological activity of each, through the ability to increase phosphorylation in the testis; both Sv and Cq rIAGs significantly elevated phosphorylation specific to their species, and in a dose-dependent manner. Mr rIAG was tested on Macrobrachium australiense (Ma), eliciting a similar response. Moreover, using bioinformatics analyses of the de novo assembled spiny lobster transcriptome, we identified a spiny lobster tyrosine kinase insulin receptor (Sv-TKIR). We validated this discovery with a receptor activation assay in COS-7 cells expressing Sv-TKIR, using a reporter SRE-LUC system designed for RTKs, with each of the rIAG proteins acting as the activation ligand. Using recombinant proteins, we aim to develop specific tools to control sexual development through the administration of IAG within the critical sexual differentiation time window. The biologically active rIAGs generated might facilitate commercially feasible solutions for the long sought techniques for sex-change induction and monosex population culture in crustaceans and shed new light on the physiological mode of action of IAG in crustaceans.

  3. The synthesis of hydroxyapatite with different crystallinities by controlling the concentration of recombinant CEMP1 for biological application.

    Science.gov (United States)

    Chen, Xiaofeng; Liu, Yu; Yang, Jie; Wu, Wenlei; Miao, Leiying; Yu, Yijun; Yang, Xuebin; Sun, Weibin

    2016-02-01

    Cementum protein 1 (CEMP1) has been reported to be cementum specific and it plays a role in the properties and structure of cementum mineralization. This study was carried out to test the hypothesis that, CEMP1 had the capacity to guide ordered HA formation as it accumulated at targeting site. Crystals induced by protein were prepared by biomimetic method. The effects of CEMP1 concentrations (0-100μg/ml) on the rate of calcium phosphate precipitation was monitored during 48h, while the formed mineral phases were assessed utilizing transmission electron microscope (TEM), selected area electron diffraction (SAED), X-ray diffraction (XRD), and Fourier transform infrared spectroscopy (FTIR). The results showed that recombinant cementum protein 1 (rhCEMP1) affected the crystal nucleation and growth. At lower rhCEMP1 concentrations (0-50μg/ml), limited mineral formation occurred and only small bundles of HA crystals were found. However, with 100μg/ml of rhCEMP1, a predominance of organized linear needle-like HA crystals was observed. FTIR revealed the characteristic bands of HA appearing at 1106cm(-1), 557cm(-1), and 598cm(-1). Therefore, increasing the concentration of rhCEMP1 can lead to the formation of ordered bundles of HA crystals in vitro.

  4. Recombinant Saccharomyces cerevisiae strain expressing a model cytochrome P450 in the rat digestive environment: viability and bioconversion activity.

    Science.gov (United States)

    Garrait, G; Jarrige, J F; Blanquet, S; Beyssac, E; Alric, M

    2007-06-01

    An innovative "biodrug" concept, based on the oral administration of living recombinant microorganisms, has recently emerged for the prevention or treatment of various diseases. An engineered Saccharomyces cerevisiae strain expressing plant P450 73A1 (cinnamate-4-hydroxylase [CA4H] activity) was used, and its survival and ability to convert trans-cinnamic acid (CIN) into p-coumaric acid (COU) were investigated in vivo. In rats, the recombinant yeast was resistant to gastric and small intestinal secretions but was more sensitive to the conditions found in the large intestine. After oral administration of yeast and CIN, the CA4H activity was shown in vivo, with COU being found throughout the rat's digestive tract and in its urine. The bioconversion reaction occurred very fast, with most of the COU being produced within the first 5 min. The gastrointestinal sac technique demonstrated that the recombinant yeast was able to convert CIN into COU (conversion rate ranging from 2 to 5%) in all the organs of the rat's digestive tract: stomach, duodenum, jejunum, ileum, cecum, and colon. These results promise new opportunities for the development of drug delivery systems based on engineered yeasts catalyzing a bioconversion reaction directly in the digestive tract.

  5. Recombinant receptor/reporter gene bioassays for assessing the estrogenic and dioxin-like activities of xenobiotics and complex mixtures

    Energy Technology Data Exchange (ETDEWEB)

    Zacharewski, T. [Univ. of Western Ontario, London, Ontario (Canada). Dept. of Pharmacology and Toxicology

    1995-12-31

    Exposure to naturally occurring or synthetic substances that possess sex steroid and/or dioxin-like activity may have long range effects on human health, reproductive fitness and environmental quality. Results from recent epidemiological studies have suggested that xenobiotics with sex steroid activity may contribute to the development of hormone-dependent cancers and disorders in the male reproductive tract as well as attenuate sperm production. However, most of these compounds, which are referred to as endocrine disruptors, are structurally dissimilar to sex steroids. Yet, based upon ambiguous assays, it has been conceded that the effects of these compounds are mediated by receptors. The authors have taken advantage of the mechanism of action of these compounds to develop recombinant receptor/reporter gene bioassays for environmental estrogens and dioxin-like compounds. The assays use an easily measurable enzyme activity (i.e. firefly luciferase), exhibit improved sensitivity and selectivity and are amenable to automation. Data will be presented demonstrating that phytoestrogens (e.g. genistein) and xenobiotics such as pesticides (e.g. DDT, Kepone), nonionic surfactants (e.g. p-nonylphenol), and precursors used in the manufacture of plastics (e.g. Bisphenol A) exhibit estrogenic activity. In addition, the assays have been used to detect estrogenic and dioxin-like activity in complex mixtures such as pulp and paper mill black liquor and effluent. These results demonstrate the utility of recombinant receptor/reporter gene bioassays for identifying substances or complex mixtures with estrogenic and/or dioxin-like activity.

  6. Phytochemical profile and biological activity of Juglans regia.

    Science.gov (United States)

    Panth, Nisha; Paudel, Keshav Raj; Karki, Rajendra

    2016-09-01

    Juglans regia Linn. (Juglandaceae), popularly known as English or Persian walnut, is a valuable medicinal plant with a potency to cure various diseases in traditional medicine. Since ancient time, different local ethnic groups have used various part of J. regia for a wide array of ailments including helminthiasis, diarrhea, sinusitis, stomach ache, arthritis, asthma, eczema, scrofula, skin disorders, diabetes mellitus, anorexia, thyroid dysfunction, cancer and infectious diseases. Biological activities of J. regia have been reported in several peer review journals and scientific attention is increasing. The present review attempts to provide comprehensive information on plant description, ethnobotanical use, toxicity, phytochemical profile, pharmacology, clinical studies and current research prospective of the J. regia. Currently, there is an immense interest on isolation/identification of active constituents from walnut and screening those active compounds for pharmacological activities. In addition, researchers are performing clinical trials as well as screening various solvent extracts or fractions of J. regia in several animal diseases models to identify promising therapeutic benefits. In the present work, we review the latest information based on published scientific investigations of J. regia.

  7. Synthesis and biological activity of hydroxycinnamoyl containing antiviral drugs

    Directory of Open Access Journals (Sweden)

    Chochkova Maya G.

    2014-01-01

    Full Text Available Seven N-hydroxycinnamoyl amides were synthesized by EDC/HOBt coupling of the corresponding substituted cinnamic acids (p-coumaric-, ferulic-, sinapic- and caffeic acids with influenza antivirals (amantadine, rimantadine and oseltamivir. DPPH (1,1-diphenyl-2-picrylhydrazyl scavenging abilities and the inhibitory effect on mushroom tyrosinase activity (using L-tyrosine as the substrate were investigated in vitro. Amongst the synthesized compounds, N-[(E-3-(3’,4’-dihydroxyphenyl-2-propenoyl]oseltamivir (1 and N-[(E-3-(3’,4’-dihydroxyphenyl-2-propenoyl]rimantadine (4, containing catechol moiety, exhibited the most potent DPPH radical-scavenging activity. Amide (1 displayed also tyrosinase inhibitory effect toward L-tyrosine as the substrate (~50%. Due to its biological activities revealed so far compound (1 can be considered as a promising candidate for a cosmetic ingredient. The synthesized compounds were also investigated for their in vitro inhibitory activity against the replication of influenza virus A (H3N2.

  8. Biological Activities of Oleanolic Acid Derivatives from Calendula officinalis Seeds.

    Science.gov (United States)

    Zaki, Ahmed; Ashour, Ahmed; Mira, Amira; Kishikawa, Asuka; Nakagawa, Toshinori; Zhu, Qinchang; Shimizu, Kuniyoshi

    2016-05-01

    Phytochemical examination of butanol fraction of Calendula officinalis seeds led to the isolation of two compounds identified as 28-O-β-D-glucopyranosyl-oleanolic acid 3-O-β-D-glucopyranosyl (1→3)-β-D-glucopyranosiduronic acid (CS1) and oleanolic acid 3-O-β-D-glucopyranosyl (1→3)-β-D-glucopyranosiduronic acid (CS2). Biological evaluation was carried out for these two compounds such as melanin biosynthesis inhibitory, hyaluronic acid production activities, anti obesity using lipase inhibition and adipocyte differentiation as well as evaluation of the protective effect against hydrogen peroxide induced neurotoxicity in neuro-2A cells. The results showed that, compound CS2 has a melanin biosynthesis stimulatory activity; however, compound CS1 has a potent stimulatory effect for the production of hyaluronic acid on normal human dermal fibroblast from adult (NHDF-Ad). Both compounds did not show any inhibitory effect on both lipase and adipocyte differentiation. Compound CS2 could protect neuro-2A cells and increased cell viability against H2 O2 . These activities (melanin biosynthesis stimulatory and protective effect against H2 O2 of CS2 and hyaluronic acid productive activities of these triterpene derivatives) have been reported for the first time. Copyright © 2016 John Wiley & Sons, Ltd.

  9. Soil degradation effect on biological activity in Mediterranean calcareous soils

    Science.gov (United States)

    Roca-Pérez, L.; Alcover-Sáez, S.; Mormeneo, S.; Boluda, R.

    2009-04-01

    Soil degradation processes include erosion, organic matter decline, compaction, salinization, landslides, contamination, sealing and biodiversity decline. In the Mediterranean region the climatological and lithological conditions, together with relief on the landscape and anthropological activity are responsible for increasing desertification process. It is therefore considered to be extreme importance to be able to measure soil degradation quantitatively. We studied soil characteristics, microbiological and biochemical parameters in different calcareous soil sequences from Valencia Community (Easter Spain), in an attempt to assess the suitability of the parameters measured to reflect the state of soil degradation and the possibility of using the parameters to assess microbiological decline and soil quality. For this purpose, forest, scrubland and agricultural soil in three soil sequences were sampled in different areas. Several sensors of the soil biochemistry and microbiology related with total organic carbon, microbial biomass carbon, soil respiration, microorganism number and enzyme activities were determined. The results show that, except microorganism number, these parameters are good indicators of a soil biological activity and soil quality. The best enzymatic activities to use like indicators were phosphatases, esterases, amino-peptidases. Thus, the enzymes test can be used as indicators of soil degradation when this degradation is related with organic matter losses. There was a statistically significant difference in cumulative O2 uptake and extracellular enzymes among the soils with different degree of degradation. We would like to thank Spanish government-MICINN for funding and support (MICINN, project CGL2006-09776).

  10. Chemistry and biological activities of flavonoids: an overview.

    Science.gov (United States)

    Kumar, Shashank; Pandey, Abhay K

    2013-01-01

    There has been increasing interest in the research on flavonoids from plant sources because of their versatile health benefits reported in various epidemiological studies. Since flavonoids are directly associated with human dietary ingredients and health, there is need to evaluate structure and function relationship. The bioavailability, metabolism, and biological activity of flavonoids depend upon the configuration, total number of hydroxyl groups, and substitution of functional groups about their nuclear structure. Fruits and vegetables are the main dietary sources of flavonoids for humans, along with tea and wine. Most recent researches have focused on the health aspects of flavonoids for humans. Many flavonoids are shown to have antioxidative activity, free radical scavenging capacity, coronary heart disease prevention, hepatoprotective, anti-inflammatory, and anticancer activities, while some flavonoids exhibit potential antiviral activities. In plant systems, flavonoids help in combating oxidative stress and act as growth regulators. For pharmaceutical purposes cost-effective bulk production of different types of flavonoids has been made possible with the help of microbial biotechnology. This review highlights the structural features of flavonoids, their beneficial roles in human health, and significance in plants as well as their microbial production.

  11. Chemistry and Biological Activities of Flavonoids: An Overview

    Directory of Open Access Journals (Sweden)

    Shashank Kumar

    2013-01-01

    Full Text Available There has been increasing interest in the research on flavonoids from plant sources because of their versatile health benefits reported in various epidemiological studies. Since flavonoids are directly associated with human dietary ingredients and health, there is need to evaluate structure and function relationship. The bioavailability, metabolism, and biological activity of flavonoids depend upon the configuration, total number of hydroxyl groups, and substitution of functional groups about their nuclear structure. Fruits and vegetables are the main dietary sources of flavonoids for humans, along with tea and wine. Most recent researches have focused on the health aspects of flavonoids for humans. Many flavonoids are shown to have antioxidative activity, free radical scavenging capacity, coronary heart disease prevention, hepatoprotective, anti-inflammatory, and anticancer activities, while some flavonoids exhibit potential antiviral activities. In plant systems, flavonoids help in combating oxidative stress and act as growth regulators. For pharmaceutical purposes cost-effective bulk production of different types of flavonoids has been made possible with the help of microbial biotechnology. This review highlights the structural features of flavonoids, their beneficial roles in human health, and significance in plants as well as their microbial production.

  12. Biological Activities of the Essential Oil from Erigeron floribundus

    Directory of Open Access Journals (Sweden)

    Riccardo Petrelli

    2016-08-01

    Full Text Available Erigeron floribundus (Asteraceae is an herbaceous plant widely used in Cameroonian traditional medicine to treat various diseases of microbial and non-microbial origin. In the present study, we evaluated the in vitro biological activities displayed by the essential oil obtained from the aerial parts of E. floribundus, namely the antioxidant, antimicrobial and antiproliferative activities. Moreover, we investigated the inhibitory effects of E. floribundus essential oil on nicotinate mononucleotide adenylyltransferase (NadD, a promising new target for developing novel antibiotics, and Trypanosoma brucei, the protozoan parasite responsible for Human African trypanosomiasis. The essential oil composition was dominated by spathulenol (12.2%, caryophyllene oxide (12.4% and limonene (8.8%. The E. floribundus oil showed a good activity against Staphylococcus aureus (inhibition zone diameter, IZD of 14 mm, minimum inhibitory concentration, MIC of 512 µg/mL. Interestingly, it inhibited the NadD enzyme from S. aureus (IC50 of 98 µg/mL, with no effects on mammalian orthologue enzymes. In addition, T. brucei proliferation was inhibited with IC50 values of 33.5 µg/mL with the essential oil and 5.6 µg/mL with the active component limonene. The essential oil exhibited strong cytotoxicity on HCT 116 colon carcinoma cells with an IC50 value of 14.89 µg/mL, and remarkable ferric reducing antioxidant power (tocopherol-equivalent antioxidant capacity, TEAC = 411.9 μmol·TE/g.

  13. Catalytically and biologically active silver nanoparticles synthesized using essential oil

    Science.gov (United States)

    Vilas, Vidya; Philip, Daizy; Mathew, Joseph

    2014-11-01

    There are numerous reports on phytosynthesis of silver nanoparticles and various phytochemicals are involved in the reduction and stabilization. Pure explicit phytosynthetic protocol for catalytically and biologically active silver nanoparticles is of importance as it is an environmentally benign green method. This paper reports the use of essential oil of Myristica fragrans enriched in terpenes and phenyl propenes in the reduction and stabilization. FTIR spectra of the essential oil and the synthesized biogenic silver nanoparticles are in accordance with the GC-MS spectral analysis reports. Nanosilver is initially characterized by an intense SPR band around 420 nm, followed by XRD and TEM analysis revealing the formation of 12-26 nm sized, highly pure, crystalline silver nanoparticles. Excellent catalytic and bioactive potential of the silver nanoparticles is due to the surface modification. The chemocatalytic potential of nanosilver is exhibited by the rapid reduction of the organic pollutant, para nitro phenol and by the degradation of the thiazine dye, methylene blue. Significant antibacterial activity of the silver colloid against Gram positive, Staphylococcus aureus (inhibition zone - 12 mm) and Gram negative, Escherichia coli (inhibition zone - 14 mm) is demonstrated by Agar-well diffusion method. Strong antioxidant activity of the biogenic silver nanoparticles is depicted through NO scavenging, hydrogen peroxide scavenging, reducing power, DPPH and total antioxidant activity assays.

  14. Biological Activities of the Essential Oil from Erigeron floribundus.

    Science.gov (United States)

    Petrelli, Riccardo; Orsomando, Giuseppe; Sorci, Leonardo; Maggi, Filippo; Ranjbarian, Farahnaz; Biapa Nya, Prosper C; Petrelli, Dezemona; Vitali, Luca A; Lupidi, Giulio; Quassinti, Luana; Bramucci, Massimo; Hofer, Anders; Cappellacci, Loredana

    2016-08-13

    Erigeron floribundus (Asteraceae) is an herbaceous plant widely used in Cameroonian traditional medicine to treat various diseases of microbial and non-microbial origin. In the present study, we evaluated the in vitro biological activities displayed by the essential oil obtained from the aerial parts of E. floribundus, namely the antioxidant, antimicrobial and antiproliferative activities. Moreover, we investigated the inhibitory effects of E. floribundus essential oil on nicotinate mononucleotide adenylyltransferase (NadD), a promising new target for developing novel antibiotics, and Trypanosoma brucei, the protozoan parasite responsible for Human African trypanosomiasis. The essential oil composition was dominated by spathulenol (12.2%), caryophyllene oxide (12.4%) and limonene (8.8%). The E. floribundus oil showed a good activity against Staphylococcus aureus (inhibition zone diameter, IZD of 14 mm, minimum inhibitory concentration, MIC of 512 µg/mL). Interestingly, it inhibited the NadD enzyme from S. aureus (IC50 of 98 µg/mL), with no effects on mammalian orthologue enzymes. In addition, T. brucei proliferation was inhibited with IC50 values of 33.5 µg/mL with the essential oil and 5.6 µg/mL with the active component limonene. The essential oil exhibited strong cytotoxicity on HCT 116 colon carcinoma cells with an IC50 value of 14.89 µg/mL, and remarkable ferric reducing antioxidant power (tocopherol-equivalent antioxidant capacity, TEAC = 411.9 μmol·TE/g).

  15. Phytochemical profi le and biological activity of Juglans regia

    Institute of Scientific and Technical Information of China (English)

    Nisha Panth; Keshav Raj Paudel; Rajendra Karki

    2016-01-01

    Juglans regia Linn. (Juglandaceae), popularly known as English or Persian walnut, is a valuable medicinal plant with a potency to cure various diseases in traditional medicine. Since ancient time, different local ethnic groups have used various part ofJ. regia for a wide array of ailments including helminthiasis, diarrhea, sinusitis, stomach ache, arthritis, asthma, eczema, scrofula, skin disorders, diabetes melitus, anorexia, thyroid dysfunction, cancer and infectious diseases. Biological activities of J. regia have been reported in several peer review journals and scientifi c attention is increasing. The present review attempts to provide comprehensive information on plant description, ethnobotanical use, toxicity, phytochemical profi le, pharmacology, clinical studies and current research prospective of theJ. regia. Currently, there is an immense interest on isolation/identifi cation of active constituents from walnut and screening those active compounds for pharmacological activities. In addition, researchers are performing clinical trials as wel as screening various solvent extracts or fractions ofJ. regiain several animal diseases models to identify promising therapeutic benefi ts. In the present work, we review the latest information based on published scientifi c investigations ofJ. regia.

  16. Base composition, selection, and phylogenetic significance of indels in the recombination activating gene-1 in vertebrates

    Directory of Open Access Journals (Sweden)

    Vences Miguel

    2009-12-01

    Full Text Available Abstract Background The Recombination Activating Proteins, RAG1 and RAG2, play a crucial role in the immune response in vertebrates. Among the nuclear markers currently used for phylogenetic purposes, Rag1 has especially enjoyed enormous popularity, since it successfully contributed to elucidating the relationships among and within a large variety of vertebrate lineages. We here report on a comparative investigation of the genetic variation, base composition, presence of indels, and selection in Rag1 in different vertebrate lineages (Actinopterygii, Amphibia, Aves, Chondrichthyes, Crocodylia, Lepidosauria, Mammalia, and Testudines through the analysis of 582 sequences obtained from Genbank. We also analyze possible differences between distinct parts of the gene with different type of protein functions. Results In the vertebrate lineages studied, Rag1 is over 3 kb long. We observed a high level of heterogeneity in base composition at the 3rd codon position in some of the studied vertebrate lineages and in some specific taxa. This result is also paralleled by taxonomic differences in the GC content at the same codon position. Moreover, positive selection occurs at some sites in Aves, Lepidosauria and Testudines. Indels, which are often used as phylogenetic characters, are more informative across vertebrates in the 5' than in the 3'-end of the gene. When the entire gene is considered, the use of indels as phylogenetic character only recovers one major vertebrate clade, the Actinopterygii. However, in numerous cases insertions or deletions are specific to a monophyletic group. Conclusions Rag1 is a phylogenetic marker of undoubted quality. Our study points to the need of carrying out a preliminary investigation on the base composition and the possible existence of sites under selection of this gene within the groups studied to avoid misleading resolution. The gene shows highly heterogeneous base composition, which affects some taxa in particular and

  17. Plasmid construction using recombination activity in the fission yeast Schizosaccharomyces pombe.

    Directory of Open Access Journals (Sweden)

    Ayako Chino

    Full Text Available BACKGROUND: Construction of plasmids is crucial in modern genetic manipulation. As of now, the common method for constructing plasmids is to digest specific DNA sequences with restriction enzymes and to ligate the resulting DNA fragments with DNA ligase. Another potent method to construct plasmids, known as gap-repair cloning (GRC, is commonly used in the budding yeast Saccharomyces cerevisiae. GRC makes use of the homologous recombination activity that occurs within the yeast cells. Due to its flexible design and efficiency, GRC has been frequently used for constructing plasmids with complex structures as well as genome-wide plasmid collections. Although there have been reports indicating GRC feasibility in the fission yeast Schizosaccharomyces pombe, this species is not commonly used for GRC as systematic studies of reporting GRC efficiency in S. pombe have not been performed till date. METHODOLOGY/PRINCIPAL FINDINGS: We investigated GRC efficiency in S. pombe in this study. We first showed that GRC was feasible in S. pombe by constructing a plasmid that contained the LEU2 auxotrophic marker gene in vivo and showed sufficient efficiency with short homology sequences (>25 bp. No preference was shown for the sequence length from the cut site in the vector plasmid. We next showed that plasmids could be constructed in a proper way using 3 DNA fragments with 70% efficiency without any specific selections being made. The GRC efficiency with 3 DNA fragments was dramatically increased >95% in lig4Delta mutant cell, where non-homologous end joining is deficient. Following this approach, we successfully constructed plasmid vectors with leu1+, ade6+, his5+, and lys1+ markers with the low-copy stable plasmid pDblet as a backbone by applying GRC in S. pombe. CONCLUSIONS/SIGNIFICANCE: We concluded that GRC was sufficiently feasible in S. pombe for genome-wide gene functional analysis as well as for regular plasmid construction. Plasmids with different

  18. Crystallization and preliminary crystallographic analysis of latent, active and recombinantly expressed aurone synthase, a polyphenol oxidase, from Coreopsis grandiflora

    Energy Technology Data Exchange (ETDEWEB)

    Molitor, Christian; Mauracher, Stephan Gerhard; Rompel, Annette, E-mail: annette.rompel@univie.ac.at [Universität Wien, Althanstrasse 14, 1090 Wien (Austria)

    2015-05-22

    Latent and active aurone synthase purified from petals of C. grandiflora (cgAUS1) were crystallized. The crystal quality of recombinantly expressed latent cgAUS1 was significantly improved by co-crystallization with the polyoxotungstate Na{sub 6}[TeW{sub 6}O{sub 24}] within the liquid–liquid phase-separation zone. Aurone synthase (AUS), a member of a novel group of plant polyphenol oxidases (PPOs), catalyzes the oxidative conversion of chalcones to aurones. Two active cgAUS1 (41.6 kDa) forms that differed in the level of phosphorylation or sulfation as well as the latent precursor form (58.9 kDa) were purified from the petals of Coreopsis grandiflora. The differing active cgAUS1 forms and the latent cgAUS1 as well as recombinantly expressed latent cgAUS1 were crystallized, resulting in six different crystal forms. The active forms crystallized in space groups P2{sub 1}2{sub 1}2{sub 1} and P12{sub 1}1 and diffracted to ∼1.65 Å resolution. Co-crystallization of active cgAUS1 with 1,4-resorcinol led to crystals belonging to space group P3{sub 1}21. The crystals of latent cgAUS1 belonged to space group P12{sub 1}1 and diffracted to 2.50 Å resolution. Co-crystallization of recombinantly expressed pro-AUS with the hexatungstotellurate(VI) salt Na{sub 6}[TeW{sub 6}O{sub 24}] within the liquid–liquid phase separation zone significantly improved the quality of the crystals compared with crystals obtained without hexatungstotellurate(VI)

  19. Minimal information: an urgent need to assess the functional reliability of recombinant proteins used in biological experiments

    Directory of Open Access Journals (Sweden)

    de Marco Ario

    2008-07-01

    Full Text Available Abstract Structural characterization of proteins used in biological experiments is largely neglected. In most publications, the information available is totally insufficient to judge the functionality of the proteins used and, therefore, the significance of identified protein-protein interactions (was the interaction specific or due to unspecific binding of misfolded protein regions? or reliability of kinetic and thermodynamic data (how much protein was in its native form?. As a consequence, the results of single experiments might not only become questionable, but the whole reliability of systems biology, built on these fundaments, would be weakened. The introduction of Minimal Information concerning purified proteins to add as metadata to the main body of a manuscript would render straightforward the assessment of their functional and structural qualities and, consequently, of results obtained using these proteins. Furthermore, accepted standards for protein annotation would simplify data comparison and exchange. This article has been envisaged as a proposal for aggregating scientists who share the opinion that the scientific community needs a platform for Minimum Information for Protein Functionality Evaluation (MIPFE.

  20. Relationships between the stereochemistry and biological activity of fungal phytotoxins.

    Science.gov (United States)

    Evidente, Antonio; Andolfi, Anna; Cimmino, Alessio

    2011-10-01

    Toxins produced by phytopathogenic fungi assume great importance because of their involvement in several plant diseases. Although such pathogens are known to have seriously damaged crops, forest, and environmental resources, they represent a very important tool to develop new environmentally friendly herbicides and fungicides. This review deals with the relationships between the biological activity of some phytotoxins produced by pathogenic fungi for major forest plants and for damaging weeds and their stereochemistry. In particular, the methods used to determine their relative and/or absolute configuration will be illustrated. These include the application of Mosher's and Murata's methods, X-ray diffractometric analysis, circular dichroism, and the use of computational methods to determine the theoretical optical rotatory power as well as the CD spectrum. The importance of determining the absolute configuration to achieve the total synthesis of some phytotoxins, interesting for their potential practical application, is also discussed.

  1. Advances in the Chemical Analysis and Biological Activities of Chuanxiong

    Directory of Open Access Journals (Sweden)

    Jin-Ao Duan

    2012-09-01

    Full Text Available Chuanxiong Rhizoma (Chuan-Xiong, CX, the dried rhizome of Ligusticum chuanxiong Hort. (Umbelliferae, is one of the most popular plant medicines in the World. Modern research indicates that organic acids, phthalides, alkaloids, polysaccharides, ceramides and cerebrosides are main components responsible for the bioactivities and properties of CX. Because of its complex constituents, multidisciplinary techniques are needed to validate the analytical methods that support CX’s use worldwide. In the past two decades, rapid development of technology has advanced many aspects of CX research. The aim of this review is to illustrate the recent advances in the chemical analysis and biological activities of CX, and to highlight new applications and challenges. Emphasis is placed on recent trends and emerging techniques.

  2. Production and biological activities of yellow pigments from Monascus fungi.

    Science.gov (United States)

    Chen, Gong; Wu, Zhenqiang

    2016-08-01

    Monascus yellow pigments (MYPs), are azaphilone compounds and one of the three main components of total Monascus pigments (MPs). Thirty-five hydrophilic or hydrophobic MYPs have been identified, with the majority being hydrophobic. Apart from screening special Monascus strains, some advanced approaches, such as extractive and high-cell-density fermentations, have been applied for developing or producing new MYPs, especially extracellular hydrophilic MYPs. The outstanding performance of MYPs in terms of resistance to photodegradation, as well as tolerance for temperature and pH, give natural MYPs reasonable prospects, compared with the orange and red MPs, for practical use in the present and future. Meanwhile, MYPs have shown promising potential for applications in the food and pharmaceutical industries based on their described bioactivities. This review briefly summarizes the reports to date on chemical structures, biological activities, biosynthetic pathways, production technologies, and physicochemical performances of MYPs. The existing problems for MYPs are discussed and research prospects proposed.

  3. Infrared Spectra and Hydrogen Bonds of Biologically Active Benzaldehydes

    Science.gov (United States)

    Tolstorozhev, G. B.; Skornyakov, I. V.; Belkov, M. V.; Shimko, A. N.; Shadyro, O. I.; Brinkevich, S. D.; Samovich, S. N.

    2013-09-01

    IR-Fourier spectra of solutions and crystals of biologically active benzaldehyde derivatives were studied. Specific features of the formation of intra- and intermolecular hydrogen bonds were analyzed. Spectral signatures that characterized participation of the hydroxyl OH group and also the OCH3 and C=O groups in the formation of intramolecular hydrogen bonds of the three different types O-H···O-H, O-H···O-CH3, and O-H···O=C were revealed. Intramolecular hydrogen bonds of the types O-H···O-H and O-H···O-CH3 were absent for benzaldehyde derivatives in the crystal phase. Only hydroxyl and carbonyl groups participated in intermolecular interactions. This resulted in the formation of linear intermolecular dimers. Seven various configurations of the linear dimers were identified in solutions and crystals.

  4. Optimization of hypocrellin B derivative amphiphilicity and biological activity

    Institute of Scientific and Technical Information of China (English)

    LIU Xin; XIE Jie; ZHANG LuYong; CHEN HongXia; GU Ying; ZHAO JingQuan

    2009-01-01

    To satisfy the dual requirements of the fluent transportation in blood and the affinity to the target tissues of vascular diseases, hypocrellin derivatives with optimized amphiphilicity are expected. In this work, 3-amino-1-propanesulfonic acid and 4-amino-1-butanesulfonic acid substituted hypocrellin B,named compounds 1 and 2, were designed, synthesized in high yields and characterized. Besides greatly strengthened red absorption, the maximum solubility of compound 2 in phosphate buffered saline (PBS) is 4.2 mg/mL which is just enough to prepare an aqueous solution for intravenous injection in clinically acceptable concentration, while the partition coefficient between n-octanol and PBS,5.6, benefits the cell-uptake and biological activity as well. Furthermore, EPR measurements reveal that the photosensitization activities of the two compounds to generate semiquinone anion radicals, superoxide anion radicals and singlet oxygen are a little bit higher than those of taurine substituted hypocrellin B (THB), but the photodynamic activities to human lung cancer A549 cells are several times that of THB, mainly due to increases in lipophilicity and cell-uptake.

  5. Immobilization biological activated carbon used in advanced drinking water treatment

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    Bacteria separated from a mature filter bed of groundwater treatment plants were incubated in a culture media containing iron and manganese. A consortium of 5 strains of bacteria removing iron and manganese were obtained by repeated enrichment culturing. It was shown from the experiments of effect factors that ironmanganese removal bacteria in the culture media containing both Fe and Mn grew better than in that containing only Fe, however, they were unable to grow in the culture media containing only Mn. When comparing the bacteria biomass in the case ofρ (DO) =2.8 mg/L andρ (DO) =9.0 mg/L, no significant difference was found.The engineering bacteria removing the organic and the bacteria removing iron and manganese were simultaneously inoculated into activated carbon reactor to treat the effluent of distribution network. The experimental results showed that by using IBAC ( Immobilization Biological Activated Carbon) treatment, the removal efficiency of iron, manganese and permanganate index was more than 98% , 96% and 55% , respectively. After the influent with turbidity of 1.5 NTU, color of 25 degree and offensive odor was treated, the turbidity and color of effluence were less than 0.5 NTU and 15 degree, respectively, and it was odorless. It is determined that the cooperation function of engineering bacteria and activated carbon achieved advanced drinking water treatment.

  6. The oxidation of methionine-54 of epoetinum alfa does not affect molecular structure or stability, but does decrease biological activity.

    Science.gov (United States)

    Labrenz, Steven R; Calmann, Melissa A; Heavner, George A; Tolman, Glen

    2008-01-01

    Erythropoietin therapy is used to treat severe anemia in renal failure and chemotherapy patients. One of these therapies based on recombinant human erythropoietin is marketed under the trade name of EPREX and utilizes epoetinum alfa as the active pharmaceutical ingredient. The effect of oxidation of methionine-54 on the structure and stability of the erythropoietin molecule has not been directly tested. We have observed partial and full chemical oxidation of methionine-54 to methionine-54 sulfoxide, accomplished using tert-Butylhydroperoxide and hydrogen peroxide, respectively. A blue shift in the fluorescence center of spectral mass wavelength was observed as a linear response to the level of methionine sulfoxide in the epoetinum alfa molecule, presumably arising from a local change in the environment near tryptophan-51, as supported by potassium iodide quenching studies. Circular dichroism studies demonstrated no change in the folded structure of the molecule with methionine oxidation. The thermal unfolding profiles of partial and completely oxidized epoetinum alfa overlap, with a T(m) of 49.5 degrees C across all levels of methionine sulfoxide content. When the protein was tested for activity, a decrease in biological activity was observed, correlating with methionine sulfoxide levels. An allosteric effect between Met54, Trp51, and residues involved in receptor binding is proposed. These results indicate that methionine oxidation has no effect on the folded structure and global thermodynamic stability of the recombinant human erythropoietin molecule. Oxidation can affect potency, but only at levels significantly in excess of those seen in EPREX.

  7. Evaluation of the biological activity of sunflower hull extracts

    Energy Technology Data Exchange (ETDEWEB)

    Taha, F. S.; Wagdy, S. M.; Hassanein, M. M. M.; Hamed, S. F.

    2012-11-01

    This work was planned with the aim of adding value to sunflower seed hulls, a waste product of the oil industry by preparing a sunflower hull phenolic extract rich in chlorogenic acid (CGA). In order to fulfill this goal, the optimization for the extraction of a phenolic extract from the hulls was investigated. The parameters studied were: type of solvent, solvent to water ratio and hull to solvent ratio. In addition, the solvent mixtures were also studied. The resulting phenolic extracts were evaluated for their biological activities. This included phenolic content determination, evaluation of the antioxidant and antimicrobial activities. Chlorogenic acid was determined in two chosen hull extracts using the UV spectrophotometric method and HPLC analysis. The anti carcinogenic activity of the two chosen extracts was tested on seven different cell line carcinomas. The results revealed that all the phenolic extracts of sunflower hull studied contain between 190-312.5 mg phenolics/ 100 g hulls. The highest phenolic extraction was achieved with 80% methanol (1:30, hull to solvent, w/v ratio) and methanol to ethanol to water (7:7:6 v/v/v) mixture with values of 312.5 and 306.5 mg phenolics/100 g hulls, respectively. The free radical scavenging activity and antioxidant activity of all the samples ranged from 33.6-72.6%. The highest antioxidant activity and free radical scavenging activity were achieved by the same extracts that possessed the highest phenolic content, namely methanol to ethanol to water extract and 80% methanol with values 71.8 and 72.6%, 68.2 and 70.9% respectively, compared to 77.9 and 76.9% respectively for TBHQ. All the phenolic extracts possessed antimicrobial activity but to different levels against different pathogenic bacteria. The two chosen extracts also possessed anti carcinogenic activity, which differed among varying cell line carcinomas. The HPLC analysis indicated that chlorogenic acid was the main phenolic acid in the extract. Thus it can

  8. Inhibition on the production of collagen type Ⅰ, Ⅲ of activated hepatic stellate cells by antisense TIMP-1 recombinant plasmid

    Institute of Scientific and Technical Information of China (English)

    Wen-Bin Liu; Chang-Qing Yang; Wei Jiang; Yi-Qing Wang; Jing-Sheng Guo; Bo-Ming He; Ji-Yao Wang

    2003-01-01

    AIM: To investigate the inhibition effects on the productionof collagen type I, Ⅲ secreted by activated rat hepatic stellatecells (rHSCs) by antisense tissue inhibitors of metalloproteinase1 (TIMP-1) recombinant plasmid through elevating interstitialcollagenase activity.METHODS: rHSCs were extracted from normal rat liverby pronase and collagenase digestion and purified bycentrifugal elutriation, and were cultured on plastic dishesuntil they were activated to a myofibroblastic phenotypeafter 7-10 days. RT-Nest-PCR and gene recombinanttechniques were used to construct the rat antisense TIMP-1 recombinant plasmids which can express in eucaryoticcells. The recombinant plasmid and the pcDNA3 emptyplasmid were transfected in rHSCs by Effectene (QIAGEN)separately. Cells were selected after growing in DMEMcontaining 400 μg/ml G418 for 2-3 weeks. Expression ofexogenous gene was assessed by Northern blot, andexpression oflIMP-1 in rHSCs was determined by Northernblot and Western blot. We tested the interstitial collagenaseactivity with FITC-labled type I collagen as substrate.Ultimately, we quantified the type Ⅰ, Ⅲ collagen byWestern blot.RESULTS: The exogenous antisense TIMP-1 recombinantplasmid could be expressed in rHSCs well, which couldblock the expression of TIMP-1 greatly, the ratio of TIMP-1/GAPDH was 0.67, 2.41, and 2.97 separately at mRNAlevel (P<0.05); the ratio of TIMP-1/β-actin was 0.31, 0.98and 1.32 separately at protein level (P<0.05); It mightelevate active and latent interstitial collagenase activity,the collagenase activity was 0.3049, 0.1411 and 0.1196respectively. (P<0.05), which led to promotion thedegradation of type Ⅰ, Ⅲ collagen, the ratio of collagen I/β-actin was 0.63, 1.78 and 1.92 separately (P<0.05); andthe ratio of collagen Ⅲ/β-actin was 0.59, 1.81 and 1.98separately (P<0.05).CONCLUSION: These data shows that the antisense TIMP-1 recombinant plasmid has the inhibitory effects on theproduction of type Ⅰ, Ⅲ collagens

  9. Influence of Cardiopulmonary Bypass on the Interaction of Recombinant Factor VIIa with Activated Platelets

    OpenAIRE

    Kjalke, Marianne; Runge, Marx; Rojkjaer, Rasmus; Steinbruchel, Daniel; Johansson, Pär I

    2009-01-01

    Recombinant factor VIIa (rFVIIa) interacts preferentially with coated platelets characterized by a high exposure of phosphatidyl serine (PS), FV, FVIII, FIX, and FX binding, and fibrinogen. Cardiopulmonary bypass (CPB) is known to impair platelet function. In this study, the influence of CPB on formation of coated platelets and the interaction of rFVIIa with the platelets were studied. Blood was either exposed to a closed CPB circuit or obtained from patients undergoing CPB-assisted cardiac s...

  10. Effect of Temperature Downshift on the Transcriptomic Responses of Chinese Hamster Ovary Cells Using Recombinant Human Tissue Plasminogen Activator Production Culture.

    Science.gov (United States)

    Bedoya-López, Andrea; Estrada, Karel; Sanchez-Flores, Alejandro; Ramírez, Octavio T; Altamirano, Claudia; Segovia, Lorenzo; Miranda-Ríos, Juan; Trujillo-Roldán, Mauricio A; Valdez-Cruz, Norma A

    2016-01-01

    Recombinant proteins are widely used as biopharmaceuticals, but their production by mammalian cell culture is expensive. Hence, improvement of bioprocess productivity is greatly needed. A temperature downshift (TDS) from 37°C to 28-34°C is an effective strategy to expand the productive life period of cells and increase their productivity (qp). Here, TDS in Chinese hamster ovary (CHO) cell cultures, initially grown at 37°C and switched to 30°C during the exponential growth phase, resulted in a 1.6-fold increase in the qp of recombinant human tissue plasminogen activator (rh-tPA). The transcriptomic response using next-generation sequencing (NGS) was assessed to characterize the cellular behavior associated with TDS. A total of 416 (q > 0.8) and 3,472 (q > 0.9) differentially expressed transcripts, with more than a 1.6-fold change at 24 and 48 h post TDS, respectively, were observed in cultures with TDS compared to those at constant 37°C. In agreement with the extended cell survival resulting from TDS, transcripts related to cell growth arrest that controlled cell proliferation without the activation of the DNA damage response, were differentially expressed. Most upregulated genes were related to energy metabolism in mitochondria, mitochondrial biogenesis, central metabolism, and avoidance of apoptotic cell death. The gene coding for rh-tPA was not differentially expressed, but fluctuations were detected in the transcripts encoding proteins involved in the secretory machinery, particularly in glycosylation. Through NGS the dynamic processes caused by TDS were assessed in this biological system.

  11. Phytochemical Analysis and Biological Activities of Cola nitida Bark.

    Science.gov (United States)

    Dah-Nouvlessounon, Durand; Adoukonou-Sagbadja, Hubert; Diarrassouba, Nafan; Sina, Haziz; Adjanohoun, Adolphe; Inoussa, Mariam; Akakpo, Donald; Gbenou, Joachim D; Kotchoni, Simeon O; Dicko, Mamoudou H; Baba-Moussa, Lamine

    2015-01-01

    Kola nut is chewed in many West African cultures and is used ceremonially. The aim of this study is to investigate some biological effects of Cola nitida's bark after phytochemical screening. The bark was collected, dried, and then powdered for the phytochemical screening and extractions. Ethanol and ethyl acetate extracts of C. nitida were used in this study. The antibacterial activity was tested on ten reference strains and 28 meat isolated Staphylococcus strains by disc diffusion method. The antifungal activity of three fungal strains was determined on the Potato-Dextrose Agar medium mixed with the appropriate extract. The antioxidant activity was determined by DPPH and ABTS methods. Our data revealed the presence of various potent phytochemicals. For the reference and meat isolated strains, the inhibitory diameter zone was from 17.5 ± 0.7 mm (C. albicans) to 9.5 ± 0.7 mm (P. vulgaris). The MIC ranged from 0.312 mg/mL to 5.000 mg/mL and the MBC from 0.625 mg/mL to >20 mg/mL. The highest antifungal activity was observed with F. verticillioides and the lowest one with P. citrinum. The two extracts have an excellent reducing free radical activity. The killing effect of A. salina larvae was perceptible at 1.04 mg/mL. The purified extracts of Cola nitida's bark can be used to hold meat products and also like phytomedicine.

  12. Phytochemical Analysis and Biological Activities of Cola nitida Bark

    Directory of Open Access Journals (Sweden)

    Durand Dah-Nouvlessounon

    2015-01-01

    Full Text Available Kola nut is chewed in many West African cultures and is used ceremonially. The aim of this study is to investigate some biological effects of Cola nitida’s bark after phytochemical screening. The bark was collected, dried, and then powdered for the phytochemical screening and extractions. Ethanol and ethyl acetate extracts of C. nitida were used in this study. The antibacterial activity was tested on ten reference strains and 28 meat isolated Staphylococcus strains by disc diffusion method. The antifungal activity of three fungal strains was determined on the Potato-Dextrose Agar medium mixed with the appropriate extract. The antioxidant activity was determined by DPPH and ABTS methods. Our data revealed the presence of various potent phytochemicals. For the reference and meat isolated strains, the inhibitory diameter zone was from 17.5±0.7 mm (C. albicans to 9.5±0.7 mm (P. vulgaris. The MIC ranged from 0.312 mg/mL to 5.000 mg/mL and the MBC from 0.625 mg/mL to >20 mg/mL. The highest antifungal activity was observed with F. verticillioides and the lowest one with P. citrinum. The two extracts have an excellent reducing free radical activity. The killing effect of A. salina larvae was perceptible at 1.04 mg/mL. The purified extracts of Cola nitida’s bark can be used to hold meat products and also like phytomedicine.

  13. Estabishment of A Human Liver Cancer Cell Line Transfected with IL-2 cDNA and Its Biologic Activity

    Institute of Scientific and Technical Information of China (English)

    孙跃明; 王学浩; 杜竞辉

    2001-01-01

    Objective To obtain IL-2 gene transfected human liver cancer cells and study IL-2 expression and its biologic activity in vivo. Methods Human liver cancer cells SMMC-7721 were cocultured with recombinant retroviral vector LNC-IL-2,and screening was performed in G418 medium.The exogenous IL-2 cDNA at the DNA,RNA,and protein levels were determined by using dot hybridization,PR-PCR and MTT methods respectively.The tumorigenesis and antitumorigenesis of the screened liver cancer cell with subcutaneous injection in nude mice were observed. Results and Conclusion The IL-2 cDNA was successfully integrated into SMMC-7721 cell genomic DNA and continuously expressed for more than 88 days.Subcutaneous vaccination of the nude mice with transfected cells revealed an obvious suppression of its tumorigenicity,and could induce antitumor activity in vivo.

  14. DISCOVERY OF THE RECOMBINING PLASMA IN THE SOUTH OF THE GALACTIC CENTER: A RELIC OF THE PAST GALACTIC CENTER ACTIVITY?

    Energy Technology Data Exchange (ETDEWEB)

    Nakashima, S.; Nobukawa, M.; Uchida, H.; Tanaka, T.; Tsuru, T. G.; Koyama, K. [Department of Physics, Graduate School of Science, Kyoto University, Kitashirakawa Oiwake-cho, Sakyo-ku, Kyoto 606-8502 (Japan); Murakami, H. [Department of Information Science, Faculty of Liberal Arts, Tohoku Gakuin University 2-1-1 Tenjinzawa, Izumi-ku, Sendai, Miyagi 981-3193 (Japan); Uchiyama, H., E-mail: shinya@cr.scphys.kyoto-u.ac.jp [Science Education, Faculty of Education, Shizuoka University, 836 Ohya, Suruga-ku, Shizuoka 422-8529 (Japan)

    2013-08-10

    We report Suzaku results for soft X-ray emission to the south of the Galactic center (GC). The emission (hereafter {sup G}C South{sup )} has an angular size of {approx}42' Multiplication-Sign 16' centered at (l, b) {approx} (0. Degree-Sign 0, - 1. Degree-Sign 4) and is located in the largely extended Galactic ridge X-ray emission (GRXE). The X-ray spectrum of GC South exhibits emission lines from highly ionized atoms. Although the X-ray spectrum of the GRXE can be well fitted with a plasma in collisional ionization equilibrium (CIE), that of GC South cannot be fitted with a plasma in CIE, leaving hump-like residuals at {approx}2.5 and 3.5 keV, which are attributable to the radiative recombination continua of the K-shells of Si and S, respectively. In fact, GC South spectrum is well fitted with a recombination-dominant plasma model; the electron temperature is 0.46 keV while atoms are highly ionized (kT = 1.6 keV) in the initial epoch, and the plasma is now in a recombining phase at a relaxation scale (plasma density Multiplication-Sign elapsed time) of 5.3 Multiplication-Sign 10{sup 11} s cm{sup -3}. The absorption column density of GC South is consistent with that toward the GC region. Thus, GC South is likely to be located in the GC region ({approx}8 kpc distance). The size of the plasma, the mean density, and the thermal energy are estimated to be {approx}97 pc Multiplication-Sign 37 pc, 0.16 cm{sup -3}, and 1.6 Multiplication-Sign 10{sup 51} erg, respectively. We discuss possible origins of the recombination-dominant plasma as a relic of past activity in the GC region.

  15. Efficient production of active recombinant Candida rugosa LIP3 lipase in Pichia pastoris and biochemical characterization of the purified enzyme.

    Science.gov (United States)

    Chang, Shu-Wei; Lee, Guan-Chiun; Shaw, Jei-Fu

    2006-08-09

    Candida rugosa lipase (CRL), an important industrial enzyme, possesses several different isoforms encoded by the high-identity lip gene family (lip1 to lip7). In this study, an additional N-terminal peptide in front of the lip3 gene was removed by PCR, and the 18 nonuniversal serine codons (CTG) of the lip3 gene were converted into universal serine codons (TCT) by means of an overlap extension PCR-based multiple-site-directed mutagenesis to express an active recombinant LIP3 in the yeast Pichia pastoris. The regional synthetic DNA fragment (339 bp) is first recombined by primer assembly with 20 overlapping nucleotides, followed by specific overlap extension PCR with outside primers containing restriction enzyme sites for directional cloning into the pGAPZalphaC vector. The results show that the production yield (0.687 unit/mL) of N-fused lip3 (nflip3) has an overall improvement of 69-fold relative to that (0.01 unit/mL) of lip3 and of 52-fold (0.47 unit/mL) of codon-optimized lip3 (colip3) relative to that (0.01 unit/mL) of non-codon-optimized lip3 (lip3), with the cultivation time set at 5 days. This finding demonstrates that the reservation of the N terminus and the regional codon optimization of the lip3 gene fragment at the 5' end can greatly increase the expression level of recombinant LIP3 in the P. pastoris system. The purified recombinant LIP3 shows distinct biochemical properties compared with other isoforms.

  16. INFLUENCE OF BIOLOGICALLY ACTIVE AGENTS ON A STRUCTURAL STATE AND THE ENZYMATIC ACTIVITY OF BLACK ORDINARY CARBONATED SOIL

    Directory of Open Access Journals (Sweden)

    Lychman V. A.

    2014-04-01

    Full Text Available The results of a long-term research of the influence of various biologically active agents (a humic preparation Lignogumat and microbiological Baikal EM fertilizer on a structural state and the enzymatic activity of ordinary carbonated black soil are presented. It has been established that biologically active substances contribute to increased enzymatic activity, humus and improve the soil structure

  17. Phenolic Compounds Characterization and Biological Activities of Citrus aurantium Bloom

    Directory of Open Access Journals (Sweden)

    Armin Oskoueian

    2012-01-01

    Full Text Available Citrus plants are known to possess beneficial biological activities for human health. In addition, ethnopharmacological application of plants is a good tool to explore their bioactivities and active compounds. This research was carried out to evaluate the phenolic and flavonoid analysis, antioxidant properties, anti inflammatory and anti cancer activity of Citrus aurantium bloom. The total phenolics and flavonoids results revealed that methanolic extract contained high total phenolics and flavonoids compared to ethanolic and boiling water extracts. The obtained total phenolics value for methanolic Citrus aurantium bloom extract was 4.55 ± 0.05 mg gallic acid equivalent (GAE/g dry weight (DW, and for total flavonoids it was 3.83 ± 0.05 mg rutin equivalent/g DW. In addition, the RP-HPLC analyses of phenolics and flavonoids indicated the presence of gallic acid, pyrogallol, syringic acid, caffeic acid, rutin, quercetin and naringin as bioactive compounds. The antioxidant activity of Citrus aurantium bloom were examined by the 1,1-diphenyl-2-picryl-hydrazyl (DPPH assay and the ferric reducing/antioxidant potential (FRAP. The free radical scavenging and ferric reducing power activities were higher for the methanolic extract of Citrus aurantium bloom at a concentration of 300 μg/mL, with values of 55.3% and 51.7%, respectively, as compared to the corresponding boiling water and ethanolic extracts, but the activities were lower than those of antioxidant standards such as BHT and α-tocopherol. Furthermore, the anti-inflammatory result of methanolic extract showed appreciable reduction in nitric oxide production of stimulated RAW 264.7 cells at the presence of plant extract. Apart from that, the anticancer activity of the methanolic extract was investigated in vitro against human cancer cell lines (MCF-7; MDA-MB-231, human colon adenocarcinoma (HT-29 and Chang cell as a normal human hepatocyte. The obtained result demonstrated the moderate to

  18. Heterologous Expression and Delivery of Biologically Active Exendin-4 by Lactobacillus paracasei L14

    Science.gov (United States)

    Zeng, Zhu; Yu, Rui; Zuo, Fanglei; Zhang, Bo; Peng, Deju; Ma, Huiqin; Chen, Shangwu

    2016-01-01

    Exendin-4, a glucagon-like protein-1 (GLP-1) receptor agonist, is an excellent therapeutic peptide drug for type 2 diabetes due to longer lasting biological activity compared to GLP-1. This study explored the feasibility of using probiotic Lactobacillus paracasei as an oral vector for recombinant exendin-4 peptide delivery, an alternative to costly chemical synthesis and inconvenient administration by injection. L. paracasei transformed with a plasmid encoding the exendin-4 gene (L. paracasei L14/pMG76e-exendin-4) with a constitutive promotor was successfully constructed and showed efficient secretion of exendin-4. The secreted exendin-4 significantly enhanced insulin secretion of INS-1 β-cells, along with an increment in their proliferation and inhibition of their apoptosis, corresponding to the effect of GLP-1 on these cells. The transcription level of the pancreatic duodenal homeobox-1 gene (PDX-1), a key transcription factor for cellular insulin synthesis and secretion, was upregulated by the treatment with secreted exendin-4, paralleling the upregulation of insulin gene expression. Caco-2 cell monolayer permeability assay showed a 34-fold increase in the transport of exendin-4 delivered by L. paracasei vs. that of free exendin-4 (control), suggesting effective facilitation of exendin-4 transport across the intestinal barrier by this delivery system. This study demonstrates that the probiotic Lactobacillus can be engineered to secrete bioactive exendin-4 and facilitate its transport through the intestinal barrier, providing a novel strategy for oral exendin-4 delivery using this lactic acid bacterium. PMID:27764251

  19. Prosequence switching: an effective strategy to produce biologically active E. coli heat-stable enterotoxin STh.

    Science.gov (United States)

    Weiglmeier, Philipp R; Berkner, Hanna; Seebahn, Angela; Vogel, Nico; Schreiber, Rainer; Wöhrl, Birgitta M; Schwarzinger, Stephan; Rösch, Paul

    2014-01-01

    Enterotoxigenic Escherichia coli (ETEC) infections account for the majority of cases of acute secretory diarrhea. The causative agents are enterotoxins secreted by ETEC, among them is the heat-stable enterotoxin, STh. STh is a 19-amino acid peptide containing three disulfide bonds that stimulates fluid secretion in the bowel by binding to the receptor domain of intestinal guanylyl cyclase C (GC-C). Since GC-C agonists have pharmacologic potential for diagnosis and treatment of disorders such as constipation-predominant irritable bowel syndrome (IBS-C), chronic constipation, and colorectal carcinoma, it is crucial to develop methods for the large-scale production of STh and related peptides. Here, we present a strategy for recombinant expression of STh that relies on the use of the prosequence of human uroguanylin to support proper folding and disulfide bond formation. The chimeric protein CysCys-STh consisting of the propeptide of uroguanylin as N-terminus and the STh peptide as C-terminus was expressed in E. coli, and an efficient purification protocol was developed. Trypsin digestion of this protein released the enterotoxin which could be obtained in high purity. NMR and mass spectrometry confirmed the identity and homogeneity of the toxin, and its biological activity was confirmed by a cell-based in vivo assay. The expression scheme introduced here represents a cost-efficient and scalable way of STh production.

  20. Activity of the Human Rhinovirus 3C Protease Studied in Various Buffers, Additives and Detergents Solutions for Recombinant Protein Production.

    Science.gov (United States)

    Ullah, Raheem; Shah, Majid Ali; Tufail, Soban; Ismat, Fouzia; Imran, Muhammad; Iqbal, Mazhar; Mirza, Osman; Rhaman, Moazur

    2016-01-01

    Proteases are widely used to remove affinity and solubility tags from recombinant proteins to avoid potential interference of these tags with the structure and function of the fusion partner. In recent years, great interest has been seen in use of the human rhinovirus 3C protease owing to its stringent sequence specificity and enhanced activity. Like other proteases, activity of the human rhinovirus 3C protease can be affected in part by the buffer components and additives that are generally employed for purification and stabilization of proteins, hence, necessitate their removal by tedious and time-consuming procedures before proteolysis can occur. To address this issue, we examined the effect of elution buffers used for common affinity based purifications, salt ions, stability/solubility and reducing agents, and detergents on the activity of the human rhinovirus 3C protease using three different fusion proteins at 4°C, a temperature of choice for purification of many proteins. The results show that the human rhinovirus 3C protease performs better at 4°C than the frequently used tobacco etch virus protease and its activity was insensitive to most of the experimental conditions tested. Though number of fusion proteins tested is limited, we expect that these finding will facilitate the use of the human rhinovirus 3C protease in recombinant protein production for pharmaceutical and biotechnological applications.

  1. Activity of the Human Rhinovirus 3C Protease Studied in Various Buffers, Additives and Detergents Solutions for Recombinant Protein Production.

    Directory of Open Access Journals (Sweden)

    Raheem Ullah

    Full Text Available Proteases are widely used to remove affinity and solubility tags from recombinant proteins to avoid potential interference of these tags with the structure and function of the fusion partner. In recent years, great interest has been seen in use of the human rhinovirus 3C protease owing to its stringent sequence specificity and enhanced activity. Like other proteases, activity of the human rhinovirus 3C protease can be affected in part by the buffer components and additives that are generally employed for purification and stabilization of proteins, hence, necessitate their removal by tedious and time-consuming procedures before proteolysis can occur. To address this issue, we examined the effect of elution buffers used for common affinity based purifications, salt ions, stability/solubility and reducing agents, and detergents on the activity of the human rhinovirus 3C protease using three different fusion proteins at 4°C, a temperature of choice for purification of many proteins. The results show that the human rhinovirus 3C protease performs better at 4°C than the frequently used tobacco etch virus protease and its activity was insensitive to most of the experimental conditions tested. Though number of fusion proteins tested is limited, we expect that these finding will facilitate the use of the human rhinovirus 3C protease in recombinant protein production for pharmaceutical and biotechnological applications.

  2. Characterisation of aroma profiles of commercial sufus by odour activity value, gas chromatography-olfactometry, aroma recombination and omission studies.

    Science.gov (United States)

    Xiao, Zuobing; Shang, Yi; Chen, Feng; Niu, Yunwei; Gu, Yongbo; Liu, Shengjiang; Zhu, Jiancai

    2015-01-01

    Sufu is a solid-state fermented product made from soya beans. For the sake of quality control and regulation purposes, it is essential to be able to identify key odorants of various commercial sufus. To identify the aroma-active compounds in sufus, gas chromatography-olfactometry/aroma extract dilution analysis (GC-O/AEDA) was performed, and odour activity value (OAV) was estimated. The correlations between aroma profiles and identified aroma-active compounds were also investigated by principal component analysis. Results showed that 35 aroma-active compounds were detected through OAV calculation, while 28 compounds were identified by using GC-O/AEDA. Quantitative descriptive analysis revealed that aroma recombination model based on OAV calculation was more similar to original sufu in terms of aroma comparing to aroma recombination model based on GC-O/AEDA. Omission experiments further confirmed that the aroma compounds, such as ethyl butanoate, ethyl 2-methylbutanoate, ethyl hexanoate, (E,E)-2,4-decadienal and 2,6-dimethylpyrazine, contributed most significantly to the characteristic aroma of a commercial sufu.

  3. A Conceptual Framework for Organizing Active Learning Experiences in Biology Instruction

    Science.gov (United States)

    Gardner, Joel; Belland, Brian R.

    2012-01-01

    Introductory biology courses form a cornerstone of undergraduate instruction. However, the predominantly used lecture approach fails to produce higher-order biology learning. Research shows that active learning strategies can increase student learning, yet few biology instructors use all identified active learning strategies. In this paper, we…

  4. Tobacco as biofactory for biologically active hPL production: a human hormone with potential applications in type-1 diabetes.

    Science.gov (United States)

    Urreta, Iratxe; Oyanguren, Iñigo; Castañón, Sonia

    2011-08-01

    Human placental lactogen (hPL) is a peptidic hormone that belongs to the short list of growth factors that could treat type-1 diabetes through pancreatic islet transplantation. Placental lactogen has the capacity to improve islet survival and function before or after transplantation. In this study, transgenic tobacco plants were used as a novel expression system for the production of recombinant hPL protein (rhPL). The expression vector pNEKhPL2 containing hPL cDNA was introduced into tobacco plants; the transcriptional activity was confirmed by real-time PCR, and the rhPL levels reached 1% of the total soluble protein (TSP) content in plants cultivated in the greenhouse. In vitro bioassays using the rat insulinoma (INS-1) cell line showed that recombinant protein was able to induce cell proliferation and activate the JAK-2/STAT-5 signal transduction pathway, demonstrating that plant cells can produce the biologically active hPL protein. To further characterize the plant expression system for hPL production, we analyzed the stability of the protein during the life cycle of tobacco plants as well as the transmission of the transgenic trait to the progeny. The recombinant protein was stably accumulated in young leaves, reaching the maximum level in the first month (6.51 μg/g of fresh weight), but showing a decreasing trend of 26% from the initial sampling time until the end of plant's life cycle. The progeny of the selected pNEKhPL2 plant showed in vitro expression levels of up to 1.1% of TSP. Our results therefore indicate that transgenic plants are a suitable expression system for hPL production.

  5. Synthesis, Characterization and Biological Activities of Organotin (IV Methylcyclohexyldithiocarbamate Compounds

    Directory of Open Access Journals (Sweden)

    Normah Awang

    2011-01-01

    Full Text Available Problem statement: The growing interest in the chemistry of sulphur donor ligands are due to their encouraging anticancer, antibacterial and antifungal activities as well as their widespread industrial application. Dithiocarbamates belong to this class and much attention has been paid to them. Approach: Novel organotin compounds with the molecular formula RmSn[S2CN(CH3(C6H11]4-m (where m = 2, R = CH3, C2H5; m = 3, R = C6H5 have been synthesized using in situ method. These compounds were characterized by elemental analysis, IR, 1H and 13C NMR spectroscopy. Results: Elemental analysis revealed that all compounds were of good purity. Infrared spectra of the compounds showed that the thioureide ν(C-N band was in the region 1450-1500 cm−1. The unsplitting band of ν(C-S in the region 974-979 cm−1 indicated the bidentate nature of the chelated dithiocarbamato legends. The 13C NMR chemical shift of the carbon atom in the N-CS2 group appeared in the range of 196.29-199.82 ppm. Single crystal analysis from one of these compounds showed that the chelating mode of the dithiocarbamate groups was isobidentate. These compounds have been screened for antibacterial activity against four bacteria; Staphylococcus aureus, Salmonella typhimurium, Pseudomonas aeruginosa and Bacillus subtilis. Only one of these compounds shows promising results against S. aureus and S. typhi. Cytotoxicity screening on human leukemic promyelocyte HL-60 cells found that two of these compounds were very active with CD50 values of 0.87 and 0.18 µg mL−1. Conclusion: The studied compounds were found to have the potential in biological activity especially in cytotoxicity where this possibly can be used for clinical trials after further research.

  6. [Composition and content of biologically active substances in rose hips].

    Science.gov (United States)

    Dubtsova, G N; Negmatulloeva, R N; Bessonov, V V; Baĭkov, V G; Sheviakova, L V; Makhova, N N; Perederiaev, O I; Bogachuk, M N; Baĭgarin, E K

    2012-01-01

    The paper studies the chemical composition of the powders obtained from the pulp with the skins and seeds of fruits of wild rose hips. Research results have shown that the main fraction of the powder is dietary fiber, powder of seeds of insoluble fiber in 1,6 and 2,3 higher than in the powder of the fruit with a thin skin and pulp, respectively. The greatest amount of carbohydrates and protein found in powders and pulp of the fruit with a thin skin, and lipids predominate in the powder from the seeds. Found that the lipid powder rosehip richest in oleic, linoleic and linolenic acids, the share of oleic acid has 6,4-19,2%, linoleic and linolenic 19,7-45,8 and 23,3-33,9% of the amount of fatty acids. Lipids powders of hips and seeds of rose have higher levels of essential linoleic acid and powder from the pulp with the skins - linolenic acid. In the study established the presence of sterols 7 fractions, the predominant of which is the beta-sitosterol. In the powder from the pulp with the skins found the greatest amount of ascorbic acid, carotenoids, and the powder of seeds - vitamin E. Carotenoids in powders are beta-carotene and lycopene. The high content of ascorbic acid, vitamin E and carotenoids in powder from wild rose hips makes them a good source of antioxidants. Therefore, we studied the possibility of using vegetable powders obtained from hips of wild rose, to enrich biologically active substances such as vitamins C, E and carotenoids, food supply, particularly of health care use. Rosehip powder from the pulp with the skins had the highest antioxidant activity, antioxidant activity of hips powders was 74% of the activity of powder from the pulp with the skins, the lowest antioxidant activity was observed in the powder from the wild rose seeds. That's way, based on the analysis of the chemical composition of rose hip powder found high levels they ascorbic acid, carotenoids, flavonoids,found their high antioxidant activity. It allows to recommend powders

  7. Expression and Purification of Active Recombinant Cathepsin C (Dipeptidyl Aminopeptidase I of Kuruma Prawn Marsupenaeus japonicus in Insect Cells

    Directory of Open Access Journals (Sweden)

    Gao-Feng Qiu

    2009-01-01

    Full Text Available Cathepsin C (CTSC is a lysosomal cysteine protease belonging to the papain superfamily. Our previous study showed that CTSC precursor (zymogen is localized exclusively in cortical rods (CRs of mature oocyte in the kuruma prawn Marsupenaeus japonicus, suggesting that CTSC might have roles on regulating release and/or formation of a jelly layer. In this study, enzymically active CTSC of the kuruma prawn was prepared by recombinant expression in the High Five insect cell line. The recombinant enzyme with a polyhistidine tag at its C-terminus was considered to be initially secreted into the culture medium as an inactive form of zymogen, because Western blot with anti-CTSC antibody detected a 51 kDa protein corresponding to CTSC precursor. After purification by affinity chromatography on nickel-iminodiacetic acid resin, the enzyme displayed three forms of 51, 31, and 30 kDa polypeptides. All of the forms can be recognized by antiserum raised against C-terminal polyhistidine tag, indicating that the 31 and 30 kDa forms were generated from 51 kDa polypeptide by removal of a portion of the N-terminus of propeptide. Following activation at pH 5.5 and 37∘C for 40 hours under native conditions, the recombinant CTSC (rCTSC exhibited increased activity against the synthetic substrate Gly-Phe-β-naphthylamide and optimal pH at around 5. The purified rCTSC will be useful for further characterization of its exact physiological role on CRs release and/or formation of a jelly layer in kuruma prawn.

  8. Factors influencing clinical outcomes of acute ischemic stroke treated with intravenous recombinant tissue plasminogen activator

    Institute of Scientific and Technical Information of China (English)

    HUANG Yin-hui; ZHUO Shi-tu; CHEN Ya-fang; LI Ming-mei; LIN You-yu; YANG Mei-li; CHEN Zhen-jie

    2013-01-01

    Background Thrombolysis with recombinant tissue plasminogen activator (rt-PA) has gained international recognition,clinical outcomes following this thrombolytic therapy varied from patient to patient.Factors affecting clinical outcomes have not been well understood yet,so this retrospective case-control study aimed to investigate factors that may influence clinical outcomes of acute ischemic stroke treated with intravenous rt-PA.Methods One hundred and one patients with acute ischemic stroke who received intravenous rt-PA thrombolysis within 4.5 hours from disease onset were included.Patients were divided into good or poor outcome group according to modified Rankin Scale (mRS) score,good outcome group:mRS score of 0-1; poor outcome group:mRS of 2-6.Stroke characteristics were compared between the two groups.Factors for stroke outcomes were analyzed via univariate analysis and Logistic regression.Results Of the 101 patients studied,patients in good outcome group (n=55) were significantly younger than patients in poor outcome group (n=46,(62.82±14.25) vs.(68.81±9.85) years,P=0.029).Good outcome group had fewer patients with diabetic history (9.09% vs.28.26%,P=0.012),fewer patients with leukoaraiosis (7.27% vs.28.26%,P=0.005) and presented with lower blood glucose level ((5.72±1.76) vs.(6.72±1.32) mmol/L,P=0.012),lower systolic blood pressure level ((135.45±19.36) vs.(148.78±19.39) mmHg,P=0.003),lower baseline NIHSS score (12.02±5.26 vs.15.78±4.98,P=0.002) and shorter onset-to-treatment time (OTT) ((2.38±1.21) vs.(2.57±1.03) hours,P=0.044) than poor outcome group.Logistic regression analysis showed that absence of diabetic history (odds ratio (OR) 0.968 (95% CI 0.941-0.996)),absence of leukoaraiosis (OR 0.835 (95% CI 0.712-0.980)),lower baseline NIHSS score (OR 0.885 (95% CI 0.793-0.989)),lower pre-thrombolysis systolic blood pressure (OR 0.962 (95% CI 0.929-0.997)),and lower blood glucose level (OR 0.699 (95% CI 0.491-0.994)) before

  9. Oat Phytochrome Is Biologically Active in Transgenic Tomatoes.

    Science.gov (United States)

    Boylan, M. T.; Quail, P. H.

    1989-08-01

    To determine the functional homology between phytochromes from evolutionarily divergent species, we used the cauliflower mosaic virus 35S promoter to express a monocot (oat) phytochrome cDNA in a dicot plant (tomato). Immunoblot analysis shows that more than 50% of the transgenic tomato plants synthesize the full-length oat phytochrome polypeptide. Moreover, leaves of light-grown transgenic plants contain appreciably less oat phytochrome than leaves from dark-adapted plants, and etiolated R1 transgenic seedlings have higher levels of spectrally active phytochrome than wild-type tomato seedlings in direct proportion to the level of immunochemically detectable oat polypeptide present. These data suggest that the heterologous oat polypeptide carries a functional chromophore, allowing reversible photoconversion between the two forms of the molecule, and that the far-red absorbing form (Pfr) is recognized and selectively degraded by the Pfr-specific degradative machinery in the dicot cell. The overexpression of oat phytochrome has pleiotropic, phenotypic consequences at all major phases of the life cycle. Adult transgenic tomato plants expressing high levels of the oat protein tend to be dwarfed, with dark green foliage and fruits. R1 transgenic seedlings have short hypocotyls with elevated anthocyanin contents. We conclude that a monocot phytochrome can be synthesized and correctly processed to a biologically active form in a dicot cell, and that the transduction pathway components that interact with the photoreceptor are evolutionarily conserved.

  10. Biological Activities of Phosphocitrate: A Potential Meniscal Protective Agent

    Directory of Open Access Journals (Sweden)

    Yubo Sun

    2013-01-01

    Full Text Available Phosphocitrate (PC inhibited meniscal calcification and the development of calcium crystal-associated osteoarthritis (OA in Hartley guinea pigs. However, the mechanisms remain elusive. This study sought to examine the biological activities of PC in the absence of calcium crystals and test the hypothesis that PC is potentially a meniscal protective agent. We found that PC downregulated the expression of many genes classified in cell proliferation, ossification, prostaglandin metabolic process, and wound healing, including bloom syndrome RecQ helicase-like, cell division cycle 7 homolog, cell division cycle 25 homolog C, ankylosis progressive homolog, prostaglandin-endoperoxide synthases-1/cyclooxygenase-1, and plasminogen activator urokinase receptor. In contrast, PC stimulated the expression of many genes classified in fibroblast growth factor receptor signaling pathway, collagen fibril organization, and extracellular structure organization, including fibroblast growth factor 7, collagen type I, alpha 1, and collagen type XI, alpha 1. Consistent with its effect on the expression of genes classified in cell proliferation, collagen fibril organization, and ossification, PC inhibited the proliferation of OA meniscal cells and meniscal cell-mediated calcification while stimulating the production of collagens. These findings indicate that PC is potentially a meniscal-protective agent and a disease-modifying drug for arthritis associated with severe meniscal degeneration.

  11. Estimates of iodine in biological materials by epithermal neutron activation analysis

    Energy Technology Data Exchange (ETDEWEB)

    Sato, T. (Tokyo Metropolitan Inst. for Neurosciences, Fuchu (Japan)); Kato, T. (Tohoku Univ., Sendai (Japan). Coll. of General Education)

    1982-01-01

    Iodine abundances in NBS biological SRMs and various organs of rats were evaluated by epithermal neutron activation analysis with a boron carbide filter. The detectability of iodine in different biological materials by this method is discussed.

  12. Effect of a biological activated carbon filter on particle counts

    Institute of Scientific and Technical Information of China (English)

    Su-hua WU; Bing-zhi DONG; Tie-jun QIAO; Jin-song ZHANG

    2008-01-01

    Due to the importance of biological safety in drinking water quality and the disadvantages which exist in traditional methods of detecting typical microorganisms such as Cryptosporidium and Giardia,it is necessary to develop an alternative.Particle counts is a qualitative measurement of the amount of dissolved solids in water.The removal rate of particle counts was previously used as an indicator of the effectiveness of a biological activated carbon(BAC)filter in removing Cryptosporidium and Giardia.The particle counts in a BAC filter effluent over one operational period and the effects of BAC filter construction and operational parameters were investigated with a 10 m3/h pilot plant.The results indicated that the maximum particle count in backwash remnant water was as high as 1296 count/ml and it needed about 1.5 h to reduce from the maximum to less than 50 count/ml.During the standard filtration period,particle counts stay constant at less than 50 count/ml for 5 d except when influ-enced by sand filter backwash remnant water.The removal rates of particle counts in the BAC filter are related to characteristics of the carbon.For example,a columned carbon and a sand bed removed 33.3% and 8.5% of particles,respectively,while the particle counts in effluent from a cracked BAC filter was higher than that of the influent.There is no significant difference among particle removal rates with different filtration rates.High post-ozone dosage(>2 mg/L)plays an important role in particle count removal;when the dosage was 3 mg/L,the removal rates by carbon layers and sand beds decreased by 17.5% and increased by 9.5%,respectively,compared with a 2 mg/L dosage.

  13. A designer hyper interleukin 11 (H11 is a biologically active cytokine

    Directory of Open Access Journals (Sweden)

    Dams-Kozlowska Hanna

    2012-03-01

    Full Text Available Abstract Background Interleukin 11 (IL-11 is a pleiotropic cytokine with anti-apoptotic, anti-inflammatory and hematopoietic potential. The IL-11 activity is determined by the expression of the IL-11R receptor alpha (IL-11Rα and the signal transducing subunit β (gp130 on the cell membrane. A recombinant soluble form of the IL-11Rα (sIL-11Rα in combination with IL-11 acts as an agonist on cells expressing the gp130 molecule. We constructed a designer cytokine Hyper IL-11 (H11, which is exclusively composed of naturally existing components. It contains the full length sIL-11Rα connected with the mature IL-11 protein using their natural sequences only. Such a construct has two major advantages: (i its components are as close as possible to the natural forms of both proteins and (ii it lacks an artificial linker what should avoid induction of antibody production. Results The H11 construct was generated, the protein was produced in a baculovirus expression system and was then purified by using ion exchange chromatography. The H11 protein displayed activity in three independent bioassays, (i it induced acute phase proteins production in HepG2 cells expressing IL-11, IL-11Rα and gp130, (ii it stimulated the proliferation of B9 cells (cells expressing IL-11Rα and gp130 and (iii proliferation of Baf/3-gp130 cells (cells not expressing IL-11 and IL-11Rα but gp130. Moreover, the preliminary data indicated that H11 was functionally distinct from Hyper-IL-6, a molecule which utilizes the same homodimer of signal transducing receptor (gp130. Conclusions The biologically active H11 may be potentially useful for treatment of thrombocytopenia, infertility, multiple sclerosis, cardiovascular diseases or inflammatory disorders.

  14. Actinobacteria from arid and desert habitats: diversity and biological activity

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    Joachim eWink

    2016-01-01

    Full Text Available Abstract The lack of new antibiotics in the pharmaceutical pipeline guides more and more researchers to leave the classical isolation procedures and to look in special niches and ecosystems. Bioprospecting of extremophilic Actinobacteria through mining untapped strains and avoiding resiolation of known biomolecules is among the most promising strategies for this purpose. With this approach, members of acidtolerant, alkalitolerant, psychrotolerant, thermotolerant, halotolerant and xerotolerant Actinobacteria have been obtained from respective habitats. Among these, little survey exists on the diversity of Actinobacteria in arid areas, which are often adapted to relatively high temperatures, salt concentrations, and radiation. Therefore, arid and desert habitats are special ecosystems which can be recruited for the isolation of uncommon Actinobacteria with new metabolic capability.At the time of this writing, members of Streptomyces, Micromonospora, Saccharothrix, Streptosporangium, Cellulomonas, Amycolatopsis, Geodermatophilus, Lechevalieria, Nocardia and Actinomadura are reported from arid habitats. However, metagenomic data present dominant members of the communities in desiccating condition of areas with limited water availability that are not yet isolated. Furthermore, significant diverse types of polyketide synthase (PKS and nonribosomal peptide synthetase (NRPS genes are detected in xerophilic and xerotolerant Actinobacteria and some bioactive compounds are reported from them. Rather than pharmaceutically active metabolites, molecules with protection activity against drying such as Ectoin and Hydroxyectoin with potential application in industry and agriculture have also been identified from xerophilic Actinobacteria. In addition, numerous biologically active small molecules are expected to be discovered from arid adapted Actinobacteria in the future. In the current survey, the diversity and biotechnological potential of Actinobacteria

  15. [Studies on acetylspiramycin. II. Biological activities of spiramycin components].

    Science.gov (United States)

    Kondo, A; Sato, K; Shuto, K; Yamashita, K; Ichikawa, S; Takahashi, K; Kita, K; Nishiie, Y; Sano, H; Yamaguchi, K

    1990-09-01

    Acetylspiramycin (ASPM) was fractionated using high performance liquid chromatography (HPLC). The peak fractions were named F1 to F7 successively in order of increasing retention times (Rt), i.e., increasing hydrophobicity, and studied for 1) antibacterial activities (MIC), 2) antibacterial potency against Bacillus subtilis ATCC 6633, 3) therapeutic effect on mice infected with Streptococcus pneumoniae III, Staphylococcus aureus Smith, 4) acute toxicity by i.p. administration to mice (LD50) and 5) cytotoxicities to fibroblasts derived from Chinese-hamster lung (CHL), cow pulmonary artery endothelial cells (CPAE) and rat hepatic cells. The results obtained are summarized below. 1. Components F1 and 4'-acetylspiramycin F2 had significantly different biological activities from those of other components: F1 showed the lowest antibacterial potency of 492 micrograms (potency)/mg, F2 showed the highest antibacterial potency of 2,040 micrograms (potency)/mg and correspondingly the lowest LD50 value of 692 mg/kg (the highest toxicity). The therapeutic effect of F2 on infections in mice was found to be the second smallest and was superior only to that of F1. The LD50 value of F1 was 1,200 mg/kg and similar to that of ASPM. 2. Antibacterial potencies of F3, F4, F5 and F6 were 1,165, 1,266, 1,374 and 1,530 micrograms (potency)/mg, respectively; fraction with the higher antibacterial activities corresponded to the longer retention times, i.e., the greater hydrophobicities. The most hydrophobic component, F7, 3-propionyl-3",4"-diacetylspiramycin, however, showed a low antibacterial potency of 1,085 micrograms (potency)/mg, next to the lowest one, F1, a fact which was in contradiction to with the sequential relation between hydrophobicities and potencies from F3 to F6.(ABSTRACT TRUNCATED AT 250 WORDS)

  16. A novel cold-active lipase from Candida albicans: cloning, expression and characterization of the recombinant enzyme.

    Science.gov (United States)

    Lan, Dong-Ming; Yang, Ning; Wang, Wen-Kai; Shen, Yan-Fei; Yang, Bo; Wang, Yong-Hua

    2011-01-01

    A novel lipase gene lip5 from the yeast Candida albicans was cloned and sequenced. Alignment of amino acid sequences revealed that 86-34% identity exists with lipases from other Candida species. The lipase and its mutants were expressed in the yeast Pichia pastoris, where alternative codon usage caused the mistranslation of 154-Ser and 293-Ser as leucine. 154-Ser to leucine resulted in loss of expression of Lip5, and 293-Ser to leucine caused a marked reduction in the lipase activity. Lip5-DM, which has double mutations that revert 154 and 293 to serine residues, showed good lipase activity, and was overexpressed and purified by (NH(4))(2)SO(4) precipitation and ion-exchange chromatography. The pure Lip5-DM was stable at low temperatures ranging from 15-35 °C and pH 5-9, with the optimal conditions being 15-25 °C and pH 5-6. The activation energy of recombinant lipase was 8.5 Kcal/mol between 5 and 25 °C, suggesting that Lip5-DM was a cold-active lipase. Its activity was found to increase in the presence of Zn(2+), but it was strongly inhibited by Fe(2+), Fe(3+), Hg(2+) and some surfactants. In addition, the Lip5-DM could not tolerate water-miscible organic solvents. Lip5-DM exhibited a preference for the short-and medium-chain length p-nitrophenyl (C4 and C8 acyl group) esters rather than the long chain length p-nitrophenyl esters (C12, C16 and C18 acyl group) with highest activity observed with the C8 derivatives. The recombinant enzyme displayed activity toward triacylglycerols, such as olive oil and safflower oil.

  17. A Novel Cold-Active Lipase from Candida albicans: Cloning, Expression and Characterization of the Recombinant Enzyme

    Directory of Open Access Journals (Sweden)

    Dong-Ming Lan

    2011-06-01

    Full Text Available A novel lipase gene lip5 from the yeast Candida albicans was cloned and sequenced. Alignment of amino acid sequences revealed that 86–34% identity exists with lipases from other Candida species. The lipase and its mutants were expressed in the yeast Pichia pastoris, where alternative codon usage caused the mistranslation of 154-Ser and 293-Ser as leucine. 154-Ser to leucine resulted in loss of expression of Lip5, and 293-Ser to leucine caused a marked reduction in the lipase activity. Lip5-DM, which has double mutations that revert 154 and 293 to serine residues, showed good lipase activity, and was overexpressed and purified by (NH42SO4 precipitation and ion-exchange chromatography. The pure Lip5-DM was stable at low temperatures ranging from 15–35 °C and pH 5–9, with the optimal conditions being 15–25 °C and pH 5–6. The activation energy of recombinant lipase was 8.5 Kcal/mol between 5 and 25 °C, suggesting that Lip5-DM was a cold–active lipase. Its activity was found to increase in the presence of Zn2+, but it was strongly inhibited by Fe2+, Fe3+, Hg2+ and some surfactants. In addition, the Lip5-DM could not tolerate water-miscible organic solvents. Lip5-DM exhibited a preference for the short- and medium-chain length p-nitrophenyl (C4 and C8 acyl group esters rather than the long chain length p-nitrophenyl esters (C12, C16 and C18 acyl group with highest activity observed with the C8 derivatives. The recombinant enzyme displayed activity toward triacylglycerols, such as olive oil and safflower oil.

  18. Bacterial Recombineering: Genome Engineering via Phage-Based Homologous Recombination.

    Science.gov (United States)

    Pines, Gur; Freed, Emily F; Winkler, James D; Gill, Ryan T

    2015-11-20

    The ability to specifically modify bacterial genomes in a precise and efficient manner is highly desired in various fields, ranging from molecular genetics to metabolic engineering and synthetic biology. Much has changed from the initial realization that phage-derived genes may be employed for such tasks to today, where recombineering enables complex genetic edits within a genome or a population. Here, we review the major developments leading to recombineering becoming the method of choice for in situ bacterial genome editing while highlighting the various applications of recombineering in pushing the boundaries of synthetic biology. We also present the current understanding of the mechanism of recombineering. Finally, we discuss in detail issues surrounding recombineering efficiency and future directions for recombineering-based genome editing.

  19. Heat-shock protein 70 from plant biofactories of recombinant antigens activate multiepitope-targeted immune responses.

    Science.gov (United States)

    Buriani, Giampaolo; Mancini, Camillo; Benvenuto, Eugenio; Baschieri, Selene

    2012-04-01

    Although a physiological role of heat-shock proteins (HSP) in antigen presentation and immune response activation has not been directly demonstrated, their use as vaccine components is under clinical trial. We have previously demonstrated that the structure of plant-derived HSP70 (pHSP70) can be superimposed to the mammalian homologue and similarly to the mammalian counterpart, pHSP70-polypeptide complexes can activate the immune system. It is here shown that pHSP70 purified from plant tissues transiently expressing the influenza virus nucleoprotein are able to induce both the activation of major histocompatibility complex class I-restricted polyclonal T-cell responses and antibody production in mice of different haplotypes without the need of adjuvant co-delivery. These results indicate that pHSP70 derived from plants producing recombinant antigens may be used to formulate multiepitope vaccines.

  20. Human Recombinant PLD2 Can Repress p65 Activity of Guinea Pigs of Chronic Asthma in vivo

    Institute of Scientific and Technical Information of China (English)

    Ling Zhu; Weibin Zou; Chuanxing Yu; Junjin Lin; Xiaoli He

    2006-01-01

    This article is to investigate the effect of human recombinant phospholipase D2 (rhPLD2) in vivo on the expression of nuclear transcription factor p65 in chronic asthma of guinea pigs. After treating the guinea pigs with chronic asthma by rhPLD2, the crude nuclear extraction was assayed with TransAM Transcription Factor Assay Kit for the activity of pulmo tissue nuclear transcription factor p65. Compared with the healthy guinea pigs, the activity of nuclear transcription factor p65 in guinea pigs of chronic asthma is much higher than that of control groups. Our results showed that rhPLD2 markedly depressed the activity of p65 when the guinea pigs were attacked by chronic asthma. Cellular & Molecular Immunology. 2006;3(4):307-310.

  1. Generation of recombinant single-chain antibodies neutralizing the cytolytic activity of vaginolysin, the main virulence factor of Gardnerella vaginalis

    Directory of Open Access Journals (Sweden)

    Pleckaityte Milda

    2011-11-01

    Full Text Available Abstract Background Gardnerella vaginalis is identified as the predominant colonist of the vaginal tract in women with bacterial vaginosis. Vaginolysin (VLY is a protein toxin released by G. vaginalis. VLY possesses cytolytic activity and is considered as a main virulence factor of G. vaginalis. Inhibition of VLY-mediated cell lysis by antibodies may have important physiological relevance. Results Single-chain variable fragments of immunoglobulins (scFvs were cloned from two hybridoma cell lines producing neutralizing antibodies against VLY and expressed as active proteins in E. coli. For each hybridoma, two variants of anti-VLY scFv consisting of either VL-VH or VH-VL linked with a 20 aa-long linker sequence (G4S4 were constructed. Recovery of scFvs from inclusion bodies with subsequent purification by metal-chelate chromatography resulted in VLY-binding proteins that were predominantly monomeric. The antigen-binding activity of purified scFvs was verified by an indirect ELISA. The neutralizing activity was investigated by in vitro hemolytic assay and cytolytic assay using HeLa cell line. Calculated apparent Kd values and neutralizing potency of scFvs were in agreement with those of parental full-length antibodies. VH-VL and VL-VH variants of scFvs showed similar affinity and neutralizing potency. The anti-VLY scFvs derived from hybridoma clone 9B4 exhibited high VLY-neutralizing activity both on human erythrocytes and cervical epithelial HeLa cells. Conclusions Hybridoma-derived scFvs with VLY-binding activity were expressed in E. coli. Recombinant anti-VLY scFvs inhibited VLY-mediated cell lysis. The monovalent scFvs showed reduced affinity and neutralizing potency as compared to the respective full-length antibodies. The loss of avidity could be restored by generating scFv constructs with multivalent binding properties. Generated scFvs is the first example of recombinant single-chain antibodies with VLY-neutralizing activity produced in

  2. Functional divergence among silkworm antimicrobial peptide paralogs by the activities of recombinant proteins and the induced expression profiles.

    Directory of Open Access Journals (Sweden)

    Wanying Yang

    Full Text Available Antimicrobial peptides are small-molecule proteins that are usually encoded by multiple-gene families. They play crucial roles in the innate immune response, but reports on the functional divergence of antimicrobial peptide gene families are rare. In this study, 14 paralogs of antimicrobial peptides belonging to cecropin, moricin and gloverin families were recombinantly expressed in pET expression systems. By antimicrobial activity tests, peptides representing paralogs in the same family of cecropin and moricin families, displayed remarkable differences against 10 tested bacteria. The evolutionary rates were relatively fast in the two families, which presented obvious functional divergence among paralogs of each family. Four peptides of gloverin family had similar antimicrobial spectrum and activity against tested bacteria. The gloverin family showed similar antimicrobial function and slow evolutionary rates. By induced transcriptional activity, genes encoding active antimicrobial peptides were upregulated at obviously different levels when silkworm pupae were infected by three types of microbes. Association analysis of antimicrobial activities and induced transcriptional activities indicated that the antimicrobial activities might be positively correlated with induced transcriptional activities in the cecropin and moricin families. These results suggest that representative BmcecB6, BmcecD and Bmmor as the major effector genes have broad antimicrobial spectrum, strong antimicrobial activity and high microbe-induced expression among each family and maybe play crucial roles in eliminating microbial infection.

  3. Biological aerosol detection with combined passive-active infrared measurements

    Science.gov (United States)

    Ifarraguerri, Agustin I.; Vanderbeek, Richard G.; Ben-David, Avishai

    2004-12-01

    A data collection experiment was performed in November of 2003 to measure aerosol signatures using multiple sensors, all operating in the long-wave infrared. The purpose of this data collection experiment was to determine whether combining passive hyperspectral and LIDAR measurements can substantially improve biological aerosol detection performance. Controlled releases of dry aerosols, including road dust, egg albumin and two strains of Bacillus Subtilis var. Niger (BG) spores were performed using the ECBC/ARTEMIS open-path aerosol test chamber located in the Edgewood Area of Aberdeen Proving Grounds, MD. The chamber provides a ~ 20' path without optical windows. Ground truth devices included 3 aerodynamic particle sizers, an optical particle size spectrometer, 6 nephelometers and a high-volume particle sampler. Two sensors were used to make measurements during the test: the AIRIS long-wave infrared imaging spectrometer and the FAL CO2 LIDAR. The AIRIS and FAL data sets were analyzed for detection performance relative to the ground truth. In this paper we present experimental results from the individual sensors as well as results from passive-active sensor fusion. The sensor performance is presented in the form of receiver operating characteristic curves.

  4. Recombination activity of light-activated copper defects in p-type silicon studied by injection- and temperature-dependent lifetime spectroscopy

    Science.gov (United States)

    Inglese, Alessandro; Lindroos, Jeanette; Vahlman, Henri; Savin, Hele

    2016-09-01

    The presence of copper contamination is known to cause strong light-induced degradation (Cu-LID) in silicon. In this paper, we parametrize the recombination activity of light-activated copper defects in terms of Shockley—Read—Hall recombination statistics through injection- and temperature dependent lifetime spectroscopy (TDLS) performed on deliberately contaminated float zone silicon wafers. We obtain an accurate fit of the experimental data via two non-interacting energy levels, i.e., a deep recombination center featuring an energy level at Ec-Et=0.48 -0.62 eV with a moderate donor-like capture asymmetry ( k =1.7 -2.6 ) and an additional shallow energy state located at Ec-Et=0.1 -0.2 eV , which mostly affects the carrier lifetime only at high-injection conditions. Besides confirming these defect parameters, TDLS measurements also indicate a power-law temperature dependence of the capture cross sections associated with the deep energy state. Eventually, we compare these results with the available literature data, and we find that the formation of copper precipitates is the probable root cause behind Cu-LID.

  5. Intramolecular Hydrogen Bond in Biologically Active o-Carbonyl Hydroquinones

    Directory of Open Access Journals (Sweden)

    Maximiliano Martínez-Cifuentes

    2014-07-01

    Full Text Available Intramolecular hydrogen bonds (IHBs play a central role in the molecular structure, chemical reactivity and interactions of biologically active molecules. Here, we study the IHBs of seven related o-carbonyl hydroquinones and one structurally-related aromatic lactone, some of which have shown anticancer and antioxidant activity. Experimental NMR data were correlated with theoretical calculations at the DFT and ab initio levels. Natural bond orbital (NBO and molecular electrostatic potential (MEP calculations were used to study the electronic characteristics of these IHB. As expected, our results show that NBO calculations are better than MEP to describe the strength of the IHBs. NBO energies (∆Eij(2 show that the main contributions to energy stabilization correspond to LPàσ* interactions for IHBs, O1…O2-H2 and the delocalization LPàπ* for O2-C2 = Cα(β. For the O1…O2-H2 interaction, the values of ∆Eij(2 can be attributed to the difference in the overlap ability between orbitals i and j (Fij, instead of the energy difference between them. The large energy for the LP O2àπ* C2 = Cα(β interaction in the compounds 9-Hydroxy-5-oxo-4,8, 8-trimethyl-l,9(8H-anthracenecarbolactone (VIII and 9,10-dihydroxy-4,4-dimethylanthracen-1(4H-one (VII (55.49 and 60.70 kcal/mol, respectively when compared with the remaining molecules (all less than 50 kcal/mol, suggests that the IHBs in VIII and VII are strongly resonance assisted.

  6. Polyphenols from Bee Pollen: Structure, Absorption, Metabolism and Biological Activity

    Directory of Open Access Journals (Sweden)

    Anna Rzepecka-Stojko

    2015-12-01

    Full Text Available Bee pollen constitutes a natural source of antioxidants such as phenolic acids and flavonoids, which are responsible for its biological activity. Research has indicated the correlation between dietary polyphenols and cardioprotective, hepatoprotective, anti-inflammatory, antibacterial, anticancerogenic, immunostimulating, antianaemic effects, as well as their beneficial influence on osseous tissue. The beneficial effects of bee pollen on health result from the presence of phenolic acids and flavonoids which possess anti-inflammatory properties, phytosterol and linolenic acid which play an anticancerogenic role, and polysaccharides which stimulate immunological activity. Polyphenols are absorbed in the alimentary tract, metabolised by CYP450 enzymes, and excreted with urine and faeces. Flavonoids and phenolic acids are characterised by high antioxidative potential, which is closely related to their chemical structure. The high antioxidant potential of phenolic acids is due to the presence and location of hydroxyl groups, a carboxyl group in the immediate vicinity of ortho-diphenolic substituents, and the ethylene group between the phenyl ring and the carboxyl group. As regards flavonoids, essential structural elements are hydroxyl groups at the C5 and C7 positions in the A ring, and at the C3′ and C4′ positions in the B ring, and a hydroxyl group at the C3 position in the C ring. Furthermore, both, the double bond between C2 and C3, and a ketone group at the C4 position in the C ring enhance the antioxidative potential of these compounds. Polyphenols have an ideal chemical structure for scavenging free radicals and for creating chelates with metal ions, which makes them effective antioxidants in vivo.

  7. Polyphenols from Bee Pollen: Structure, Absorption, Metabolism and Biological Activity.

    Science.gov (United States)

    Rzepecka-Stojko, Anna; Stojko, Jerzy; Kurek-Górecka, Anna; Górecki, Michał; Kabała-Dzik, Agata; Kubina, Robert; Moździerz, Aleksandra; Buszman, Ewa

    2015-12-04

    Bee pollen constitutes a natural source of antioxidants such as phenolic acids and flavonoids, which are responsible for its biological activity. Research has indicated the correlation between dietary polyphenols and cardioprotective, hepatoprotective, anti-inflammatory, antibacterial, anticancerogenic, immunostimulating, antianaemic effects, as well as their beneficial influence on osseous tissue. The beneficial effects of bee pollen on health result from the presence of phenolic acids and flavonoids which possess anti-inflammatory properties, phytosterol and linolenic acid which play an anticancerogenic role, and polysaccharides which stimulate immunological activity. Polyphenols are absorbed in the alimentary tract, metabolised by CYP450 enzymes, and excreted with urine and faeces. Flavonoids and phenolic acids are characterised by high antioxidative potential, which is closely related to their chemical structure. The high antioxidant potential of phenolic acids is due to the presence and location of hydroxyl groups, a carboxyl group in the immediate vicinity of ortho-diphenolic substituents, and the ethylene group between the phenyl ring and the carboxyl group. As regards flavonoids, essential structural elements are hydroxyl groups at the C5 and C7 positions in the A ring, and at the C3' and C4' positions in the B ring, and a hydroxyl group at the C3 position in the C ring. Furthermore, both, the double bond between C2 and C3, and a ketone group at the C4 position in the C ring enhance the antioxidative potential of these compounds. Polyphenols have an ideal chemical structure for scavenging free radicals and for creating chelates with metal ions, which makes them effective antioxidants in vivo.

  8. SYNTHESIS, CHARACTERIZATION AND BIOLOGICAL ACTIVITY OF SOME NOVEL ARYL AND HETROARYL CHALCONE ANALOGUES

    Directory of Open Access Journals (Sweden)

    Tribhuvan Singh

    2012-07-01

    Full Text Available A new series of Heterocyclic chalcones showed diversified biological activities. In view of potential biological activities of Heterocyclic chalcones derivative were prepared by claisen-Schmidt condensation technique. The compound were screened for anti-inflammatory and antibacterial activity.

  9. Neutralization of biological activity and inhibition of receptor binding by antibodies against human thrombopoietin.

    Science.gov (United States)

    Tahara, T; Kuwaki, T; Matsumoto, A; Morita, H; Watarai, H; Inagaki, Y; Ohashi, H; Ogami, K; Miyazaki, H; Kato, T

    1998-01-01

    Thrombopoietin (TPO) is a recently isolated cytokine that primarily regulates megakaryocytopoiesis and thrombopoiesis. We recently reported the development of a variety of antibodies (Abs) to synthetic peptides of human (h)TPO and to recombinant human TPO (rhTPO). In this study, we characterized the Abs and mapped immunologically distinct areas of the molecule. Among the five different antipeptide polyclonal Abs, only one, raised against synthetic peptide D8 to Q28, neutralized the TPO-dependent growth of FDCP-2 cells expressing human Mpl (FDCP-hMpl5 cells). One out of seven anti-rhTPO monoclonal Abs, designated as TN1, also showed neutralizing activity. TN1 was found to be specifically reactive with two proteolytic fragments, residues S1 to R117 and A60 to K122 of hTPO, indicating that the epitope(s) of TN1 is localized in residues A60 to R117 of the molecule. These two neutralizing Abs inhibited the binding of biotinylated rhTPO to FDCP-hMpl5 cells. On the other hand, the other Abs, which reacted with five polypeptides of S47 to D62, L108 to A126, N172 to A190, S262 to T284, and P306 to G332 of hTPO, did not show either the neutralizing activity or the ability to inhibit the binding of biotinylated rhTPO to the cell surface hMpl. These findings indicate that two regions, residues D8 to Q28 and A60 to R117 of hTPO, may contain the domains associated with its receptor, C-Mpl. These Abs characterized here are valuable for studying the structural analysis and the biological function of hTPO mediated by its receptor.

  10. Assessment of the adjuvant activity of mesoporous silica nanoparticles in recombinant Mycoplasma hyopneumoniae antigen vaccines

    Directory of Open Access Journals (Sweden)

    Veridiana Gomes Virginio

    2017-01-01

    Full Text Available The adjuvant potential of two mesoporous silica nanoparticles (MSNs, SBa-15 and SBa-16, was assessed in combination with a recombinant HSP70 surface polypeptide domain from Mycoplasma hyopneumoniae, the etiological agent of porcine enzootic pneumonia (PEP. The recombinant antigen (HSP70212-600, previously shown as immunogenic in formulation with classic adjuvants, was used to immunize BALB/c mice in combination with SBa-15 or SBa-16 MSNs, and the effects obtained with these formulations were compared to those obtained with alum, the adjuvant traditionally used in anti-PEP bacterins. The HSP70212-600 + SBa-15 vaccine elicited a strong humoral immune response, with high serum total IgG levels, comparable to those obtained using HSP70212-600 + alum. The HSP70212-600 + SBa-16 vaccine elicited a moderate humoral immune response, with lower levels of total IgG. The cellular immune response was assessed by the detection of IFN-γ, IL-4 and IL-10 in splenocyte culture supernatants. The HSP70212-600 + SBa-15 vaccine increased IFN-γ, IL-4 and IL-10 levels, while no stimulation was detected with the HSP70212-600 + SBa-16 vaccine. The HSP70212-600 + SBa-15 vaccine induced a mixed Th1/Th2-type response, with an additional IL-10 mediated anti-inflammatory effect, both of relevance for an anti-PEP vaccine. Alum adjuvant controls stimulated an unspecific cellular immune response, with similar levels of cytokines detected in mice immunized either with HSP70212-600 + alum or with the adjuvant alone. The better humoral and cellular immune responses elicited in mice indicated that SBa-15 has adjuvant potential, and can be considered as an alternative to the use of alum in veterinary vaccines. The use of SBa-15 with HSP70212-600 is also promising as a potential anti-PEP subunit vaccine formulation.

  11. Investigating the Use of Inquiry & Web-Based Activities with Inclusive Biology Learners

    Science.gov (United States)

    Bodzin, Alec M.; Waller, Patricia L.; Edwards, Lana; Darlene Kale, Santoro

    2007-01-01

    A Web-integrated biology program is used to explore how to best assist inclusive high school students to learn biology with inquiry-based activities. Classroom adaptations and instructional strategies teachers may use to assist in promoting biology learning with inclusive learners are discussed.

  12. Homing of radiolabelled recombinant interleukin-2 activated natural killer cells and their efficacy in adoptive immunotherapy against murine fibrosarcoma

    Indian Academy of Sciences (India)

    Anuradha Rai; Ashim K Chakravarty

    2007-12-01

    Natural killer (NK) cells are spontaneously cytotoxic against tumour target cells. Their number was found to be four times more in the spleen of tumour-bearing Swiss albino mice. After activation with recombinant interleukin-2 (rIL-2), NK cells were tested and found to seek out the tumour site when injected intravenously in tumour-bearing mice. Their potential for fighting tumours in vivo was further seen following adoptive transfer of rIL-2 activated NK (A-NK) cells in tumour-bearing mice. After surgical removal of tumour load, adoptive transfer of A-NK cells inhibited tumour recurrence in 92.3% cases, thereby suggesting the use of this protocol for therapeutic purposes to obtain a better outcome.

  13. Structure-activity relationship of a recombinant hybrid Manganese superoxide dismutase of Staphylococcus saprophyticus/S. equorum.

    Science.gov (United States)

    Retnoningrum, Debbie S; Arumsari, Sekar; Artarini, Anita; Ismaya, Wangsa T

    2017-05-01

    Recombinant hybrid Manganese superoxide dismutase from Staphyloccus saphropyticus/S. equorum (rMnSODSeq) exhibits stability at high temperatures. The enzyme occurs as a dimer that dissociates around 52°C prior to unfolding of the monomer around 64°C, demonstrating contribution of the dimeric form to stability. Here, structure - activity relationship of rMnSODSeq was evaluated on the basis of its activity and stability in the presence of inhibitors, NaCl, denaturants, detergents, reducing agents, and at different pH values. The activity was evaluated at both 37°C and 52°C, which the latter is the temperature for dissociation of the dimer. Dimer to monomer transition coincided with significant decrease in residual activity at 52°C. However, the activity assay results at 52°C and 37°C suggest spontaneous re-association of the monomer into dimer. Intriguingly, various new species with melting temperature (TM) values other than those of the dimer or monomer were observed. These species displayed medium to comparable level of residual activities to the native at 37°C. This report suggests that dimer to monomer transition may be not the only explanation for activity loss or decrease.

  14. Electrodermal screening of biologically active points for upper gastrointestinal bleeding.

    Science.gov (United States)

    Tseng, Ying-Jung; Hu, Wen-Long; Hung, I-Ling; Hsieh, Chia-Jung; Hung, Yu-Chiang

    2014-01-01

    The purpose of this case-control study was to investigate the relationship between the electrical resistance of the skin at biologically active points (BAPs) on the main meridians and upper gastrointestinal bleeding (UGIB). Electrical resistance to direct current at 20 BAPs on the fingers and toes of 100 patients with (38 men, 12 women; mean age [range], 58.20 ± 19.62 [18-83] years) and without (27 men, 23 women; 49.54 ± 12.12 [22-74] years) UGIB was measured through electrodermal screening (EDS), based on the theory of electroacupuncture according to Voll (EAV). Data were compared through analysis of variance (ANOVA), receiver operating characteristic (ROC) curve analysis, and logistic regression. The initial readings were lower in the UGIB group, indicating blood and energy deficiency due to UGIB. Significant differences in indicator drop values were observed at nine BAPs (p < 0.05) on the bilateral small intestine, bilateral stomach, bilateral circulation, bilateral fibroid degeneration, and right lymph meridians. The area under the ROC curve values of the BAPs on the bilateral small intestine and stomach meridians were larger than 0.5, suggesting the diagnostic accuracy of EDS for UGIB on the basis of the indicator drop of these BAPs. Logistic regression revealed that when the indicator drop of the BAP on the left stomach meridian increased by one score, the risk of UGIB increased by about 1.545-3.523 times. In conclusion, the change in the electrical resistance of the skin measured by EDS at the BAPs on the bilateral small intestine and stomach meridians provides specific information on UGIB.

  15. Dissolution of emboli in rats with experimental cerebral thromboembolism by recombinant human tissue plasminogen activator (TD-2061)

    Energy Technology Data Exchange (ETDEWEB)

    Hara, T.; Iwamoto, M.; Ogawa, H.; Yamamoto, A.; Tomikawa, M. (Research Institute, Daiichi Pharmaceutical Co., Ltd., Tokyo, (Japan))

    1990-08-15

    Tissue plasminogen activator (t-PA) is frequently administered clinically as thrombolytic therapy. We injected recombinant t-PA into rats with cerebral {sup 125}I-labeled blood clot emboli to evaluate the dissolutive effect of recombinant human single-chain t-PA (rt-PA; TD-2061) on such emboli and to examine the possibility of improving neurological damage in patients with cerebral thrombosis. When rt-PA was given intravenously at a dose of 350,000 IU/kg 2 minutes before embolization, radioactivity in the affected cerebral hemisphere decreased to 20% of that in the vehicle control 2 hours after embolization. A significant decrease in radioactivity in the cerebral hemisphere was also found on the administration of 700,000 IU/kg of rt-PA 30 or 60 minutes after embolization, but not when rt-PA was administered 2 minutes after embolization. Marked inhibition of abnormal behavior such as hemiplegia was seen on treatment with rt-PA 2 minutes before embolization, but not at all when rt-PA treatment was given 30 or 60 minutes after embolization. The findings suggest that rt-PA can dissolve blood clot emboli in cerebral vessels and that prompt thrombolytic therapy is important to minimize neurological dysfunction in cases of cerebral thromboembolism.

  16. Phospholipase C Produced by Clostridium botulinum Types C and D:Comparison of Gene, Enzymatic, and Biological Activities with Those of Clostridium perfringens Alpha-toxin

    Directory of Open Access Journals (Sweden)

    Sakurai,Jun

    2013-02-01

    Full Text Available Clostridium botulinum type C and D strains recently have been found to produce PLC on egg yolk agar plates. To characterize the gene, enzymatic and biological activities of C. botulinum PLCs (Cb-PLCs, the cb-plc genes from 8 strains were sequenced, and 1 representative gene was cloned and expressed as a recombinant protein. The enzymatic and hemolytic activities of the recombinant Cb-PLC were measured and compared with those of the Clostridium perfringens alpha-toxin. Each of the eight cb-plc genes encoded a 399 amino acid residue protein preceded by a 27 residue signal peptide. The protein consists of 2 domains, the N- and C-domains, and the overall amino acid sequence identity between Cb-PLC and alpha-toxin was greater than 50%, suggesting that Cb-PLC is homologous to the alpha-toxin. The key residues in the N-domain were conserved, whereas those in the C-domain which are important in membrane interaction were different than in the alpha-toxin. As expected, Cb-PLC could hydrolyze egg yolk phospholipid, p-nitrophenylphosphorylcholine, and sphingomyelin, and also exhibited hemolytic activity;however, its activities were about 4- to over 200-fold lower than those of alpha-toxin. Although Cb-PLC showed weak enzymatic and biological activities, it is speculated that Cb-PLC might play a role in the pathogenicity of botulism or for bacterial survival.

  17. Phospholipase C produced by Clostridium botulinum types C and D: comparison of gene, enzymatic, and biological activities with those of Clostridium perfringens alpha-toxin.

    Science.gov (United States)

    Fatmawati, Ni Nengah Dwi; Sakaguchi, Yoshihiko; Suzuki, Tomonori; Oda, Masataka; Shimizu, Kenta; Yamamoto, Yumiko; Sakurai, Jun; Matsushita, Osamu; Oguma, Keiji

    2013-01-01

    Clostridium botulinum type C and D strains recently have been found to produce PLC on egg yolk agar plates. To characterize the gene, enzymatic and biological activities of C. botulinum PLCs (Cb-PLCs), the cb-plc genes from 8 strains were sequenced, and 1 representative gene was cloned and expressed as a recombinant protein. The enzymatic and hemolytic activities of the recombinant Cb-PLC were measured and compared with those of the Clostridium perfringens alpha-toxin. Each of the eight cb-plc genes encoded a 399 amino acid residue protein preceded by a 27 residue signal peptide. The protein consists of 2 domains, the N- and C-domains, and the overall amino acid sequence identity between Cb-PLC and alpha-toxin was greater than 50%, suggesting that Cb-PLC is homologous to the alpha-toxin. The key residues in the N-domain were conserved, whereas those in the C-domain which are important in membrane interaction were different than in the alpha-toxin. As expected, Cb-PLC could hydrolyze egg yolk phospholipid, p-nitrophenylphosphorylcholine, and sphingomyelin, and also exhibited hemolytic activity;however, its activities were about 4- to over 200-fold lower than those of alpha-toxin. Although Cb-PLC showed weak enzymatic and biological activities, it is speculated that Cb-PLC might play a role in the pathogenicity of botulism or for bacterial survival.

  18. Expression, purification and characterization of a functional, recombinant, cold-active lipase (LipA) from psychrotrophic Yersinia enterocolitica.

    Science.gov (United States)

    Ji, Xiuling; Li, Shan; Wang, Baoqiang; Zhang, Qi; Lin, Lianbing; Dong, Zhiyang; Wei, Yunlin

    2015-11-01

    A novel cold-active lipase gene encoding 294 amino acid residues was obtained from the Yersinia enterocolitica strain KM1. Sequence alignment and phylogenetic analysis revealed that this novel lipase is a new member of the bacterial lipase family I.1. The lipase shares the conserved GXSXG motif and catalytic triad Ser85-Asp239-His261. The recombinant protein LipA was solubly and heterogeneously expressed in Escherichia coli, purified by Ni-affinity chromatography, and then characterized. LipA was active over a broad range spanning 15-60°C with an optimum activity at 25°C and across a wide pH range from 5.0 to 11.0 with an optimum activity at pH 7.5. The molecular weight was estimated to be 34.2 KDa. The lipase could be activated by Mg(2+) and a low concentration (10%) of ethanol, dimethyl sulfoxide, methanol and acetonitrile, whereas it was strongly inhibited by Zn(2+), Cu(2+) and Mn(2+). This cold-active lipase may be a good candidate for detergents and biocatalysts at low temperature.

  19. Recombinant Nox4 cytosolic domain produced by a cell or cell-free base systems exhibits constitutive diaphorase activity

    Energy Technology Data Exchange (ETDEWEB)

    Nguyen, Minh Vu Chuong, E-mail: mvchuong@yahoo.fr [GREPI AGIM FRE 3405 CNRS-UJF, Universite Joseph Fourier, Grenoble (France); Zhang, Leilei [GREPI AGIM FRE 3405 CNRS-UJF, Universite Joseph Fourier, Grenoble (France); Lhomme, Stanislas; Mouz, Nicolas [PX' Therapeutics, MINATEC/Batiment de Haute Technologie, Grenoble (France); Lenormand, Jean-Luc [HumProTher Laboratory, TheReX/TIMC-IMAG UMR 5525 CNRS UJF, Universite Joseph Fourier, UFR de Medecine, Domaine de la Merci, 38706 La Tronche (France); Lardy, Bernard; Morel, Francoise [GREPI AGIM FRE 3405 CNRS-UJF, Universite Joseph Fourier, Grenoble (France)

    2012-03-16

    Highlights: Black-Right-Pointing-Pointer A comparison of two bacterial cell and cell-free protein expression systems is presented. Black-Right-Pointing-Pointer Soluble and active truncated Nox4 proteins are produced. Black-Right-Pointing-Pointer Nox4 has a constitutive diaphorase activity which is independent of cytosolic factors. Black-Right-Pointing-Pointer Isoform Nox4B is unable to initiate the first electronic transfer step. Black-Right-Pointing-Pointer Findings contribute to the understanding of the mechanism of Nox4 oxidase activity. -- Abstract: The membrane protein NADPH (nicotinamide adenine dinucleotide phosphate) oxidase Nox4 constitutively generates reactive oxygen species differing from other NADPH oxidases activity, particularly in Nox2 which needs a stimulus to be active. Although the precise mechanism of production of reactive oxygen species by Nox2 is well characterized, the electronic transfer throughout Nox4 remains unclear. Our study aims to investigate the initial electronic transfer step (diaphorase activity) of the cytosolic tail of Nox4. For this purpose, we developed two different approaches to produce soluble and active truncated Nox4 proteins. We synthesized soluble recombinant proteins either by in vitro translation or by bacteria induction. While proteins obtained by bacteria induction demonstrate an activity of 4.4 {+-} 1.7 nmol/min/nmol when measured against iodonitro tetrazolium chloride and 20.5 {+-} 2.8 nmol/min/nmol with cytochrome c, the soluble proteins produced by cell-free expression system exhibit a diaphorase activity with a turn-over of 26 {+-} 2.6 nmol/min/nmol when measured against iodonitro tetrazolium chloride and 48 {+-} 20.2 nmol/min/nmol with cytochrome c. Furthermore, the activity of the soluble proteins is constitutive and does not need any stimulus. We also show that the cytosolic tail of the isoform Nox4B lacking the first NADPH binding site is unable to demonstrate any diaphorase activity pointing out the

  20. Biologic

    CERN Document Server

    Kauffman, L H

    2002-01-01

    In this paper we explore the boundary between biology and the study of formal systems (logic). In the end, we arrive at a summary formalism, a chapter in "boundary mathematics" where there are not only containers but also extainers ><, entities open to interaction and distinguishing the space that they are not. The boundary algebra of containers and extainers is to biologic what boolean algebra is to classical logic. We show how this formalism encompasses significant parts of the logic of DNA replication, the Dirac formalism for quantum mechanics, formalisms for protein folding and the basic structure of the Temperley Lieb algebra at the foundations of topological invariants of knots and links.

  1. Evaluation of the biological activity of sunflower hull extracts

    Directory of Open Access Journals (Sweden)

    Taha, F. S.

    2012-06-01

    Full Text Available This work was planned with the aim of adding value to sunflower seed hulls, a waste product of the oil industry by preparing a sunflower hull phenolic extract rich in chlorogenic acid (CGA. In order to fulfill this goal, the optimization for the extraction of a phenolic extract from the hulls was investigated. The parameters studied were: type of solvent, solvent to water ratio and hull to solvent ratio. In addition, the solvent mixtures were also studied. The resulting phenolic extracts were evaluated for their biological activities. This included phenolic content determination, evaluation of the antioxidant and antimicrobial activities. Chlorogenic acid was determined in two chosen hull extracts using the UV spectrophotometric method and HPLC analysis. The anticarcinogenic activity of the two chosen extracts was tested on seven different cell line carcinomas. The results revealed that all the phenolic extracts of sunflower hull studied contain between 190-312.5 mg phenolics/ 100 g hulls. The highest phenolic extraction was achieved with 80% methanol (1:30, hull to solvent, w/v ratio and methanol to ethanol to water (7:7:6 v/v/v mixture with values of 312.5 and 306.5 mg phenolics/100 g hulls, respectively. The free radical scavenging activity and antioxidant activity of all the samples ranged from 33.6-72.6%. The highest antioxidant activity and free radical scavenging activity were achieved by the same extracts that possessed the highest phenolic content, namely methanol to ethanol to water extract and 80% methanol with values 71.8 and 72.6%, 68.2 and 70.9% respectively, compared to 77.9 and 76.9% respectively for TBHQ. All the phenolic extracts possessed antimicrobial activity but to different levels against different pathogenic bacteria. The two chosen extracts also possessed anticarcinogenic activity, which differed among varying cell line carcinomas. The HPLC analysis indicated that chlorogenic acid was the main phenolic acid in the

  2. Recombinant mouse PAP has pH-dependent ectonucleotidase activity and acts through A(1-adenosine receptors to mediate antinociception.

    Directory of Open Access Journals (Sweden)

    Nathaniel A Sowa

    Full Text Available Prostatic acid phosphatase (PAP is expressed in nociceptive neurons and functions as an ectonucleotidase. When injected intraspinally, the secretory isoforms of human and bovine PAP protein have potent and long-lasting antinociceptive effects that are dependent on A(1-adenosine receptor (A(1R activation. In this study, we purified the secretory isoform of mouse (mPAP using the baculovirus expression system to determine if recombinant mPAP also had antinociceptive properties. We found that mPAP dephosphorylated AMP, and to a much lesser extent, ADP at neutral pH (pH 7.0. In contrast, mPAP dephosphorylated all purine nucleotides (AMP, ADP, ATP at an acidic pH (pH 5.6. The transmembrane isoform of mPAP had similar pH-dependent ectonucleotidase activity. A single intraspinal injection of mPAP protein had long-lasting (three day antinociceptive properties, including antihyperalgesic and antiallodynic effects in the Complete Freund's Adjuvant (CFA inflammatory pain model. These antinociceptive effects were transiently blocked by the A(1R antagonist 8-cyclopentyl-1, 3-dipropylxanthine (CPX, suggesting mPAP dephosphorylates nucleotides to adenosine to mediate antinociception just like human and bovine PAP. Our studies indicate that PAP has species-conserved antinociceptive effects and has pH-dependent ectonucleotidase activity. The ability to metabolize nucleotides in a pH-dependent manner could be relevant to conditions like inflammation where tissue acidosis and nucleotide release occur. Lastly, our studies demonstrate that recombinant PAP protein can be used to treat chronic pain in animal models.

  3. Systems Biology Graphical Notation: Activity Flow language Level 1 Version 1.2.

    Science.gov (United States)

    Mi, Huaiyu; Schreiber, Falk; Moodie, Stuart; Czauderna, Tobias; Demir, Emek; Haw, Robin; Luna, Augustin; Le Novère, Nicolas; Sorokin, Anatoly; Villéger, Alice

    2015-09-04

    The Systems Biological Graphical Notation (SBGN) is an international community effort for standardized graphical representations of biological pathways and networks. The goal of SBGN is to provide unambiguous pathway and network maps for readers with different scientific backgrounds as well as to support efficient and accurate exchange of biological knowledge between different research communities, industry, and other players in systems biology. Three SBGN languages, Process Description (PD), Entity Relationship (ER) and Activity Flow (AF), allow for the representation of different aspects of biological and biochemical systems at different levels of detail. The SBGN Activity Flow language represents the influences of activities among various entities within a network. Unlike SBGN PD and ER that focus on the entities and their relationships with others, SBGN AF puts the emphasis on the functions (or activities) performed by the entities, and their effects to the functions of the same or other entities. The nodes (elements) describe the biological activities of the entities, such as protein kinase activity, binding activity or receptor activity, which can be easily mapped to Gene Ontology molecular function terms. The edges (connections) provide descriptions of relationships (or influences) between the activities, e.g., positive influence and negative influence. Among all three languages of SBGN, AF is the closest to signaling pathways in biological literature and textbooks, but its well-defined semantics offer a superior precision in expressing biological knowledge.

  4. Viral and murine interleukin-10 are correctly processed and retain their biological activity when produced in tobacco

    Directory of Open Access Journals (Sweden)

    Avesani Linda

    2009-03-01

    Full Text Available Abstract Background Interleukin-10 (IL-10 is a potent anti-inflammatory cytokine, with therapeutic applications in several autoimmune and inflammatory diseases. Oral administration of this cytokine alone, or in combination with disease-associated autoantigens could confer protection form the onset of a specific autoimmune disease through the induction of oral tolerance. Transgenic plants are attractive systems for production of therapeutic proteins because of the ability to do large scale-up at low cost, and the low maintenance requirements. They are highly amenable to oral administration and could become effective delivery systems without extensive protein purification. We investigated the ability of tobacco plants to produce high levels of biologically-active viral and murine IL-10. Results Three different subcellular targeting strategies were assessed in transient expression experiments, and stable transgenic tobacco plants were generated with the constructs that yielded the highest accumulation levels by targeting the recombinant proteins to the endoplasmic reticulum. The best yields using this strategy in T1 plants were 10.8 and 37.0 μg/g fresh leaf weight for viral and murine IL-10, respectively. The recombinant proteins were purified from transgenic leaf material and characterized in terms of their N-glycan composition, dimerization and biological activity in in vitro assays. Both molecules formed stable dimers, were able to activate the IL-10 signaling pathway and to induce specific anti-inflammatory responses in mouse J774 macrophage cells. Conclusion Tobacco plants are able to correctly process viral and murine IL-10 into biologically active dimers, therefore representing a suitable platform for the production for these cytokines. The accumulation levels obtained are high enough to allow delivery of an immunologically relevant dose of IL-10 in a reasonable amount of leaf material, without extensive purification. This study paves the

  5. Application of synchrotron-radiation-based x-ray microprobe techniques for the analysis of recombination activity of metals precipitated at Si/SiGe misfit dislocations

    Energy Technology Data Exchange (ETDEWEB)

    Vyvenko, O F [University of California, LBNL, 1 Cyclotron Rd, Berkeley, CA 94720 (United States); Buonassisi, T [University of California, LBNL, 1 Cyclotron Rd, Berkeley, CA 94720 (United States); Istratov, A A [University of California, LBNL, 1 Cyclotron Rd, Berkeley, CA 94720 (United States); Weber, E R [University of California, LBNL, 1 Cyclotron Rd, Berkeley, CA 94720 (United States); Kittler, M [IHP, Im Technologiepark 25, D-15236 Frankfurt (Oder) (Germany); Seifert, W [IHP, Im Technologiepark 25, D-15236 Frankfurt (Oder) (Germany)

    2002-12-09

    In this study we report application of synchrotron-radiation-based x-ray microprobe techniques (the x-ray-beam-induced current (XBIC) and x-ray fluorescence ({mu}-XRF) methods) to the analysis of the recombination activity and space distribution of copper and iron in the vicinity of dislocations in silicon/silicon-germanium structures. A combination of these two techniques enables one to study the chemical nature of the defects and impurities and their recombination activity in situ and to map metal clusters with a micron-scale resolution. XRF analysis revealed that copper formed clearly distinguishable precipitates along the misfit dislocations. A proportional dependence between the XBIC contrast and the number of copper atoms in the precipitates was established. In hydrogen-passivated iron-contaminated samples we observed clusters of iron precipitates which had no recombination activity detectable by the XBIC technique as well as iron clusters which were not completely passivated.

  6. 重组EpoAB-NGF模拟肽的神经营养功能%Preliminary Study on Neurotrophic Activity of Recombinant Peptide Mimics EpoAB-NGF

    Institute of Scientific and Technical Information of China (English)

    潘勇昭; 陈虹; 王欣; 马晓骊; 黄秉仁

    2011-01-01

    Objective: To investigate the neurotrophic activity of recombinant fusion protein epopeptide ABnerve growth factor analog peptide (EpoAB-NGF9/12). Methods: The DNA fragments encoding the sequences of EpoAB-NGF9 and EpoAB-NGF12 polypeptide were amplified by PCR and cloned into the pET-42a prokaryotic expression vector. The recombinant plasmids of pET-42a-EpoAB-NGF9/12 were transformed into E. coli BL21 (DE3) and exogenous protein expression was induced by IPTG. Fusion proteins GST-EpoAB-NGF9/12 were purified by affinity chromatography. The biological activities were determined using the experiment of inducing axon outgrowth of PC12 cells and flow cytometry analysis of apoptotic rate in R2L1 cells. Results: Apparent molecular weight of fusion proteins GST-EpoAB-NGF9/12 were approximate 30kDa. Immunoblot analysis showed that the fusion proteins were immunoreactive with anti-GST antibody. Fusion proteins stimulate the differentiation and promote the axon outgrowth in PC12 cells. Cell apoptosis was induced in (31.7 ±0.60)% of R2L1 control cells by serum free incubation, whereas cell apoptosis rate were in ( 25.2 ± 3.52 ) % or ( 25.7 ± 1.46 ) % by adding GST-EpoAB-NGF9 or GST-EpoAB-NGF12 into serum free condition respectively. The result indicates that fusion proteins were enabling to prevent cell death in R2L1 cells. Conclusion: These findings in cell biology region suggest that recombinant fusion protein containing Epo-NGF peptide mimics have the neurotrophic effects similar to that of NGF.%目的:研究重组融合蛋白红细胞生成素AB肽-神经生长因子模拟肽(EpoAB-NGF9和EpoAB-NGF/12)的神经营养作用.方法:构建pET-42a-EpoAB-NGF9/12原核表达质粒,转化大肠杆菌BL21(DE3),以IPTG诱导表达,亲和层析纯化重组蛋白,显微镜观察PC12细胞诱导分化,流式细胞仪检测凋亡细胞.结果:大肠杆菌表达的重组融合蛋白GST-EpoAB-NGF9/12分子量约为30kDa,抗GST抗体免疫印迹反应呈阳

  7. Telomerase repeat amplification protocol (TRAP) activity upon recombinant expression and purification of human telomerase in a bacterial system.

    Science.gov (United States)

    Hansen, Debra T; Thiyagarajan, Thirumagal; Larson, Amy C; Hansen, Jeffrey L

    2016-07-01

    Telomerase biogenesis is a highly regulated process that solves the DNA end-replication problem. Recombinant expression has so far been accomplished only within a eukaryotic background. Towards structural and functional analyses, we developed bacterial expression of human telomerase. Positive activity by the telomerase repeat amplification protocol (TRAP) was identified in cell extracts of Escherichia coli expressing a sequence-optimized hTERT gene, the full-length hTR RNA with a self-splicing hepatitis delta virus ribozyme, and the human heat shock complex of Hsp90, Hsp70, p60/Hop, Hsp40, and p23. The Hsp90 inhibitor geldanamycin did not affect post-assembly TRAP activity. By various purification methods, TRAP activity was also obtained upon expression of only hTERT and hTR. hTERT was confirmed by tandem mass spectrometry in a ∼120 kDa SDS-PAGE fragment from a TRAP-positive purification fraction. TRAP activity was also supported by hTR constructs lacking the box H/ACA small nucleolar RNA domain. End-point TRAP indicated expression levels within 3-fold of that from HeLa carcinoma cells, which is several orders of magnitude below detection by the direct assay. These results represent the first report of TRAP activity from a bacterium and provide a facile system for the investigation of assembly factors and anti-cancer therapeutics independently of a eukaryotic setting.

  8. Recombinant jacalin-like plant lectins are produced at high levels in Nicotiana benthamiana and retain agglutination activity and sugar specificity.

    Science.gov (United States)

    Fernandez-del-Carmen, Asun; Juárez, Paloma; Presa, Silvia; Granell, Antonio; Orzáez, Diego

    2013-02-20

    The plant kingdom is an underexplored source of valuable proteins which, like plant lectins, display unique interacting specificities. Furthermore, plant protein diversity remains under-exploited due to the low availability and heterogeneity of native sources. All these hurdles could be overcome with recombinant production. A narrow phylogenetic gap between the native source and the recombinant platform is likely to facilitate proper protein processing and stability; therefore, the plant cell chassis should be specially suited for the recombinant production of many plant native proteins. This is illustrated herein with the recombinant production of two representatives of the plant jacalin-related lectin (JRLs) protein family in Nicotiana benthamiana using state-of-the-art magnICON technology. Mannose-specific Banlec JRL was produced at very high levels in leaves, reaching 1.0mg of purified protein per gram of fresh weight and showing strong agglutination activity. Galactose-specific jacalin JRL, with its complicated processing requirements, was also successfully produced in N. benthamiana at levels of 0.25 mg of purified protein per gram of fresh weight. Recombinant Jacalin (rJacalin) proved efficient in the purification of human IgA1, and was able to discriminate between plant-made and native IgA1 due to their differential glycosylation status. Together, these results show that the plant cell factory should be considered a primary option in the recombinant production of valuable plant proteins.

  9. Bioactivity of marine organisms. Part 3. Screening of marine algae of Indian coast for biological activity

    Digital Repository Service at National Institute of Oceanography (India)

    Kamat, S.Y.; Wahidullah, S.; Naik, C.G.; DeSouza, L.; Jayasree, V.; Ambiye, V.; Bhakuni, D.S.; Goel, A.K.; Garg, H.S.; Srimal, R.C.

    Ethanolic extracts from Indian marine algae have been tested for anti-viral, anti-bacterial, anti-fungal, anti-fertility, hypoglycaemic and a wide range of pharmacological activities. Of 34 species investigated 17 appeared biologically active. Six...

  10. Biological Activity of Peanut (Arachis hypogaea) Phytoalexins and Selected Natural and Synthetic Stilbenoids

    Science.gov (United States)

    2011-02-11

    for their effects on economically important plant pathogenic fungi of the genera Colletotrichum, Botrytis, Fusarium, and Phomopsis. We further...hypogaea, peanuts, groundnuts, stilbenoids, resveratrol, phytoalexins, biological activity, antifungal, antitumor, anticancer , cytotoxic, antioxidant...prenylated flavonoids have been identified as constituents in plants, and display biological activities, such as anticancer , antiandrogen, anti-Leishmania, and

  11. Efficacy of recombinant tissue-type plasminogen activator thrombolysis and primary coronary stenting after acute myocardial infarction

    Institute of Scientific and Technical Information of China (English)

    陈步星; 王伟民; 赵红; 胡大一; 徐成斌; 赵明中; 卢明瑜; 刘健; 吴淳

    2003-01-01

    Objective To compare the efficacy of low dose recombinant tissue-type plasminogen activator (rt-PA) thrombolysis with primary coronary stenting after acute myocardial infarction.Methods Of 261 patients with first acute myocardial infarction, 131 were given low dose rt-PA intravenous thrombolysis, and 130 primary coronary stenting.Results The age, time from onset of chest pain to hospital presentation and infarct location between these two groups were comparable. The patency rate of the infarct-related artery (IRA) in patients in the thrombolysis group was significantly lower than that of patients in the primary stenting group (P0.05).Conclusion Comparing with low dose rt-PA thrombolytic therapy after acute myocardial infarction, primary coronary stenting has a higher patency rate of the IRA, better cardiac function and shorter hospitalization time.

  12. Characterization of the highly active fragment of glyceraldehyde-3-phosphate dehydrogenase gene promoter for recombinant protein expression in Pleurotus ostreatus.

    Science.gov (United States)

    Yin, Chaomin; Zheng, Liesheng; Zhu, Jihong; Chen, Liguo; Ma, Aimin

    2015-03-01

    Developing efficient native promoters is important for improving recombinant protein expression by fungal genetic engineering. The promoter region of glyceraldehyde-3-phosphate dehydrogenase gene in Pleurotus ostreatus (Pogpd) was isolated and optimized by upstream truncation. The activities of these promoters with different lengths were further confirmed by fluorescence, quantitative real-time PCR and Western blot analysis. A truncated Pogpd-P2 fragment (795 bp) drove enhanced green fluorescence protein (egfp) gene expression in P. ostreatus much more efficiently than full-length Pogpd-P1. Further truncating Pogpd-P2 to 603, 403 and 231 bp reduced the eGFP expression significantly. However, the 403-bp fragment between -356 bp and the start codon was the minimal but sufficient promoter element for eGFP expression. Compact native promoters for genetic engineering of P. ostreatus were successfully developed and validated in this study. This will broaden the preexisting repertoire of fungal promoters for biotechnology application.

  13. Guidance on patient Identification and Administration of Recombinant Human Activated protein C for the Treatment of Severe Sepsis

    Directory of Open Access Journals (Sweden)

    Gary Garber

    2002-01-01

    Full Text Available Approximately one-third of cases of severe sepsis result in death. Endogenous activated protein C (ApC plays a key role in the regulation of the inflammation, fibrinolysis and coagulation associated with severe sepsis. In a recently published phase III trial, protein C Worldwide Evaluation in Severe Sepsis (pROWESS, intravenous administration of recombinant human ApC (rhApC 24 µg/kg/h for 96 h to patients with severe sepsis resulted in a 6.1% reduction in absolute mortality and a 19.4% reduction in the relative risk of death from any cause within 28 days (number needed to treat = 16. This dose is now being applied in clinical practice.

  14. Activity in mice of recombinant BCG-EgG1Y162 vaccine for Echinococcus granulosus infection.

    Science.gov (United States)

    Ma, Xiumin; Zhao, Hui; Zhang, Fengbo; Zhu, Yuejie; Peng, Shanshan; Ma, Haimei; Cao, Chunbao; Xin, Yan; Yimiti, Delixiati; Wen, Hao; Ding, Jianbing

    2016-01-01

    Cystic hydatid disease is a zoonotic parasitic disease caused by Echinococcus granulosus which is distributed worldwide. The disease is difficult to treat with surgery removal is the only cure treatment. In the high endemic areas, vaccination of humans is believed a way to protect communities from the disease. In this study we vaccinated BALB/c mice with rBCG-EgG1Y162, and then detected the level of IgG and IgE specifically against the recombinant protein by ELISA, rBCG-EgG1Y162 induced strong and specific cellular and humoral immune responses. In vitro study showed that rBCG-EgG1Y162 vaccine not only promote splenocytes proliferation but also active T cell. In addition, the rBCG-EgG1Y162 induced a protection in the mice against secondary infection of Echinococcus granulosus.

  15. EFFECT ON BIOLOGICAL BEHAVIOR OF CHEMOTHERAPY-RESISTANT TUMOR CELLS BY HUMAN WILD-TYPE P53, GM-CSF AND B7-1 GENES VIA RECOMBINANT ADENOVIRUS

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    Objective: To explore the effect on biological behavior of chemotherapy-resistant tumor cells by human wild-type p53, GM-CSF and B7-1 genes mediated via recombinant adenovirus. Methods: p53-abnormal KB-v200 (VCR resistant) and KB-s (VCR sensitive) cell lines were used as model tumor cells, which are resistant and sensitive to chemotherapeutic drugs respectively. After infected with recombinant adenovirus carrying human wild-type p53, GM-CSF and B7-1 genes, changes in biological behavior (including drug sensitivity) of these two kinds of gene-transduced cancer cells were observed. Results: Both of the cell lines were susceptible to adenovirus, all of three exogenous genes (p53, GM-CSF and B7-1) could be effectively expressed in these cell lines, their growth was suppressed, and apoptosis was induced. The drug-pumping-out function of Pgp glycoprotein on the cytomembrane of drug-resistant KB-v200 cells was markedly affected 48h after transfection of the recombinant adenovirus, revealed by increase of the amount of rhodamine 123 accumulation in the cells. The MTT assay also indicated the reversal of their sensitivity to VCR drugs. In vivo experiment in nude mice it was demonstrated reduction of tumorigenicity of the KB-v200 cells or KB-s cells infected with the recombinant adenovirus, and increase of their sensitivity to VCR. Conclusion: The clinical application of this recombinant adenovirus carrying agents might be more effective in treatment of tumors with multidrug resistance (MDR).

  16. Cancer, viruses, and mass migration: Paul Berg's venture into eukaryotic biology and the advent of recombinant DNA research and technology, 1967-1980.

    Science.gov (United States)

    Yi, Doogab

    2008-01-01

    The existing literature on the development of recombinant DNA technology and genetic engineering tends to focus on Stanley Cohen and Herbert Boyer's recombinant DNA cloning technology and its commercialization starting in the mid-1970s. Historians of science, however, have pointedly noted that experimental procedures for making recombinant DNA molecules were initially developed by Stanford biochemist Paul Berg and his colleagues, Peter Lobban and A. Dale Kaiser in the early 1970s. This paper, recognizing the uneasy disjuncture between scientific authorship and legal invention in the history of recombinant DNA technology, investigates the development of recombinant DNA technology in its full scientific context. I do so by focusing on Stanford biochemist Berg's research on the genetic regulation of higher organisms. As I hope to demonstrate, Berg's new venture reflected a mass migration of biomedical researchers as they shifted from studying prokaryotic organisms like bacteria to studying eukaryotic organisms like mammalian and human cells. It was out of this boundary crossing from prokaryotic to eukaryotic systems through virus model systems that recombinant DNA technology and other significant new research techniques and agendas emerged. Indeed, in their attempt to reconstitute 'life' as a research technology, Stanford biochemists' recombinant DNA research recast genes as a sequence that could be rewritten thorough biochemical operations. The last part of this paper shifts focus from recombinant DNA technology's academic origins to its transformation into a genetic engineering technology by examining the wide range of experimental hybridizations which occurred as techniques and knowledge circulated between Stanford biochemists and the Bay Area's experimentalists. Situating their interchange in a dense research network based at Stanford's biochemistry department, this paper helps to revise the canonized history of genetic engineering's origins that emerged during

  17. The Use of Low-Dose Recombinant Tissue Plasminogen Activator to Treat a Preterm Infant with an Intrauterine Spontaneous Arterial Thromboembolis

    Directory of Open Access Journals (Sweden)

    Yaşar Demirelli

    2015-12-01

    Full Text Available Neonatal thromboembolic events are rare, and only a few cases of intrauterine spontaneous arterial thromboembolisms have been reported in the literature. Thrombolytic therapy with recombinant tissue plasminogen activator is usually the preferred treatment because it has a short half-life, fewer systemic side effects, and a strong, specific affinity for fibrin. Protocols vary from center to center, but there is still no consensus regarding the proper dosage or treatment duration. Herein, we present the case of an intrauterine spontaneous arterial thromboembolism in a preterm infant that completely resolved after being treated with low-dose recombinant tissue plasminogen activator (0.02 mg/kg/h.

  18. Sequential combination of two intravenous thrombolytics (recombinant tissue plasminogen activator/tenecteplase) in a patient with stroke and cardioembolic basilar artery occlusion.

    Science.gov (United States)

    Smadja, Didier; Olindo, Stéphane; Saint-Vil, Martine; Chausson, Nicolas

    2009-01-01

    Stroke caused by acute occlusion of basilar artery (AOBA) produces high risk of death. In eligible patients, thrombolysis significantly reduces mortality and disability rate. In most hospitals, thrombolysis is limited to intravenous (IV) route of recombinant tissue plasminogen activator, without any therapeutic alternative in cases of treatment failure. We report a case of cardioembolic AOBA, not responsive to a conventional regimen of IV recombinant tissue plasminogen activator. A sequential combination of IV tenecteplase (0.4 mg/kg) led to a complete recanalization of basilar artery, with a very good clinical outcome. The potential for a combination of two successive IV regimens should be evaluated in AOBA.

  19. Design, Synthesis and Biological Activities of Novel Benzoyl Hydrazines Containing Pyrazole

    Institute of Scientific and Technical Information of China (English)

    闫涛; 于淑晶; 刘鹏飞; 刘卓; 王宝雷; 熊丽霞; 李正名

    2012-01-01

    In search of environmentally benign compounds with high biological activity, low toxicity and low resistance, 8 novel benzoyl hydrazines containing pyrazole were designed and synthesized. All compounds were characterized by I H NMR spectra and HRMS. The preliminary results of biological activity assessment indicated that most of title compounds exhibited certain insecticidal activities against M),thimna separata Walker at 200 mg L-1 but excellent fungicidal activities against six fungus at 50 mg L-1, which were better than the control.

  20. Using Active Learning to Teach Concepts and Methods in Quantitative Biology.

    Science.gov (United States)

    Waldrop, Lindsay D; Adolph, Stephen C; Diniz Behn, Cecilia G; Braley, Emily; Drew, Joshua A; Full, Robert J; Gross, Louis J; Jungck, John A; Kohler, Brynja; Prairie, Jennifer C; Shtylla, Blerta; Miller, Laura A

    2015-11-01

    This article provides a summary of the ideas discussed at the 2015 Annual Meeting of the Society for Integrative and Comparative Biology society-wide symposium on Leading Students and Faculty to Quantitative Biology through Active Learning. It also includes a brief review of the recent advancements in incorporating active learning approaches into quantitative biology classrooms. We begin with an overview of recent literature that shows that active learning can improve students' outcomes in Science, Technology, Engineering and Math Education disciplines. We then discuss how this approach can be particularly useful when teaching topics in quantitative biology. Next, we describe some of the recent initiatives to develop hands-on activities in quantitative biology at both the graduate and the undergraduate levels. Throughout the article we provide resources for educators who wish to integrate active learning and technology into their classrooms.

  1. Impact of cysteine variants on the structure, activity, and stability of recombinant human α-galactosidase A.

    Science.gov (United States)

    Qiu, Huawei; Honey, Denise M; Kingsbury, Jonathan S; Park, Anna; Boudanova, Ekaterina; Wei, Ronnie R; Pan, Clark Q; Edmunds, Tim

    2015-09-01

    Recombinant human α-galactosidase A (rhαGal) is a homodimeric glycoprotein deficient in Fabry disease, a lysosomal storage disorder. In this study, each cysteine residue in rhαGal was replaced with serine to understand the role each cysteine plays in the enzyme structure, function, and stability. Conditioned media from transfected HEK293 cells were assayed for rhαGal expression and enzymatic activity. Activity was only detected in the wild type control and in mutants substituting the free cysteine residues (C90S, C174S, and the C90S/C174S). Cysteine-to-serine substitutions at the other sites lead to the loss of expression and/or activity, consistent with their involvement in the disulfide bonds found in the crystal structure. Purification and further characterization confirmed that the C90S, C174S, and the C90S/C174S mutants are enzymatically active, structurally intact and thermodynamically stable as measured by circular dichroism and thermal denaturation. The purified inactive C142S mutant appeared to have lost part of its alpha-helix secondary structure and had a lower apparent melting temperature. Saturation mutagenesis study on Cys90 and Cys174 resulted in partial loss of activity for Cys174 mutants but multiple mutants at Cys90 with up to 87% higher enzymatic activity (C90T) compared to wild type, suggesting that the two free cysteines play differential roles and that the activity of the enzyme can be modulated by side chain interactions of the free Cys residues. These results enhanced our understanding of rhαGal structure and function, particularly the critical roles that cysteines play in structure, stability, and enzymatic activity.

  2. CBP and p300 histone acetyltransferases contribute to homologous recombination by transcriptionally activating the BRCA1 and RAD51 genes.

    Directory of Open Access Journals (Sweden)

    Hideaki Ogiwara

    Full Text Available Histone acetylation at DNA double-strand break (DSB sites by CBP and p300 histone acetyltransferases (HATs is critical for the recruitment of DSB repair proteins to chromatin. Here, we show that CBP and p300 HATs also function in DSB repair by transcriptionally activating the BRCA1 and RAD51 genes, which are involved in homologous recombination (HR, a major DSB repair system. siRNA-mediated depletion of CBP and p300 impaired HR activity and downregulated BRCA1 and RAD51 at the protein and mRNA levels. Chromatin immunoprecipitation assays showed that CBP and p300 bind to the promoter regions of the BRCA1 and RAD51 genes, and that depletion of CBP and/or p300 reduces H3 and H4 acetylation and inhibits binding of the transcription factor E2F1 to these promoters. Depletion of CBP and p300 impaired DNA damage-induced phosphorylation and chromatin binding of the single-strand DNA-binding protein RPA following BRCA1-mediated DNA end resection. Consistent with this, subsequent phosphorylation of CHK1 and activation of the G2/M damage checkpoint were also impaired. These results indicate that the HATs CBP and p300 play multiple roles in the activation of the cellular response to DSBs.

  3. CBP and p300 histone acetyltransferases contribute to homologous recombination by transcriptionally activating the BRCA1 and RAD51 genes.

    Science.gov (United States)

    Ogiwara, Hideaki; Kohno, Takashi

    2012-01-01

    Histone acetylation at DNA double-strand break (DSB) sites by CBP and p300 histone acetyltransferases (HATs) is critical for the recruitment of DSB repair proteins to chromatin. Here, we show that CBP and p300 HATs also function in DSB repair by transcriptionally activating the BRCA1 and RAD51 genes, which are involved in homologous recombination (HR), a major DSB repair system. siRNA-mediated depletion of CBP and p300 impaired HR activity and downregulated BRCA1 and RAD51 at the protein and mRNA levels. Chromatin immunoprecipitation assays showed that CBP and p300 bind to the promoter regions of the BRCA1 and RAD51 genes, and that depletion of CBP and/or p300 reduces H3 and H4 acetylation and inhibits binding of the transcription factor E2F1 to these promoters. Depletion of CBP and p300 impaired DNA damage-induced phosphorylation and chromatin binding of the single-strand DNA-binding protein RPA following BRCA1-mediated DNA end resection. Consistent with this, subsequent phosphorylation of CHK1 and activation of the G2/M damage checkpoint were also impaired. These results indicate that the HATs CBP and p300 play multiple roles in the activation of the cellular response to DSBs.

  4. Improving Free Radical Scavenging Activity of Soy Isoflavone Glycosides Daidzin and Genistin by 3'-Hydroxylation Using Recombinant Escherichia coli.

    Science.gov (United States)

    Chiang, Chien-Min; Wang, Dong-Sheng; Chang, Te-Sheng

    2016-12-15

    The present study describes the biotransformation of a commercially available crude extract of soy isoflavones, which contained significant amounts of the soy isoflavone glycosides daidzin and genistin, by recombinant Escherichia coli expressing tyrosinase from Bacillus megaterium. Two major products were isolated from the biotransformation and identified as 3'-hydroxydaidzin and 3'-hydroxygenistin, respectively, based on their mass and nuclear magnetic resonance spectral data. The two 3'-hydroxyisoflavone glycosides showed potent 2,2-diphenyl-1-picrylhydrazyl free radical scavenging activity with IC50 values of 7.4 and 9.8 μM for 3'-hydroxydaidzin and 3'-hydroxygenistin, respectively. The free radical scavenging activities of the two 3'-hydroxyisoflavone glycosides were, respectively, 120 and 72 times higher than the activity of their precursors, daidzin and genistin, and were also stronger than the activity of ascorbic acid, which showed an IC50 value of 15.1 μM. This is the first report of the bio-production and potential antioxidant applications of both 3'-hydroxydaidzin and 3'-hydroxygenistin.

  5. Presence of estrogenic activity from emission of fossil fuel combustion as detected by a recombinant yeast bioassay

    Science.gov (United States)

    Wang, Jingxian; Wu, Wenzhong; Henkelmann, Bernhard; You, Li; Kettrup, Antonius; Schramm, Karl-Werner

    Estrogenic activities of emission samples generated by fossil fuel combustion were investigated with human estrogen receptor (ER) recombinant yeast bioassay. The results showed that there were weak but clear estrogenic activities in combustion emissions of fossil fuels including coal, petroleum, and diesel. The estrogenic relative potency (RP) of fossil fuel combustion was the highest in petroleum-fired car, followed by coal-fired stove, diesel-fired agrimotor, coal-fired electric power station. On the other hand, the estrogenic relative inductive efficiency (RIE) was the highest in coal-fired stove and coal-fired electric power station, followed by petroleum-fired car and diesel-fired agrimotor. The estrogenic activities in the sub-fractions from chromatographic separation of emitted materials were also determined. The results indicated that different chemical fractions in these complex systems have different estrogenic potencies. The GC/MS analysis of the emission showed that there were many aromatic carbonyls, big molecular alcohol, PAHs and derivatives, and substituted phenolic compounds and derivatives which have been reported as environmental estrogens. The existence of estrogenic substances in fossil fuel combustion demands further investigation of their potential adverse effects on human and on the ecosystem. The magnitude of pollution due to global usage of fossil fuels makes it imperative to understand the issue of fossil fuel-derived endocrine activities and the associated health risks, particularly the aggregated risks stemmed from exposure to toxicants of multiple sources.

  6. Endogenous and recombinant type I interferons and disease activity in multiple sclerosis.

    Directory of Open Access Journals (Sweden)

    Finn Sellebjerg

    Full Text Available Although treatment of multiple sclerosis (MS with the type I interferon (IFN IFN-β lowers disease activity, the role of endogenous type I IFN in MS remains controversial. We studied CD4+ T cells and CD4+ T cell subsets, monocytes and dendritic cells by flow cytometry and analysed the relationship with endogenous type I IFN-like activity, the effect of IFN-β therapy, and clinical and magnetic resonance imaging (MRI disease activity in MS patients. Endogenous type I IFN activity was associated with decreased expression of the integrin subunit CD49d (VLA-4 on CD4+CD26(high T cells (Th1 helper cells, and this effect was associated with less MRI disease activity. IFN-β therapy reduced CD49d expression on CD4+CD26(high T cells, and the percentage of CD4+CD26(high T cells that were CD49d(high correlated with clinical and MRI disease activity in patients treated with IFN-β. Treatment with IFN-β also increased the percentage of CD4+ T cells expressing CD71 and HLA-DR (activated T cells, and this was associated with an increased risk of clinical disease activity. In contrast, induction of CD71 and HLA-DR was not observed in untreated MS patients with evidence of endogenous type IFN I activity. In conclusion, the effects of IFN-β treatment and endogenous type I IFN activity on VLA-4 expression are similar and associated with control of disease activity. However, immune-activating effects of treatment with IFN-β may counteract the beneficial effects of treatment and cause an insufficient response to therapy.

  7. Recombination instability

    DEFF Research Database (Denmark)

    D'Angelo, N.

    1967-01-01

    A recombination instability is considered which may arise in a plasma if the temperature dependence of the volume recombination coefficient, alpha, is sufficiently strong. Two cases are analyzed: (a) a steady-state plasma produced in a neutral gas by X-rays or high energy electrons; and (b) an af...

  8. ACTIVE AND PARTICIPATORY METHODS IN BIOLOGY: PROBLEM-SOLVING

    Directory of Open Access Journals (Sweden)

    Adela NEMEŞ

    2010-01-01

    Full Text Available We face with considerable challenge of developing students’ problem solving skills in our difficult environment. Good problem solving skills empower managers in their professional and personal lives. Problem solving skills are valued by academics and employers. The informations in Biology are often presented in abstract forms without contextualisation. Creative problem-solving process involves a few steps, which together provide a structured procedure for identifying challenges, generating ideas and implementing innovative solutions: identifying the problem, searching for possible solutions, selecting the most optimal solution and implementing a possible solution. Each aspect of personality has a different orientation to problem solving, different criteria for judging the effectiveness of the process and different associated strengths. Using real-world data in sample problems will also help facilitate the transfer process, since students can more easily identify with the context of a given situation. The paper describes the use of the Problem-Solving in Biology and the method of its administration. It also presents the results of a study undertaken to evaluate the value in teaching Biology. Problem-solving is seen as an essential skill that is developed in biology education.

  9. 78 FR 66751 - Office of Science Policy, Office of Biotechnology Activities; Recombinant or Synthetic Nucleic...

    Science.gov (United States)

    2013-11-06

    ... HUMAN SERVICES National Institutes of Health Office of Science Policy, Office of Biotechnology... Biotechnology Activities (OBA) is updating Appendix B (Classification of Human Etiologic Agents on the Basis of... mail to the Office of Biotechnology Activities, National Institutes of Health, 6705 Rockledge...

  10. Phlorotannins from Ecklonia cava (Phaeophyceae): biological activities and potential health benefits.

    Science.gov (United States)

    Wijesekara, Isuru; Yoon, Na Young; Kim, Se-Kwon

    2010-01-01

    The importance of bioactive derivatives as functional ingredients has been well recognized due to their valuable health beneficial effects. Therefore, isolation and characterization of novel functional ingredients with biological activities from seaweeds have gained much attention. Ecklonia cava Kjellman is an edible seaweed, which has been recognized as a rich source of bioactive derivatives mainly, phlorotannins. These phlorotannins exhibit various beneficial biological activities such as antioxidant, anticancer, antidiabetic, anti-human immunodeficiency virus, antihypertensive, matrix metalloproteinase enzyme inhibition, hyaluronidase enzyme inhibition, radioprotective, and antiallergic activities. This review focuses on biological activities of phlorotannins with potential health beneficial applications in functional foods, pharmaceuticals, and cosmeceuticals.

  11. Preparation and immune activity analysis of H5N1 subtype avian influenza virus recombinant protein-based vaccine.

    Science.gov (United States)

    Xie, Q M; Ji, J; Du, L Q; Cao, Y C; Wei, L; Xue, C Y; Qin, J P; Ma, J Y; Bi, Y Z

    2009-08-01

    Avian influenza is a severe disease among farmed poultry and free-living birds and a constant threat to the commercial chicken industry around the world. Hemagglutinin (HA) is the major immunogen on the envelope of influenza A virus and is the predominant inducer of neutralizing antibody. To obtain the bioactive antigen proteins in large quantities, a new protein expression vector pBCX was constructed, which is based on the pET32a vector. The HA gene of the H5N1 subtype of avian influenza virus (AIV) was inserted into the pBCX vector and expressed efficiently in Escherichia coli BL21 (DE3). Fused expression of the exogenous gene and msyB produced a 97-kDa msyB-HA fusion protein. Sodium dodecyl sulfate-PAGE combined with scanning analysis demonstrated that the msyB-HA fusion protein accounted for 29.5% of the total bacterial protein, 90.5% being soluble. The msyB-HA fusion protein was purified with nondenaturing 50% Ni-NTA column chromatography, and the result showed that 24 mg of purified msyB-HA fusion protein could be obtained from 1 L of induced expression bacterial culture medium. The comparative results in the present study showed that pBCX was superior to pET32a as a protein expression vector. Western blotting showed the recombinant msyB-HA (rHA) to have better antigenic activity, which may be the result from the better posttranslation protein modification and folding in the pBCX expression system. With the rHA fusion protein as antigen, we successfully prepared and screened specific monoclonal antibodys against the H5N1 subtype AIV, which indicated that the rHA had antigen epitopes and biofunctions. The immune test confirmed that the rHA protein vaccine could also induce high neutralizing antibodies, and the AIV challenge test proved that the rHA protein-based vaccine could prevent the corresponding infection. This study demonstrates that the recombinant HA protein produced by the pBCX expression system could be used as a recombinant protein-based vaccine

  12. Epidemiological, biological and histological characterization of patients with indeterminate third-generation recombinant immunoblot assay antibody results for hepatitis C virus.

    Science.gov (United States)

    Ríos, M; Diago, M; Rivera, P; Tuset, C; Cors, R; García, V; Carbonel, P; Gonzalez, C

    2006-03-01

    We studied the epidemiological, laboratory and histological characteristics of a group of patients with positive antibodies against hepatitis C virus (HCV) as determined by third-generation enzyme-linked immunosorbent assay (ELISA), and with indeterminate HCV antibody positivity as established by third-generation recombinant immunoblot assay (RIBA-3). The results obtained were compared with those recorded in a group of RIBA-3-positive patients. Both groups correspond to blood donors in whom the prevalence of hepatitis C is low. There were no statistically significant intergroup differences in mean age, or in the presence of infection risk factors. RNA positivity was much more frequent in the RIBA-positive group (71%vs 10%; P < 0.05), as was transaminase elevation during the 3 years of follow-up (54%vs 13%; P < 0.05). In 46% of the RIBA-indeterminate patients the liver biopsy proved normal, or only liver steatosis or minimal changes were detected, while 33% had persistent chronic hepatitis, and 21% showed active chronic hepatitis. A mean Knodell index score of 2.28 was recorded; 50% of the subjects showed no fibrosis, 46% grade 1 fibrosis (fibrous portal expansion), 4% grade 2 fibrosis (bridging fibrosis), and none grade 3 fibrosis (liver cirrhosis). In the RIBA-positive group, a greater percentage of patients had active chronic hepatitis, a greater Knodell index, and increased-grade fibrosis. It can be concluded that the RIBA-3-indeterminate group is epidemiologically similar to the RIBA-3-positive series, although with a lesser prevalence of laboratory test alterations, a lower viral replication index, and more likely to have benign disease - particularly in subjects without viral replication.

  13. Endogenous and recombinant type I interferons and disease activity in multiple sclerosis

    DEFF Research Database (Denmark)

    Sellebjerg, Finn; Krakauer, Martin; Limborg, Signe;

    2012-01-01

    with endogenous type I IFN-like activity, the effect of IFN-ß therapy, and clinical and magnetic resonance imaging (MRI) disease activity in MS patients. Endogenous type I IFN activity was associated with decreased expression of the integrin subunit CD49d (VLA-4) on CD4+CD26(high) T cells (Th1 helper cells......Although treatment of multiple sclerosis (MS) with the type I interferon (IFN) IFN-ß lowers disease activity, the role of endogenous type I IFN in MS remains controversial. We studied CD4+ T cells and CD4+ T cell subsets, monocytes and dendritic cells by flow cytometry and analysed the relationship...... the percentage of CD4+ T cells expressing CD71 and HLA-DR (activated T cells), and this was associated with an increased risk of clinical disease activity. In contrast, induction of CD71 and HLA-DR was not observed in untreated MS patients with evidence of endogenous type IFN I activity. In conclusion...

  14. Adjuvant activity of chicken interleukin-12 co-administered with infectious bursal disease virus recombinant VP2 antigen in chickens.

    Science.gov (United States)

    Su, Bor Sheu; Chiu, Hua Hsien; Lin, Cheng Chung; Shien, Jui Hung; Yin, Hsien Sheng; Lee, Long Huw

    2011-02-15

    A recombinant fowlpox virus (rFPV/VP2) expressing infectious bursal diseases virus (IBDV) VP2 gene has been constructed. After purification and identification of rFPV/VP2, the adjuvant activity of the recombinant chicken IL-12 (rchIL-12), synthesized by our previous construct of rFPV/chIL-12, in rFPV/VP2-expressed rVP2 antigen was assessed in one-week-old specific-pathogen free chickens. The results indicated that rchIL-12 alone or rchIL-12 plus mineral oil (MO) co-administered with rVP2 antigen significantly enhanced the production of serum neutralization (SN) antibody against IBDV, compared to those with MO alone. The SN titers in groups receiving rVP2 antigen with MO alone were more inconsistent after vaccination. On the other hand, rchIL-12 significantly stimulated IFN-γ production in serum and in splenocyte cultured supernatant, suggesting that rchIL-12 alone or plus MO significantly induced a cell-mediated immune response. Finally, bursal lesion protection from very virulent IBDV (vvIBDV) challenge in chickens receiving rVP2 antigen with rchIL-12 alone or plus MO was much more effective than that with MO alone at two weeks after boosting. Taken together, rchIL-12 alone augmented in vivo the induction of a primary and also a secondary SN antibody production and a cell-mediated immunity against IBDV rVP2 antigen, which conferred the enhancement of bursal lesion protective efficacy from vvIBDV challenge. These data indicated that a potential for chIL-12 as immunoadjuvant for chicken vaccine development such as IBDV rVP2 antigen.

  15. Highly efficient expression, purification of recombinant LTB protein and its activity against mucosal immunoadjuvant by nasal immunization

    Institute of Scientific and Technical Information of China (English)

    王静; 李琳琳; 郑瑾; 于军; 杨筱凤; 耿宜平; 来宝长; 王一理; 司履生

    2003-01-01

    Objective To develop an efficient expression, purification system of recombinant Escherichia coli heat-labile enterotoxin B subunit (rLTB) and study its activity against mucosal immunoadjuvant by nasal immunization.Methods A recombinant, pMMB68-LTB was generated by cloning the LTB cDNA fragment into an expression vector (pMMB68) and transformed it into the host strain marine vibrio VSP60. The relevant target protein was identified using SDS-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blot. Sephacryl S-100 gel filtration chromatography was carried out for purification of rLTB in engineering bacteria VSP60. BALB/c mice received hen egg lysozyme (HEL) alone or combined with rLTB by nasal administration. After three times immunization, IgG and IgA antibody levels in serum or small intestine wash samples were determined using ELISA.Results rLTB protein was highly expressed in VSP60. After gel filtration with Sephacryl S-100, the purity of rLTB reached 98.1%, the yield rate was about 52%. After immunization, IgG and IgA antibody responses specific to HEL in system and mucosa of HEL+rLTB groups were significantly increased, compared with the HEL alone group (P<0.001).Conclusions A set of protocols for large-scale rLTB preparation has been established, which is simple, efficient and applicable. The rLTB protein we prepared was proved to be a powerful mucocal adjuvant, which could greatly enhance systemic and mucosal immune responses to nasally co-administered antigen.

  16. Yeast homologous recombination-based promoter engineering for the activation of silent natural product biosynthetic gene clusters.

    Science.gov (United States)

    Montiel, Daniel; Kang, Hahk-Soo; Chang, Fang-Yuan; Charlop-Powers, Zachary; Brady, Sean F

    2015-07-21

    Large-scale sequencing of prokaryotic (meta)genomic DNA suggests that most bacterial natural product gene clusters are not expressed under common laboratory culture conditions. Silent gene clusters represent a promising resource for natural product discovery and the development of a new generation of therapeutics. Unfortunately, the characterization of molecules encoded by these clusters is hampered owing to our inability to express these gene clusters in the laboratory. To address this bottleneck, we have developed a promoter-engineering platform to transcriptionally activate silent gene clusters in a model heterologous host. Our approach uses yeast homologous recombination, an auxotrophy complementation-based yeast selection system and sequence orthogonal promoter cassettes to exchange all native promoters in silent gene clusters with constitutively active promoters. As part of this platform, we constructed and validated a set of bidirectional promoter cassettes consisting of orthogonal promoter sequences, Streptomyces ribosome binding sites, and yeast selectable marker genes. Using these tools we demonstrate the ability to simultaneously insert multiple promoter cassettes into a gene cluster, thereby expediting the reengineering process. We apply this method to model active and silent gene clusters (rebeccamycin and tetarimycin) and to the silent, cryptic pseudogene-containing, environmental DNA-derived Lzr gene cluster. Complete promoter refactoring and targeted gene exchange in this "dead" cluster led to the discovery of potent indolotryptoline antiproliferative agents, lazarimides A and B. This potentially scalable and cost-effective promoter reengineering platform should streamline the discovery of natural products from silent natural product biosynthetic gene clusters.

  17. Presence of estrogenic activity from emission of fossil fuel combustion as detected by a recombinant yeast bioassay

    Energy Technology Data Exchange (ETDEWEB)

    Wang, J.X.; Wu, W.Z.; Henkelmann, B.; You, L.; Kettrup, A.; Schramm, K.W. [GSF, Neuherberg (Germany). National Research Center for Environmental & Health

    2003-07-01

    Estrogenic activities of emission samples generated by fossil fuel combustion were investigated with human estrogen receptor recombinant yeast bioassay. The results showed that there were weak but clear estrogenic activities in combustion emissions of fossil fuels including coal, petroleum, and diesel. The estrogenic relative potency of fossil fuel combustion was the highest in petroleum-fired car, followed by coal-fired stove, diesel-fired agrimotor, coal-fired electric power station. On the other hand, the estrogenic relative inductive efficiency was the highest in coal-fired stove and coal-fired electric power station, followed by petroleum-fired car and diesel-fired agrimotor. The estrogenic activities in the sub-fractions from chromatographic separation of emitted materials measured. The results indicated that different chemical fractions in these complex systems have different estrogenic potencies. The GC/MS analysis of the emission showed that there were many aromatic carbonyls, big molecular alcohol, PAHs and derivatives, and substituted phenolic compounds and derivatives which have been reported as environmental estrogens. The existence of estrogenic substances in fossil fuel combustion demands further investigation of their potential adverse effects on human and on the ecosystem.

  18. Physicochemical and porosity characteristics of thermally regenerated activated carbon polluted with biological activated carbon process.

    Science.gov (United States)

    Dong, Lihua; Liu, Wenjun; Jiang, Renfu; Wang, Zhansheng

    2014-11-01

    The characteristics of thermally regenerated activated carbon (AC) polluted with biological activated carbon (BAC) process were investigated. The results showed that the true micropore and sub-micropore volume, pH value, bulk density, and hardness of regenerated AC decreased compared to the virgin AC, but the total pore volume increased. XPS analysis displayed that the ash contents of Al, Si, and Ca in the regenerated AC respectively increased by 3.83%, 2.62% and 1.8%. FTIR spectrum showed that the surface functional groups of virgin and regenerated AC did not change significantly. Pore size distributions indicated that the AC regeneration process resulted in the decrease of micropore and macropore (D>10 μm) volume and the increase of mesopore and macropore (0.1 μmbiological waste (spent AC) from BAC process.

  19. Top3 processes recombination intermediates and modulates checkpoint activity after DNA damage

    DEFF Research Database (Denmark)

    Mankouri, Hocine W; Hickson, Ian D

    2006-01-01

    Mutation of TOP3 in Saccharomyces cerevisiae causes poor growth, hyperrecombination, and a failure to fully activate DNA damage checkpoints in S phase. Here, we report that overexpression of a dominant-negative allele of TOP3, TOP3(Y356F), which lacks the catalytic (decatenation) activity of Top3......, causes impaired S-phase progression and the persistence of abnormal DNA structures (X-shaped DNA molecules) after exposure to methylmethanesulfonate. The impaired S-phase progression is due to a persistent checkpoint-mediated cell cycle delay and can be overridden by addition of caffeine. Hence......, the catalytic activity of Top3 is not required for DNA damage checkpoint activation, but it is required for normal S-phase progression after DNA damage. We also present evidence that the checkpoint-mediated cell cycle delay and persistence of X-shaped DNA molecules resulting from overexpression of TOP3(Y356F...

  20. Recombinant lectins: an array of tailor-made glycan-interaction biosynthetic tools.

    Science.gov (United States)

    Oliveira, Carla; Teixeira, José A; Domingues, Lucília

    2013-03-01

    Lectins are a heterogeneous group of proteins found in plants, animals and microorganisms, which possess at least one non-catalytic domain that binds reversibly to specific mono- or oligosaccharides. The range of lectins and respective biological activities is unsurprising given the immense diversity and complexity of glycan structures and the multiple modes of interaction with proteins. Recombinant DNA technology has been traditionally used for cloning and characterizing newly discovered lectins. It has also been employed as a means of producing pure and sequence-defined lectins for different biotechnological applications. This review focuses on the production of recombinant lectins in heterologous organisms, and highlighting the Escherichia coli and Pichia pastoris expression systems, which are the most employed. The choice of expression host depends on the lectin. Non-glycosylated recombinant lectins are produced in E. coli and post-translational modified recombinant lectins are produced in eukaryotic organisms, namely P. pastoris and non-microbial hosts such as mammalian cells. Emphasis is given to the applications of the recombinant lectins especially (a) in cancer diagnosis and/or therapeutics, (b) as anti-microbial, anti-viral, and anti-insect molecules or (c) in microarrays for glycome profiling. Most reported applications are from recombinant plant lectins. These applications benefit from the tailor-made design associated with recombinant production and will aid in unraveling the complex biological mechanisms of glycan-interactions, bringing recombinant lectins to the forefront of glycobiology. In conclusion, recombinant lectins are developing into valuable biosynthetic tools for biomedical research.

  1. Mango ginger (Curcuma amada Roxb.) – A promising spice for phytochemicals and biological activities

    Indian Academy of Sciences (India)

    R S Policegoudra; S M Aradhya; L Singh

    2011-09-01

    Mango ginger (Curcuma amada Roxb.) is a unique spice having morphological resemblance with ginger but imparts a raw mango flavour. The main use of mango ginger rhizome is in the manufacture of pickles and culinary preparations. Ayurveda and Unani medicinal systems have given much importance to mango ginger as an appetizer, alexteric, antipyretic, aphrodisiac, diuretic, emollient, expectorant and laxative and to cure biliousness, itching, skin diseases, bronchitis, asthma, hiccough and inflammation due to injuries. The biological activities of mango ginger include antioxidant activity, antibacterial activity, antifungal activity, anti-inflammatory activity, platelet aggregation inhibitory activity, cytotoxicity, antiallergic activity, hypotriglyceridemic activity, brine-shrimp lethal activity, enterokinase inhibitory activity, CNS depressant and analgesic activity. The major chemical components include starch, phenolic acids, volatile oils, curcuminoids and terpenoids like difurocumenonol, amadannulen and amadaldehyde. This article brings to light the major active components present in C. amada along with their biological activities that may be important from the pharmacological point of view.

  2. Combined treatment with recombinant tissue plasminogen activator and dexamethasone phosphate-containing liposomes improves neurological outcome and restricts lesion progression after embolic stroke in rats

    NARCIS (Netherlands)

    Tiebosch, I.A.; Crielaard, B.J.; Bouts, M.J.; Salas-Perdomo, A.; Lammers, T.G.G.M.; Planas, A.M.; Storm, G.; Dijkhuizen, R.M.

    2012-01-01

    Abstract Variable efficacies have been reported for glucocorticoid drugs as anti-inflammatory treatment after stroke. We applied an alternative drug delivery strategy, by injection of dexamethasone phosphate-containing liposomes in combination with recombinant tissue plasminogen activator (rtPA), i

  3. Recombinant host cells and nucleic acid constructs encoding polypeptides having cellulolytic enhancing activity

    Energy Technology Data Exchange (ETDEWEB)

    Schnorr, Kirk; Kramer, Randall

    2017-03-28

    The present invention relates to isolated polypeptides having cellulolytic enhancing activity and isolated polynucleotides encoding the polypeptides. The invention also relates to nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as methods of producing and using the polypeptides.

  4. Biological Opinion Compliance Report and Permit Activity Report

    Science.gov (United States)

    1999-03-01

    T.W. Custer. 1986. Hatching Success of Caspian Terns Nesting on the Lower Laguna Madre , Texas , USA. Colonial Birds. 9(1). Pp 86-89. Persons working with...U.S. Fish and Wildlife Service on November 14,’ 1 990. Initial District implementation of that Biological Opinion began in 1 992, following the...known as the "Red Book." The Red Book was reviewed by the U.S. Fish and Wildlife Service and approved for implementation by the Missouri River Division

  5. 111Indium Labelling of Recombinant Activated Coagulation Factor VII: In Vitro and Preliminary In Vivo Studies in Healthy Rats

    Directory of Open Access Journals (Sweden)

    Amarnadh Nalla

    2012-01-01

    Full Text Available The aim of this study is to investigate whether 111Indium-labelled recombinant FVIIa (rFVIIa could be a potential radiopharmaceutical for localization of bleeding sources. DTPA-conjugated rFVIIa was radiolabelled with 111In chloride. In vitro binding efficiency of 111In-DTPA-rFVIIa to F1A2-Mab-sepharose was 99% in buffer, while it was 88–82% in serum. The binding efficiency of 111In-DTPA-rFVIIa to TF (1–209-sepharose was 48% in buffer whereas 39%–36% in serum, respectively. In vivo experiment was conducted in healthy rats, and gamma camera images were taken immediately after iv. administration of 1.6–1.8 MBq 111In-DTPA-rFVIIa up to 120–130 min. Five min after administration of 111In-DTPA-rFVIIa, percentage of 111In activity was 6.0% in the cardiac region and 24.5% in the liver region. After 2 hours activity was decreased to 3.3% in heart while it had increased to 42.0% in the liver. The 111In-DTPA-rFVIIa might be a potential radiopharmaceutical for visualisation of tissues with significant TF expression such as acute bleeding lesions in the gastrointestinal tract.

  6. S-S synapsis during class switch recombination is promoted by distantly located transcriptional elements and activation-induced deaminase.

    Science.gov (United States)

    Wuerffel, Robert; Wang, Lili; Grigera, Fernando; Manis, John; Selsing, Erik; Perlot, Thomas; Alt, Frederick W; Cogne, Michel; Pinaud, Eric; Kenter, Amy L

    2007-11-01

    Molecular mechanisms underlying synapsis of activation-induced deaminase (AID)-targeted S regions during class switch recombination (CSR) are poorly understood. By using chromosome conformation capture techniques, we found that in B cells, the Emicro and 3'Ealpha enhancers were in close spatial proximity, forming a unique chromosomal loop configuration. B cell activation led to recruitment of the germline transcript (GLT) promoters to the Emicro:3'Ealpha complex in a cytokine-dependent fashion. This structure facilitated S-S synapsis because Smicro was proximal to Emicro and a downstream S region was corecruited with the targeted GLT promoter to Emicro:3'Ealpha. We propose that GLT promoter association with the Emicro:3'Ealpha complex creates an architectural scaffolding that promotes S-S synapsis during CSR and that these interactions are stabilized by AID. Thus, the S-S synaptosome is formed as a result of the self-organizing transcription system that regulates GLT expression and may serve to guard against spurious chromosomal translocations.

  7. Mediator facilitates transcriptional activation and dynamic long-range contacts at the IgH locus during class switch recombination

    Science.gov (United States)

    Thomas-Claudepierre, Anne-Sophie; Robert, Isabelle; Rocha, Pedro P.; Raviram, Ramya; Schiavo, Ebe; Heyer, Vincent; Bonneau, Richard; Luo, Vincent M.; Reddy, Janardan K.; Borggrefe, Tilman; Skok, Jane A.

    2016-01-01

    Immunoglobulin (Ig) class switch recombination (CSR) is initiated by the transcription-coupled recruitment of activation-induced cytidine deaminase (AID) to Ig switch regions (S regions). During CSR, the IgH locus undergoes dynamic three-dimensional structural changes in which promoters, enhancers, and S regions are brought to close proximity. Nevertheless, little is known about the underlying mechanisms. In this study, we show that Med1 and Med12, two subunits of the mediator complex implicated in transcription initiation and long-range enhancer/promoter loop formation, are dynamically recruited to the IgH locus enhancers and the acceptor regions during CSR and that their knockdown in CH12 cells results in impaired CSR. Furthermore, we show that conditional inactivation of Med1 in B cells results in defective CSR and reduced acceptor S region transcription. Finally, we show that in B cells undergoing CSR, the dynamic long-range contacts between the IgH enhancers and the acceptor regions correlate with Med1 and Med12 binding and that they happen at a reduced frequency in Med1-deficient B cells. Our results implicate the mediator complex in the mechanism of CSR and are consistent with a model in which mediator facilitates the long-range contacts between S regions and the IgH locus enhancers during CSR and their transcriptional activation. PMID:26903242

  8. Mediator facilitates transcriptional activation and dynamic long-range contacts at the IgH locus during class switch recombination.

    Science.gov (United States)

    Thomas-Claudepierre, Anne-Sophie; Robert, Isabelle; Rocha, Pedro P; Raviram, Ramya; Schiavo, Ebe; Heyer, Vincent; Bonneau, Richard; Luo, Vincent M; Reddy, Janardan K; Borggrefe, Tilman; Skok, Jane A; Reina-San-Martin, Bernardo

    2016-03-07

    Immunoglobulin (Ig) class switch recombination (CSR) is initiated by the transcription-coupled recruitment of activation-induced cytidine deaminase (AID) to Ig switch regions (S regions). During CSR, the IgH locus undergoes dynamic three-dimensional structural changes in which promoters, enhancers, and S regions are brought to close proximity. Nevertheless, little is known about the underlying mechanisms. In this study, we show that Med1 and Med12, two subunits of the mediator complex implicated in transcription initiation and long-range enhancer/promoter loop formation, are dynamically recruited to the IgH locus enhancers and the acceptor regions during CSR and that their knockdown in CH12 cells results in impaired CSR. Furthermore, we show that conditional inactivation of Med1 in B cells results in defective CSR and reduced acceptor S region transcription. Finally, we show that in B cells undergoing CSR, the dynamic long-range contacts between the IgH enhancers and the acceptor regions correlate with Med1 and Med12 binding and that they happen at a reduced frequency in Med1-deficient B cells. Our results implicate the mediator complex in the mechanism of CSR and are consistent with a model in which mediator facilitates the long-range contacts between S regions and the IgH locus enhancers during CSR and their transcriptional activation.

  9. Engineering the cellular protein secretory pathway for enhancement of recombinant tissue plasminogen activator expression in Chinese hamster ovary cells: effects of CERT and XBP1s genes.

    Science.gov (United States)

    Rahimpour, Azam; Vaziri, Behrouz; Moazzami, Reza; Nematollahi, Leila; Barkhordari, Farzaneh; Kokabee, Leila; Adeli, Ahmad; Mahboudi, Fereidoun

    2013-08-01

    Cell line development is the most critical and also the most time-consuming step in the production of recombinant therapeutic proteins. In this regard, a variety of vector and cell engineering strategies have been developed for generating high-producing mammalian cells; however, the cell line engineering approach seems to show various results on different recombinant protein producer cells. In order to improve the secretory capacity of a recombinant tissue plasminogen activator (t-PA)-producing Chinese hamster ovary (CHO) cell line, we developed cell line engineering approaches based on the ceramide transfer protein (CERT) and X-box binding protein 1 (XBP1) genes. For this purpose, CERT S132A, a mutant form of CERT that is resistant to phosphorylation, and XBP1s were overexpressed in a recombinant t-PA-producing CHO cell line. Overexpression of CERT S132A increased the specific productivity of t-PA-producing CHO cells up to 35%. In contrast, the heterologous expression of XBP1s did not affect the t-PA expression rate. Our results suggest that CERTS132A- based secretion engineering could be an effective strategy for enhancing recombinant t- PA production in CHO cells.

  10. Activities of wildtype and mutant p53 in suppression of homologous recombination as measured by a retroviral vector system

    Energy Technology Data Exchange (ETDEWEB)

    Lu Xiongbin; Lozano, Guillermina; Donehower, Lawrence A

    2003-01-28

    DNA repair of double strand breaks, interstrand DNA cross-links, and other types of DNA damage utilizes the processes of homologous recombination and non-homologous end joining to repair the damage. Aberrant homologous recombination is likely to be responsible for a significant fraction of chromosomal deletions, duplications, and translocations that are observed in cancer cells. To facilitate measurement of homologous recombination frequencies in normal cells, mutant cells, and cancer cells, we have developed a high titer retroviral vector containing tandem repeats of mutant versions of a GFP-Zeocin resistance fusion gene and an intact neomycin resistance marker. Recombination between the tandem repeats regenerates a functional GFP-Zeo{sup R} marker that can be easily scored. This retroviral vector was used to assess homologous recombination frequencies in human cancer cells and rodent fibroblasts with differing dosages of wild type or mutant p53. Absence of wild type p53 stimulated spontaneous and ionizing radiation-induced homologous recombination, confirming previous studies. Moreover, p53{sup +/-} mouse fibroblasts show elevated levels of homologous recombination compared to their p53{sup +/+} counterparts following retroviral vector infection, indicating that p53 is haploinsufficient for suppression of homologous recombination. Transfection of vector-containing p53 null Saos-2 cells with various human cancer-associated p53 mutants revealed that these altered p53 proteins retain some recombination suppression function despite being totally inactive for transcriptional transactivation. The retroviral vector utilized in these studies may be useful in performing recombination assays on a wide array of cell types, including those not readily transfected by normal vectors.

  11. Activities of wildtype and mutant p53 in suppression of homologous recombination as measured by a retroviral vector system.

    Science.gov (United States)

    Lu, Xiongbin; Lozano, Guillermina; Donehower, Lawrence A

    2003-01-28

    DNA repair of double strand breaks, interstrand DNA cross-links, and other types of DNA damage utilizes the processes of homologous recombination and non-homologous end joining to repair the damage. Aberrant homologous recombination is likely to be responsible for a significant fraction of chromosomal deletions, duplications, and translocations that are observed in cancer cells. To facilitate measurement of homologous recombination frequencies in normal cells, mutant cells, and cancer cells, we have developed a high titer retroviral vector containing tandem repeats of mutant versions of a GFP-Zeocin resistance fusion gene and an intact neomycin resistance marker. Recombination between the tandem repeats regenerates a functional GFP-Zeo(R) marker that can be easily scored. This retroviral vector was used to assess homologous recombination frequencies in human cancer cells and rodent fibroblasts with differing dosages of wild type or mutant p53. Absence of wild type p53 stimulated spontaneous and ionizing radiation-induced homologous recombination, confirming previous studies. Moreover, p53(+/-) mouse fibroblasts show elevated levels of homologous recombination compared to their p53(+/+) counterparts following retroviral vector infection, indicating that p53 is haploinsufficient for suppression of homologous recombination. Transfection of vector-containing p53 null Saos-2 cells with various human cancer-associated p53 mutants revealed that these altered p53 proteins retain some recombination suppression function despite being totally inactive for transcriptional transactivation. The retroviral vector utilized in these studies may be useful in performing recombination assays on a wide array of cell types, including those not readily transfected by normal vectors.

  12. Recombinant Helicobacter pylori catalase

    Institute of Scientific and Technical Information of China (English)

    Yang Bai; Ya-Li Zhang; Jian-Feng Jin; Ji-De Wang; Zhao-Shan Zhang

    2003-01-01

    AIM: To construct a recombinant strain which highly expresses catalase of Helicobacter pylori(H.pylori) and assay the activity of H. pylori catalase.METHODS: The catalase DNA was amplified from H. pylori chromosomal DNA with PCR techniques and inserted into the prokaryotie expression vector pET-22b (+), and then was transformed into the BL21 (DE3) E. coli strain which expressed catalase recombinant protein. The activity of H.pylori catalase was assayed by the Beers & Sizers.RESULTS: DNA sequence analysis showed that the sequence of catalase DNA was the same as GenBank's research. The catalase recombinant protein amounted to 24.4 % of the total bacterial protein after induced with IPTG for 3 hours at 37 ℃ and the activity of H. pylori catalase was high in the BL21 (DE3) E. coli strain.CONCLUSION: A clone expressing high activity H. pylori catalase is obtained, laying a good foundation for further studies.

  13. Evaluation of in vivo biological activity profile of isoorientin.

    Science.gov (United States)

    Küpeli, Esra; Aslan, Mustafa; Gürbüz, Ilhan; Yesilada, Erdem

    2004-01-01

    Anti-nociceptive, anti-inflammatory and gastroprotective activities of the known C-glycosyl flavonoid, isoorientin, were studied in rats and mice. For the anti-nociceptive activity assessment the p-benzoquinone-induced writing test, for the anti-inflammatory activity the carrageenan-induced hind paw edema model in mice, and for the gastroprotective activity the EtOH-induced ulcerogenesis model in rats were used. Isoorientin was shown to possess significant anti-nociceptive and anti-inflammatory activities at 15 mg/kg and 30 mg/kg doses, without inducing any apparent acute toxicity as well as gastric damage. However, the compound did not possess any significant gastroprotective activity against EtOH-induced ulcerogenesis.

  14. Recombination monitor

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, S. Y. [Brookhaven National Lab. (BNL), Upton, NY (United States); Blaskiewicz, M. [Brookhaven National Lab. (BNL), Upton, NY (United States)

    2017-02-03

    This is a brief report on LEReC recombination monitor design considerations. The recombination produced Au78+ ion rate is reviewed. Based on this two designs are discussed. One is to use the large dispersion lattice. It is shown that even with the large separation of the Au78+ beam from the Au79+ beam, the continued monitoring of the recombination is not possible. Accumulation of Au78+ ions is needed, plus collimation of the Au79+ beam. In another design, it is shown that the recombination monitor can be built based on the proposed scheme with the nominal lattice. From machine operation point of view, this design is preferable. Finally, possible studies and the alternative strategies with the basic goal of the monitor are discussed.

  15. Synthesis and biological activity of benzamide DNA minor groove binders.

    Science.gov (United States)

    Khan, Gul Shahzada; Pilkington, Lisa I; Barker, David

    2016-02-01

    A range of di- and triaryl benzamides were synthesised to investigate the effect of the presence and nature of a polar sidechain, bonding and substitution patterns and functionalisation of benzylic substituents. These compounds were tested for their antiproliferative activity as well as their DNA binding activity. The most active compounds in all assays were unsymmetrical triaryl benzamides with a bulky or alkylating benzylic substituent and a polar amino sidechain.

  16. Biological activity of fucoidan from leafy bladderwort (Utricularia aurea Lour.)

    OpenAIRE

    Chotigeat, W.; Phongdara, A.; Choosawad, D.

    2005-01-01

    Fucoidan extracted from aquatic plant Utricularia aurea was 1.3% of dry weight and comprised glucuronic acid 62.5% and fucose 4.98% of the crude extract and 28.74% of the fucose was sulfate. The fucoidan was investigated for antibacterial activity. The minimal inhibitory concentration (MIC) of crude fucoidan against Vibrio harveyi and Escherichia coli was 20 and 10 mg/ml. Moreover, the fucoidan had anticoagulant activity. The crude fucoidan had activated partial thromboplastin time (APTT) at ...

  17. Studies on Synthesis and Biological Activities of Novel Triazole Compounds Containing 1,3-Dioxolane

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    Nine title compounds were synthesized. Their strucures were identified by means of IR, EA, 1H NMR and MS. The results from the primary biological test show that all the compounds have some activitiies of fungicide and plant growth regulator. When R group is 2,4-Cl2C6H3, compound 2 or compound 4 shows better biological activities.

  18. Novel Plant-Derived Recombinant Human Interferons with Broad Spectrum Antiviral Activity

    Science.gov (United States)

    2011-10-14

    compared the antiviral activities of more than 1400 plant-derived, hybrid IFNs against three RNA viruses and one DNA virus from four different families...highly pathogenic viruses with varying sensitivities to type I IFN. In particular, the DNA virus , MPXV, was not expected to be as susceptible to the...K.M., Callis, R.T., Stephen, E.L., 1980. Lassa virus infection of rhesus monkeys: pathogenesis and treatment with ribavirin. J. Infect. Dis. 141, 580

  19. Plasmodium falciparum Plasmepsin V (PfPMV): Insights into recombinant expression, substrate specificity and active site structure.

    Science.gov (United States)

    Boonyalai, Nonlawat; Sittikul, Pichamon; Yuvaniyama, Jirundon

    2015-05-01

    Plasmepsin V from Plasmodium falciparum (PfPMV) is responsible for the cleavage of the Plasmodium export element (PEXEL) motif at the N-terminus of several hundreds of the exported proteins. PfPMV is necessary for parasite viability and has become a novel promising target for antimalarial therapy. The first recombinant expression of soluble, active PfPMV as thioredoxin fusion proteins is reported herein. Two truncated forms of PfPMV were fused to thioredoxin (Trx) to generate Trx-PfPMVp37 and Trx-PfPMVm84. The fusion proteins were successfully purified using Ni(2+) affinity chromatography in combination with ATP treatment to eliminate Escherichia coli HSP60 contaminant. Trx-PfPMVm84 could hydrolyze the PEXEL-containing peptides more efficiently than Trx-PfPMVp37. Interestingly, both Trx-PfPMVs preferred to cleave PfEMP2 peptide over HRPII peptide. The replacement of Ser with Val or Glu at P1' position created a substrate with 75% reduction in the enzyme activity, whereas the substitution of Ile with Lys or Glu at P2 position reduced the cleavage efficiency by 30%. The activity of Trx-PfPMVm84 was inhibited by PMSF and nelfinavir but not by pepstatin A. After the removal of Trx domain, activities of both enzymes toward PfEMP2 and HRPII peptides were fitted to the Michaelis-Menten model to determine kinetic parameters. The Km values toward both peptides were apparently much lower than the previously reported data although with similar kcat values. Along with an improved PfPMV preparation protocol, these findings have provided insights into its substrate specificity at P2 and P1' positions as well as interactions among the enzyme, substrates, and inhibitors.

  20. TPM analyses reveal that FtsK contributes both to the assembly and the activation of the XerCD-dif recombination synapse.

    Science.gov (United States)

    Diagne, Cheikh Tidiane; Salhi, Maya; Crozat, Estelle; Salomé, Laurence; Cornet, Francois; Rousseau, Philippe; Tardin, Catherine

    2014-02-01

    Circular chromosomes can form dimers during replication and failure to resolve those into monomers prevents chromosome segregation, which leads to cell death. Dimer resolution is catalysed by a highly conserved site-specific recombination system, called XerCD-dif in Escherichia coli. Recombination is activated by the DNA translocase FtsK, which is associated with the division septum, and is thought to contribute to the assembly of the XerCD-dif synapse. In our study, direct observation of the assembly of the XerCD-dif synapse, which had previously eluded other methods, was made possible by the use of Tethered Particle Motion, a single molecule approach. We show that XerC, XerD and two dif sites suffice for the assembly of XerCD-dif synapses in absence of FtsK, but lead to inactive XerCD-dif synapses. We also show that the presence of the γ domain of FtsK increases the rate of synapse formation and convert them into active synapses where recombination occurs. Our results represent the first direct observation of the formation of the XerCD-dif recombination synapse and its activation by FtsK.

  1. Molecular cloning, expression of a galectin gene in Pacific white shrimp Litopenaeus vannamei and the antibacterial activity of its recombinant protein.

    Science.gov (United States)

    Cha, Gui-Hong; Liu, Yuan; Peng, Ting; Huang, Ming-Zhu; Xie, Chen-Ying; Xiao, Yu-Chao; Wang, Wei-Na

    2015-10-01

    Galectins play crucial roles in innate immune responses in invertebrate by recognizing and eliminating microinvaders. In this study, a cDNA encoding a galectin in the Pacific white shrimp (L. vannamei) was identified and characterized. A recombinant variant of this lectin, rLvgalectin, was expressed in the model organism P. pastoris and its expression was confirmed by Western blot. Biochemical assays indicated that the recombinant protein antibacterial rLvgalectin activity and was expressed in all of the organism's tested tissues Injection of the bacterium V. alginolyticus into L. vannamei induced hemocytes upregulation of Lvgalectin. The recombinant Lvgalectin protein (rLvgalectin) could bind various microorganism including Gram-positive bacteria, Gram-negative bacteria and yeast. And it revealed antimicrobial activity against the test Gram-positive bacteria, Gram-negative bacteria, but did not inhibit the growth of fungus Pichia pastoris. Moreover, rLvgalectin could significantly enhance the clearance activity of V. alginolyticus in vivo. In vivo challenge experiments showed that the recombinant rLvgalectin protein can significantly reduce the mortalities of V. alginolyticus injection. Furthermore, Compared to their wild-type counterparts, Lvgalectin-silenced shrimp exhibited increased mortality and hemocyte apoptosis, decreased bacterial clearance ability and total hemocyte counts, and stronger expression of Lvp53, LvproPO, LvPEN3, and LvCrustin following V. alginolyticus challenge. Taken together, these results suggest that galectin is important in the innate immune response of shrimp to pathogens infection.

  2. Biological insights into the expression of translation initiation factors from recombinant CHOK1SV cell lines and their relationship to enhanced productivity.

    Science.gov (United States)

    Mead, Emma J; Masterton, Rosalyn J; Feary, Marc; Obrezanova, Olga; Zhang, Lin; Young, Robert; Smales, C Mark

    2015-12-15

    Translation initiation is on the critical pathway for the production of monoclonal antibodies (mAbs) by mammalian cells. Formation of a closed loop structure comprised of mRNA, a number of eukaryotic initiation factors (eIFs) and ribosomal proteins has been proposed to aid re-initiation of translation and therefore increase global translational efficiency. We have determined mRNA and protein levels of the key components of the closed loop, eIFs (eIF3a, eIF3b, eIF3c, eIF3h, eIF3i and eIF4G1), poly(A)-binding protein (PABP) 1 and PABP-interacting protein 1 (PAIP1), across a panel of 30 recombinant mAb-producing GS-CHOK1SV cell lines with a broad range of growth characteristics and production levels of a model recombinant mAb. We have used a multi-level statistical approach to investigate the relationship between key performance indicators (cell growth and recombinant antibody productivity) and the intracellular amounts of target translation initiation factor proteins and the mRNAs encoding them. We show that high-producing cell lines maintain amounts of the translation initiation factors involved in the formation of the closed loop mRNA, maintaining these proteins at appropriate levels to deliver enhanced recombinant protein production. We then utilize knowledge of the amounts of these factors to build predictive models for and use cluster analysis to identify, high-producing cell lines. The present study therefore defines the translation initiation factor amounts that are associated with highly productive recombinant GS-CHOK1SV cell lines that may be targets for screening highly productive cell lines or to engineer new host cell lines with the potential for enhanced recombinant antibody productivity.

  3. BIOLOGICAL VALUE OF PUNY FRUITS RELATED TO THEIR ANTIRADICAL ACTIVITY

    Directory of Open Access Journals (Sweden)

    M. BALOGHOVÁ

    2013-12-01

    Full Text Available All analysed species of puny fruits (red currant (Ribes rubrum L variant Jonkheervan Tets, white currant (Ribes vulgare L. variant Blanka, black currant (Ribesnigrum L. variant Eva, blueberry (Vaccinium myrtilis variant Berkeley, elderberry(Sambucus nigra L. variant Sambo, hawthorn (Crataegus oxyacantha, mulberry(Morus nigra L. genotypes M152 and M047 are natural sources of anthocyanpigments and vitamin C with a high antiradical activity. Due to the fact that thehighest antiradical activity is not accompanied by the highest content of anthocyansand vitamin C in puny fruits, we suppose that the antiradical activity of plantmaterials is also connected with the presence of other compounds with antioxidantand antiradical activity. From our results follows that all studied puny fruits with ahigh antiradical activity increase the antioxidant value of human nutrition and alsoits prophylactic and medicinal effect.

  4. Phytochemical prospection and biological activity of Duroia macrophylla (Rubiaceae

    Directory of Open Access Journals (Sweden)

    Daiane Martins

    2014-12-01

    Full Text Available Context: Duroia macrophylla (Rubiaceae is endemic from the Amazon Rainforest. Aims: To perform phytochemical profile of Duroia macrophylla extracts and to evaluate them as antioxidant, insecticidal and cytotoxic. Methods: Dichloromethane and methanol extracts of leaves and branches (collected three times were subjected to phytochemical screening by comparative thin layer chromatography and NMR analyses. The extracts were assayed to antioxidant (DPPH and Fe-phenanthroline, at 10 μg/mL, insecticidal on Sitophilus zeamais (by ingestion of stored grains and contact, both at 10 mg/mL and toxic activities on Artemia salina (1000 μg/mL. Results: There were found evidences of terpenes, phenolic substances (phenols and flavonoids and alkaloids, with differences between the vegetal part, collection period and solvent used. Antioxidant evaluations showed three of twelve were active and two were considered moderately active, with a relationship dependently of concentration. All methanol extracts showed the presence of phenolic substances (phenols and flavonoids but one showed only phenols. For insecticidal activity, there were three most active extracts, two of which showed only presence of terpenes and the other, besides terpenes, phenolic substances (phenols and flavonoids. For Artemia salina toxicity assay, the five most active were all from the 2nd and 3rd collections. Conclusions: The active extracts of D. macrophylla in each test were different. Three methanol extracts showed antioxidant activity; three extracts showed insecticidal activity and the presence of terpenic substances and five extracts presented cytotoxic activity, but it was not possible to correlate it with any specific secondary metabolite.

  5. Tests of biological activity of metabolites from Penicillium expansum (Link Thom various isolates

    Directory of Open Access Journals (Sweden)

    Halina Borecka

    2013-12-01

    Full Text Available Aqrobacterium tumefaciens and cucumber, mustard and linseeds were compared as test organisms for evaluation of the biological activity of patulin. It was found that the reaction of cucumber seeds and linseed to the patulin concentrations was more pronounced than that of mustard and Aqrobacterium tumefaciens. The activity of metabolites produced by Penicillium expansum was investigated with the use of cucumber seeds. As measure of activity served the percentage of radicule growth inhibition was compared with the growth in control seeds. The biological activity of the metabolites was specific for the isolates, those from apples being more active. Thirty two isolates from pears and 34 from apples were examined.

  6. Activation of the cellular unfolded protein response by recombinant adeno-associated virus vectors.

    Directory of Open Access Journals (Sweden)

    Balaji Balakrishnan

    Full Text Available The unfolded protein response (UPR is a stress-induced cyto-protective mechanism elicited towards an influx of large amount of proteins in the endoplasmic reticulum (ER. In the present study, we evaluated if AAV manipulates the UPR pathways during its infection. We first examined the role of the three major UPR axes, namely, endoribonuclease inositol-requiring enzyme-1 (IRE1α, activating transcription factor 6 (ATF6 and PKR-like ER kinase (PERK in AAV infected cells. Total RNA from mock or AAV infected HeLa cells were used to determine the levels of 8 different ER-stress responsive transcripts from these pathways. We observed a significant up-regulation of IRE1α (up to 11 fold and PERK (up to 8 fold genes 12-48 hours after infection with self-complementary (scAAV2 but less prominent with single-stranded (ssAAV2 vectors. Further studies demonstrated that scAAV1 and scAAV6 also induce cellular UPR in vitro, with AAV1 vectors activating the PERK pathway (3 fold while AAV6 vectors induced a significant increase on all the three major UPR pathways [6-16 fold]. These data suggest that the type and strength of UPR activation is dependent on the viral capsid. We then examined if transient inhibition of UPR pathways by RNA interference has an effect on AAV transduction. siRNA mediated silencing of PERK and IRE1α had a modest effect on AAV2 and AAV6 mediated gene expression (∼1.5-2 fold in vitro. Furthermore, hepatic gene transfer of scAAV2 vectors in vivo, strongly elevated IRE1α and PERK pathways (2 and 3.5 fold, respectively. However, when animals were pre-treated with a pharmacological UPR inhibitor (metformin during scAAV2 gene transfer, the UPR signalling and its subsequent inflammatory response was attenuated concomitant to a modest 2.8 fold increase in transgene expression. Collectively, these data suggest that AAV vectors activate the cellular UPR pathways and their selective inhibition may be beneficial during AAV mediated gene transfer.

  7. Human recombinant anti-thyroperoxidase autoantibodies: in vitro cytotoxic activity on papillary thyroid cancer expressing TPO

    OpenAIRE

    Rebuffat, S A; Morin, M.; Nguyen, B; Castex, F; Robert, B.; Péraldi-Roux, S

    2010-01-01

    Background: Thyroid cancers are difficult to treat due to their limited responsiveness to chemo- and radiotherapy. There is thus a great interest in and a need for alternative therapeutic approaches. Results: We studied the cytotoxic activity of anti-thyroperoxidase autoantibodies (anti-TPO aAbs, expressed in baculovirus/insect cell (B4) and CHO cells (B4′) or purified from patients' sera) against a papillary thyroid cancer (NPA) cell line. Anti-TPO aAbs from patients' sera led to a partial d...

  8. Synthesis and Antiplatelet Activity of Antithrombotic Thiourea Compounds: Biological and Structure-Activity Relationship Studies

    Directory of Open Access Journals (Sweden)

    André Luiz Lourenço

    2015-04-01

    Full Text Available The incidence of hematological disorders has increased steadily in Western countries despite the advances in drug development. The high expression of the multi-resistance protein 4 in patients with transitory aspirin resistance, points to the importance of finding new molecules, including those that are not affected by these proteins. In this work, we describe the synthesis and biological evaluation of a series of N,N'-disubstituted thioureas derivatives using in vitro and in silico approaches. New designed compounds inhibit the arachidonic acid pathway in human platelets. The most active thioureas (compounds 3d, 3i, 3m and 3p displayed IC50 values ranging from 29 to 84 µM with direct influence over in vitro PGE2 and TXA2 formation. In silico evaluation of these compounds suggests that direct blockage of the tyrosyl-radical at the COX-1 active site is achieved by strong hydrophobic contacts as well as electrostatic interactions. A low toxicity profile of this series was observed through hemolytic, genotoxic and mutagenic assays. The most active thioureas were able to reduce both PGE2 and TXB2 production in human platelets, suggesting a direct inhibition of COX-1. These results reinforce their promising profile as lead antiplatelet agents for further in vivo experimental investigations.

  9. In plant activation: an inducible, hyperexpression platform for recombinant protein production in plants.

    Science.gov (United States)

    Dugdale, Benjamin; Mortimer, Cara L; Kato, Maiko; James, Tess A; Harding, Robert M; Dale, James L

    2013-07-01

    In this study, we describe a novel protein production platform that provides both activation and amplification of transgene expression in planta. The In Plant Activation (INPACT) system is based on the replication machinery of tobacco yellow dwarf mastrevirus (TYDV) and is essentially transient gene expression from a stably transformed plant, thus combining the advantages of both means of expression. The INPACT cassette is uniquely arranged such that the gene of interest is split and only reconstituted in the presence of the TYDV-encoded Rep/RepA proteins. Rep/RepA expression is placed under the control of the AlcA:AlcR gene switch, which is responsive to trace levels of ethanol. Transgenic tobacco (Nicotiana tabacum cv Samsun) plants containing an INPACT cassette encoding the β-glucuronidase (GUS) reporter had negligible background expression but accumulated very high GUS levels (up to 10% total soluble protein) throughout the plant, within 3 d of a 1% ethanol application. The GUS reporter was replaced with a gene encoding a lethal ribonuclease, barnase, demonstrating that the INPACT system provides exquisite control of transgene expression and can be adapted to potentially toxic or inhibitory compounds. The INPACT gene expression platform is scalable, not host-limited, and has been used to express both a therapeutic and an industrial protein.

  10. Antigenicity and Hemaglutination Activity of a Recombinant Hemagglutinin-Neuraminidase of Paramyxovirus Tianjin Strain

    Institute of Scientific and Technical Information of China (English)

    Mei LI; Li-jun YUAN; Li-ying SHI; Xiao-mian LI; Qing WANG; Wen-xiu WANG

    2008-01-01

    Paramyxovirus Tianjin strain, a new genotype of Sendal virus, was isolated from the lungs of common cotton-eared marmoset that died of severe respiratory infection in the marmoset colonies. The 19.28% IgM positive rate in the young children with acute respiratory tract infection suggested a close relationship between Tianjin strain and humans. Hemagglutinin-neuraminidase (HN) is its major transmembrane glycoprotein responsible for viral attachment, penetration and release. To clear the relationship between HN structure and function of paramyxovirus Tianjin strain, rHN1, rHN2 and rHN3 overlapping the ectodomain of HN protein were expressed. Their antigenicity and hemaglutination activity, as well as cross reactivity to standard antisera against influenza virus type A, type B were analyzed. The results indicated expressed rHNs have the natural antigenicity.The segment rHN2 possesses more linear epitopes exposed on the surface of the native I-IN protein than found in segments rHN3 and rHN1. The hemagglutination activity of segment rHN3 is higher than that of segments rHN2 and rHN1, and partially dependent on the three-dimensional conformation of HN3 protein. Cross-reactivity between rHNs and standard antisera against influenza virus type A, type B suggested that rHNs might not be the best alternative as specific antigens to detect virus in clinicalserum specimens.

  11. Capturing Biological Activity in Natural Product Fragments by Chemical Synthesis.

    Science.gov (United States)

    Crane, Erika A; Gademann, Karl

    2016-03-14

    Natural products have had an immense influence on science and have directly led to the introduction of many drugs. Organic chemistry, and its unique ability to tailor natural products through synthesis, provides an extraordinary approach to unlock the full potential of natural products. In this Review, an approach based on natural product derived fragments is presented that can successfully address some of the current challenges in drug discovery. These fragments often display significantly reduced molecular weights, reduced structural complexity, a reduced number of synthetic steps, while retaining or even improving key biological parameters such as potency or selectivity. Examples from various stages of the drug development process up to the clinic are presented. In addition, this process can be leveraged by recent developments such as genome mining, antibody-drug conjugates, and computational approaches. All these concepts have the potential to identify the next generation of drug candidates inspired by natural products.

  12. Chemical Constituents of Descurainia sophia L. and its Biological Activity

    Directory of Open Access Journals (Sweden)

    Nawal H. Mohamed

    2009-01-01

    Full Text Available Seven coumarin compounds were isolated for the first time from the aerial parts of DescurainiaSophia L. identified as scopoletine, scopoline, isoscopoline, xanthtoxol, xanthtoxin, psoralene and bergaptane.Three flavonoids namely kaempferol, quercetine and isorhamnetine and three terpenoid compounds -sitosterol-amyrine and cholesterol were also isolated and identified by physical and chemical methods; melting point, Rfvalues, UV and 1H NMR spectroscopy. Qualitative and quantitative analyses of free and protein amino acidsusing amino acid analyzer were performed. The plant contains 15 amino acids as free and protein amino acidswith different range of concentrations. Fatty acid analysis using GLC, revealed the presence of 10 fatty acids,the highest percentage was palmitic acid (27.45 % and the lowest was lauric acid (0.13%. Biological screeningof alcoholic extract showed that the plant is highly safe and has analgesic, antipyretic and anti-inflammatoryeffects.

  13. Chemical Constituents and Biological Activities of Artemisia herba-alba

    Directory of Open Access Journals (Sweden)

    Abou El-Hamd H. Mohamed

    2010-01-01

    Full Text Available Artemisia, one of the larger genera in the family Asteraceae and the largest genus in the tribe Anthemideae, comprises from 200 to more than 500 taxa at the specific or subspecific level. Many Artemisia species have a high economic value in several fields, as food plants and as antihelminthic and antimalaria in medicine. Artemisia herba-alba was known for its therapeutic and medicinal properties, it was used in both traditional and modern medicine. Several papers have been published on the chemical composition of specimens of A. herba-alba. The aim of this work is to review all available scientific literature published on A. herba-alba. The focus will be on the chemical constitutions which have been identified from this species, in addition to all of the reported biological activites of this species have been included as well as the pharmacology and toxicology

  14. Isolation, biological activity, synthesis, and medicinal chemistry of the pederin/mycalamide family of natural products.

    Science.gov (United States)

    Mosey, R Adam; Floreancig, Paul E

    2012-09-01

    This review highlights the broad range of science that has arisen from the isolation of pederin, the mycalamides, theopederins, and onnamides, and psymberin. Specific topics include structure determination, biological activity, synthesis, and analog preparation and analysis.

  15. Environmental Contaminants Monitoring in Selected Wetlands of Wyoming: Biologically Active Elements Study

    Data.gov (United States)

    US Fish and Wildlife Service, Department of the Interior — Sediment, water and biota were collected from selected wetlands in Wyoming for the Biologically Active Elements (BAE) Study in 1988, 1989 and 1990 to identify...

  16. Bioactive Components and Functional Properties of Biologically Activated Cereal Grains: A Bibliographic Review.

    Science.gov (United States)

    Singh, Arashdeep; Sharma, Savita

    2015-10-14

    Whole grains provide energy, nutrients, fibres and bioactive compounds that may synergistically contribute to their protective effects. A wide range of these compounds is affected by germination. While some compounds, such as β-glucans are degraded, others, like antioxidants and total phenolics are increased by means of biological activation of grains. The water and oil absorption capacity as well as emulsion and foaming capacity of biologically activated grains are also improved. Application of biological activation of grains is of emerging interest, which may significantly enhance the nutritional, functional and bioactive content of grains, as well as improve palatability of grain foods in a natural way. Therefore, biological activation of cereals can be a way to produce food grains enriched with health promoting compounds and enhanced functional attributes.

  17. Therapeutic Uses and Pharmacological Properties of Garlic, Shallot, and Their Biologically Active Compounds

    Directory of Open Access Journals (Sweden)

    Peyman Mikaili

    2013-10-01

    Garlic and shallots are safe and rich sources of biologically active compounds with low toxicity. Further studies are needed to confirm the safety and quality of the plants to be used by clinicians as therapeutic agents.

  18. Biological activity of fucoidan from leafy bladderwort (Utricularia aurea Lour.

    Directory of Open Access Journals (Sweden)

    Chotigeat, W.

    2005-12-01

    Full Text Available Fucoidan extracted from aquatic plant Utricularia aurea was 1.3% of dry weight and comprised glucuronic acid 62.5% and fucose 4.98% of the crude extract and 28.74% of the fucose was sulfate. The fucoidan was investigated for antibacterial activity. The minimal inhibitory concentration (MIC of crude fucoidan against Vibrio harveyi and Escherichia coli was 20 and 10 mg/ml. Moreover, the fucoidan had anticoagulant activity. The crude fucoidan had activated partial thrombo