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Sample records for lys-274 arg mutant

  1. Uv-visible spectroscopy of bacteriorhodopsin mutants: substitution of Arg-82, Asp-85, Tyr-185, and Asp-212 results in abnormal light-dark adaptation.

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    Duñach, M; Marti, T; Khorana, H G; Rothschild, K J

    1990-12-01

    The light-dark adaptation reactions of a set of bacteriorhodopsin (bR) mutants that affect function and color of the chromophore were examined by using visible absorption spectroscopy. The absorbance spectra of the mutants Arg-82 in equilibrium Ala (Gln), Asp-85 in equilibrium Ala (Asn, Glu), Tyr-185 in equilibrium Phe, and Asp-212 in equilibrium Ala (Asn, Glu) were measured at different pH values during and after illumination. None of these mutants exhibited a normal dark-light adaptation, which in wild-type bR causes a red shift of the visible absorption maximum from 558 nm (dark-adapted bR) to 568 nm (light-adapted bR). Instead a reversible light reaction occurs in the Asp-85 and Asp-212 mutants from a blue form with lambda max near 600 nm to a pink form with lambda max near 480 nm. This light-induced shift explains the appearance of a reversed light adaptation previously observed for the Asp-212 mutants. In the case of the Tyr-185 and Arg-82 mutants, light causes a purple-to-blue transformation similar to the effect of lowering the pH. However, the blue forms observed in these mutants are not identical to those formed by acid titration or deionization of wild-type bR. It is suggested that in all of these mutants, the chromophore has lost the ability to undergo the normal 13-cis, 15-syn to all-trans, 15-anti light-driven isomerization, which occurs in native bR. Instead these mutants may have as stable forms all-trans,syn and 13-cis,anti chromophores, which are not allowed in native bR, except transiently.

  2. Roles of distal arginine in activity and stability of Coprinus cinereus peroxidase elucidated by kinetic and NMR analysis of the Arg51Gln, -Asn, -Leu, and -Lys mutants.

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    Schiødt, Christine B; Veitch, Nigel C; Welinder, Karen G

    2007-02-01

    In heme peroxidases, a distal His residue plays an essential role in the initial two electron oxidation of resting state enzyme to compound I by hydrogen peroxide. A distal Arg residue assists in this process. The contributions of the charge, H-bonding capacity, size, and mobility of this Arg residue to Coprinus cinereus peroxidase (CIP) reactivity and stability have been examined by substituting Arg51 with Gln (retains H-bond donor at N epsilon position), Asn (small size, H-bond donor and acceptor), Leu (similar to Asn, but hydrophobic), and Lys (charge and H-bond donor, but at N zeta position). UV-visible spectroscopy was used to monitor pH-linked heme changes, compound I formation and reduction, fluoride binding, and thermostability. (1)H NMR spectroscopy enabled heme pocket differences in both resting and cyanide-ligated states of the enzymes to be evaluated and compared with wild-type CIP. We found that the H-bonding capacity of distal Arg is key to fast compound I formation and ligand binding to heme, whereas charge is important for lowering the pK(a) of distal His and for the binding and stabilisation of anionic ligands at heme iron. The properties of the distal Arg residue in CIP, cytochrome c peroxidase (CCP) and horseradish peroxidase (HRP) differ significantly in their pH induced transitions and dynamics.

  3. Role of αArg145 and βArg263 in the active site of penicillin acylase of Escherichia coli

    NARCIS (Netherlands)

    Alkema, Wynand B.L.; Prins, Antoon K.; de Vries, Erik; Janssen, Dick B.

    2002-01-01

    The active site of penicillin acylase of Eschcrichia coli contains two conserved arginine residues, The function of these arainines, alphaArg(145) and betaArg(263), was studied by site-directed mutagenesis and kinetic analysis of the mutant enzymes. The mutants alphaArg(145)-->Leu (alphaArg145Leu),

  4. Regulation of the histone acetyltransferase activity of hMOF via autoacetylation of Lys274

    Institute of Scientific and Technical Information of China (English)

    Bingfa Sun; Shunling Guo; Qingyu Tang; Chen Li; Rong Zeng; Zhiqi Xiong; Chen Zhong; Jianping Ding

    2011-01-01

    Dear Editor, Males-absent-on-the-first (MOF, also called MYST1 or KAT8) is a histone acetyltransferase (HAT) belonging to the MOZ, Ybf2/Sas3, Sas2 and Tip60 (MYST) family.MOF has been shown to possess a specific HAT activity towards Lysl6 of histone H4 (H4K16) [1].Homozygous knockout of MOF in mice results in loss of H4K16 acetylation and embryonic lethality, indicating that MOF and H4K16 acetylation are essential for embryogenesis and genome stability in mammals [2].Downregulation of human MOF (hMOF) leads to dramatic nuclear morphological deformation and inhibition of cell cycle progression [3], and has recently been correlated with primary breast carcinoma and medulloblastoma [4].

  5. Stationary phase expression of the arginine biosynthetic operon argCBH in Escherichia coli

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    Sun Yuan

    2006-02-01

    Full Text Available Abstract Background Arginine biosynthesis in Escherichia coli is elevated in response to nutrient limitation, stress or arginine restriction. Though control of the pathway in response to arginine limitation is largely modulated by the ArgR repressor, other factors may be involved in increased stationary phase and stress expression. Results In this study, we report that expression of the argCBH operon is induced in stationary phase cultures and is reduced in strains possessing a mutation in rpoS, which encodes an alternative sigma factor. Using strains carrying defined argR, and rpoS mutations, we evaluated the relative contributions of these two regulators to the expression of argH using operon-lacZ fusions. While ArgR was the main factor responsible for modulating expression of argCBH, RpoS was also required for full expression of this biosynthetic operon at low arginine concentrations (below 60 μM L-arginine, a level at which growth of an arginine auxotroph was limited by arginine. When the argCBH operon was fully de-repressed (arginine limited, levels of expression were only one third of those observed in ΔargR mutants, indicating that the argCBH operon is partially repressed by ArgR even in the absence of arginine. In addition, argCBH expression was 30-fold higher in ΔargR mutants relative to levels found in wild type, fully-repressed strains, and this expression was independent of RpoS. Conclusion The results of this study indicate that both derepression and positive control by RpoS are required for full control of arginine biosynthesis in stationary phase cultures of E. coli.

  6. Mutation of the Erwinia amylovora argD gene causes arginine auxotrophy, nonpathogenicity in apples, and reduced virulence in pears.

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    Ramos, Laura S; Lehman, Brian L; Peter, Kari A; McNellis, Timothy W

    2014-11-01

    Fire blight is caused by Erwinia amylovora and is the most destructive bacterial disease of apples and pears worldwide. In this study, we found that E. amylovora argD(1000)::Tn5, an argD Tn5 transposon mutant that has the Tn5 transposon inserted after nucleotide 999 in the argD gene-coding region, was an arginine auxotroph that did not cause fire blight in apple and had reduced virulence in immature pear fruits. The E. amylovora argD gene encodes a predicted N-acetylornithine aminotransferase enzyme, which is involved in the production of the amino acid arginine. A plasmid-borne copy of the wild-type argD gene complemented both the nonpathogenic and the arginine auxotrophic phenotypes of the argD(1000)::Tn5 mutant. However, even when mixed with virulent E. amylovora cells and inoculated onto immature apple fruit, the argD(1000)::Tn5 mutant still failed to grow, while the virulent strain grew and caused disease. Furthermore, the pCR2.1-argD complementation plasmid was stably maintained in the argD(1000)::Tn5 mutant growing in host tissues without any antibiotic selection. Therefore, the pCR2.1-argD complementation plasmid could be useful for the expression of genes, markers, and reporters in E. amylovora growing in planta, without concern about losing the plasmid over time. The ArgD protein cannot be considered an E. amylovora virulence factor because the argD(1000)::Tn5 mutant was auxotrophic and had a primary metabolism defect. Nevertheless, these results are informative about the parasitic nature of the fire blight disease interaction, since they indicate that E. amylovora cannot obtain sufficient arginine from apple and pear fruit tissues or from apple vegetative tissues, either at the beginning of the infection process or after the infection has progressed to an advanced state.

  7. Expression and Characterization of ArgR, An Arginine Regulatory Protein in Corynebacterium crenatum

    Institute of Scientific and Technical Information of China (English)

    CHEN Xue Lan; ZHANG Bin; TANG Li; JIAO Hai Tao; XU Heng Yi; XU Feng; XU Hong; WEI Hua; XIONG Yong Hua

    2014-01-01

    Objective Corynebacterium crenatum MT, a mutant from C. crenatum AS 1.542 with a lethal argR gene, exhibits high arginine production. To confirm the effect of ArgR on arginine biosynthesis in C. crenatum, an intact argR gene from wild-type AS 1.542 was introduced into C. crenatum MT, resulting in C. crenatum MT. sp, and the changes of transcriptional levels of the arginine biosynthetic genes and arginine production were compared between the mutant strain and the recombinant strain. Methods Quantitative real-time polymerase chain reaction was employed to analyze the changes of the related genes at the transcriptional level, electrophoretic mobility shift assays were used to determine ArgR binding with the argCJBDF, argGH, and carAB promoter regions, and arginine production was determined with an automated amino acid analyzer. Results Arginine production assays showed a 69.9%reduction in arginine from 9.01±0.22 mg/mL in C. crenatum MT to 2.71±0.13 mg/mL (P Conclusion The arginine biosynthetic genes in C. crenatum are clearly controlled by the negative regulator ArgR, and intact ArgR in C. crenatum MT results in a significant descrease in arginine production.

  8. ArgR-dependent repression of arginine and histidine transport genes in Escherichia coli K-12.

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    Caldara, Marina; Minh, Phu Nguyen Le; Bostoen, Sophie; Massant, Jan; Charlier, Daniel

    2007-10-19

    In Escherichia coli L-arginine is taken up by three periplasmic binding protein-dependent transport systems that are encoded by two genetic loci: the artPIQM-artJ and argT-hisJQMP gene clusters. The transcription of the artJ, artPIQM and hisJQMP genes and operons is repressed by liganded ArgR, whereas argT, encoding the LAO (lysine, arginine, ornithine) periplasmic binding protein, is insensitive to the repressor. Here we characterize the repressible Esigma70 P artJ, P artP and P hisJ promoters and demonstrate that the cognate operators consist of two 18 bp ARG boxes separated by 3 bp. Determination of the energy landscape of the ArgR-operator contacts by missing contact probing and mutant studies indicated that each box of a pair contributes to complex formation in vitro and to the repressibility in vivo, but to a different extent. The organization of the ARG boxes and promoter elements in the control regions of the uptake genes is distinct from that of the arginine biosynthetic genes. The hisJQMP operon is the first member of the E. coli ArgR regulon, directly repressed by liganded ArgR, where none of the core promoter elements overlaps the ARG boxes. Single round in vitro transcription assays and DNase I footprinting experiments indicate that liganded ArgR inhibits P artJ and P artP promoter activity by steric exclusion of the RNA polymerase. In contrast, ArgR-mediated repression of P hisJ by inhibition of RNA polymerase binding appears to occur through topological changes of the promoter region.

  9. Neutron crystallography of photoactive yellow protein reveals unusual protonation state of Arg52 in the crystal.

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    Yonezawa, Kento; Shimizu, Nobutaka; Kurihara, Kazuo; Yamazaki, Yoichi; Kamikubo, Hironari; Kataoka, Mikio

    2017-08-24

    Because of its high pKa, arginine (Arg) is believed to be protonated even in the hydrophobic environment of the protein interior. However, our neutron crystallographic structure of photoactive yellow protein, a light sensor, demonstrated that Arg52 adopts an electrically neutral form. We also showed that the hydrogen bond between the chromophore and Glu46 is a so-called low barrier hydrogen bond (LBHB). Because both the neutral Arg and LBHB are unusual in proteins, these observations remain controversial. To validate our findings, we carried out neutron crystallographic analysis of the E46Q mutant of PYP. The resultant structure revealed that the proportion of the cationic form is higher in E46Q than in WT, although the cationic and neutral forms of Arg52 coexist in E46Q. These observations were confirmed by the occupancy of the deuterium atom bound to the N η1 atom combined with an alternative conformation of the N(η2)D2 group comprising sp(2) hybridisation. Based on these results, we propose that the formation of the LBHB decreases the proton affinity of Arg52, stabilizing the neutral form in the crystal.

  10. Comparative stability of dihydrofolate reductase mutants in vitro and in vivo.

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    Leontiev, V V; Uversky, V N; Gudkov, A T

    1993-01-01

    Dihydrofolate reductase mutants with amino acid replacements in the active center (Thr35-->Asp mutant, Arg57-->His mutant and the mutant with triple replacement Thr35-->Asp, Asn37-->Ser, Arg57-->His) were obtained by site-directed mutagenesis. The stabilization effect of trimethoprim and NADP.H on the protein tertiary structure in vitro has been investigated. In the case of mutants with a 'weak' tertiary structure (Thr35-->Asp35 and the triple mutant) the separate addition of ligands does not affect their stability. The simultaneous addition of these ligands to Thr35-->Asp35 and the triple mutant leads to the large increase in their stability. A distinct correlation was found between the in vitro studied stability of the mutant proteins to the urea- or heat-induced denaturation and the level of proteolytic degradation of these mutants previously observed in vivo.

  11. Chemical rescue and inhibition studies to determine the role of Arg301 in phosphite dehydrogenase.

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    John E Hung

    Full Text Available Phosphite dehydrogenase (PTDH catalyzes the NAD(+-dependent oxidation of phosphite to phosphate. This reaction requires the deprotonation of a water nucleophile for attack on phosphite. A crystal structure was recently solved that identified Arg301 as a potential base given its proximity and orientation to the substrates and a water molecule within the active site. Mutants of this residue showed its importance for efficient catalysis, with about a 100-fold loss in k cat and substantially increased K m,phosphite for the Ala mutant (R301A. The 2.35 Å resolution crystal structure of the R301A mutant with NAD(+ bound shows that removal of the guanidine group renders the active site solvent exposed, suggesting the possibility of chemical rescue of activity. We show that the catalytic activity of this mutant is restored to near wild-type levels by the addition of exogenous guanidinium analogues; Brønsted analysis of the rates of chemical rescue suggests that protonation of the rescue reagent is complete in the transition state of the rate-limiting step. Kinetic isotope effects on the reaction in the presence of rescue agents show that hydride transfer remains at least partially rate-limiting, and inhibition experiments show that K i of sulfite with R301A is ∼400-fold increased compared to the parent enzyme, similar to the increase in K m for phosphite in this mutant. The results of our experiments indicate that Arg301 plays an important role in phosphite binding as well as catalysis, but that it is not likely to act as an active site base.

  12. Fmoc-Arg(Pbf)-OH的合成%The synthesis of Fmoc-Arg(Pbf)-OH

    Institute of Scientific and Technical Information of China (English)

    洪镛裕; 刘超程; 赵宏伟

    2006-01-01

    研究了精氨酸用9-芴甲氧羰基(Fmoc)和2,2,4,6,7-五甲基苯并呋喃-5-磺酰基(Pbf)双保护的Fmoc-Arg(Pbf)-OH的制备过程.在引入Pbf时,加入了相转移催化剂四乙基溴化铵(TEBA),提高了反应效率,减少了Pbf-Cl的水解;成盐时改为将Cbz-Arg(Pbf)-OH溶液加入到环己胺溶液中,使成盐质量有所提高,将该步收率由文献的59%提高到72%.

  13. Occurrence and temporal variation of antibiotic resistance genes (ARGs) in shrimp aquaculture: ARGs dissemination from farming source to reared organisms.

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    Su, Haochang; Liu, Shan; Hu, Xiaojuan; Xu, Xiangrong; Xu, Wujie; Xu, Yu; Li, Zhuojia; Wen, Guoliang; Liu, Yousheng; Cao, Yucheng

    2017-12-31

    Considerable attention has been paid to the occurrence and abundance of antibiotic resistance genes (ARGs) in aquatic environments. However, the temporal variation and dissemination of ARGs in aquaculture environments and reared organisms need further study. This study investigated the abundance and diversity of ARGs and bacterial community in water source, shrimp pond water, sediment, and shrimps during the rearing period in Pearl River Delta region, South China. The results showed that sul1, qnrD, cmlA, and floR were the predominant ARGs in the aquaculture samples. A trend of decreasing abundance of ARGs was observed for pond water samples during the rearing period, whereas an increasing trend was observed in the sediment and shrimp samples. The total concentration of ARGs in water source was significantly higher than that in shrimp pond water (pshrimps were 4.48-19.0 times higher than those in juvenile shrimps. Similar to water source and pond water, cmlA and sul1 were the predominant ARGs in shrimp intestinal tract. The bacterial community in the shrimp intestinal tract changed greatly from juvenile to adult. The results of the present study indicated that the abundances of ARGs in aquaculture varied temporally during the rearing period. Water source was an important medium disseminating ARGs to the aquaculture environments and reared organisms. Sul1 could be used as a potential indicator for ARGs in both water and sediment in aquaculture in the estuary of the Pearl River Delta, South China. This study represents a case study for the temporal variation of abundance and dissemination of ARGs in aquaculture and is a reference for potential risks to food safety and human health. Copyright © 2017 Elsevier B.V. All rights reserved.

  14. Functional characterizations of residues Arg-158 and Tyr-170 of the mosquito-larvicidal Bacillus thuringiensis Cry4Ba

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    Leetachewa, Somphob; Moonsom, Saengduen; Chaisri, Urai; Khomkhum, Narumol; Yoonim, Nonglak; Wang, Ping; Angsuthanasombat, Chanan

    2014-01-01

    The insecticidal activity of Bacillus thuringiensis (Bt) Cry toxins involves toxin stabilization, oligomerization, passage across the peritrophic membrane (PM), binding to midgut receptors and pore-formation. The residues Arg-158 and Tyr-170 have been shown to be crucial for the toxicity of Bt Cry4Ba. We characterized the biological function of these residues. In mosquito larvae, the mutants R158A/E/Q (R158) could hardly penetrate the PM due to a significantly reduced ability to alter PM permeability; the mutant Y170A, however, could pass through the PM, but degraded in the space between the PM and the midgut epithelium. Further characterization by oligomerization demonstrated that Arg-158 mutants failed to form correctly sized high-molecular weight oligomers. This is the first report that Arg-158 plays a role in the formation of Cry4Ba oligomers, which are essential for toxin passage across the PM. Tyr-170, meanwhile, is involved in toxin stabilization in the toxic mechanism of Cry4Ba in mosquito larvae. [BMB Reports 2014; 47(10): 546-551] PMID:24286331

  15. Mutagenesis of residue betaArg-246 in the phosphate-binding subdomain of catalytic sites of Escherichia coli F1-ATPase.

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    Ahmad, Zulfiqar; Senior, Alan E

    2004-07-23

    Residues responsible for phosphate binding in F(1)F(0)-ATP synthase catalytic sites are of significant interest because phosphate binding is believed linked to proton gradient-driven subunit rotation. From x-ray structures, a phosphate-binding subdomain is evident in catalytic sites, with conserved betaArg-246 in a suitable position to bind phosphate. Mutations betaR246Q, betaR246K, and betaR246A in Escherichia coli were found to impair oxidative phosphorylation and to reduce ATPase activity of purified F(1) by 100-fold. In contrast to wild type, ATPase of mutants was not inhibited by MgADP-fluoroaluminate or MgADP-fluoroscandium, showing the Arg side chain is required for wild-type transition state formation. Whereas 7-chloro-4-nitrobenzo-2-oxa-1,3-diazole (NBD-Cl) inhibited wild-type ATPase essentially completely, ATPase in mutants was inhibited maximally by approximately 50%, although reaction still occurred at residue betaTyr-297, proximal to betaArg-246 in the phosphate-binding pocket. Inhibition characteristics supported the conclusion that NBD-Cl reacts in betaE (empty) catalytic sites, as shown previously by x-ray structure analysis. Phosphate protected against NBD-Cl inhibition in wild type but not in mutants. The results show that phosphate can bind in the betaE catalytic site of E. coli F(1) and that betaArg-246 is an important phosphate-binding residue.

  16. Rubber Tree (Hevea brasiliensis Muell. Arg).

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    Venkatachalam, Perumal; Jayashree, Radha; Rekha, Karumamkandathil; Sushmakumari, Sreedharannair; Sobha, Sankaren; Kumari Jayasree, Parukkuttyamma; Kala, Radha Gopikkuttanunithan; Thulaseedharan, Arjunan

    2006-01-01

    Rubber tree (Hevea brasiliensis Muell. Arg.) is an important industrial crop for natural rubber production. At present, more than 9.5 million hectares in about 40 countries are devoted to rubber tree cultivation with a production about 6.5 million tons of dry rubber each year. The world supply of natural rubber is barely keeping up with a global demand for 12 million tons of natural rubber in 2020. Tapping panel dryness (TPD) is a complex physiological syndrome widely found in rubber tree plantations, which causes severe yield and crop losses in natural rubber producing countries. Currently, there is no effective prevention or treatment for this serious malady. As it is a perennial tree crop, the integration of specific desired traits through conventional breeding is both time-consuming and labour-intensive. Genetic transformation with conventional breeding is certainly a more promising tool for incorporation of agronomically important genes that could improve existing Hevea genotype. This chapter provides an Agrobacterium-mediated transformation protocol for rubber tree using immature anther-derived calli as initial explants. We have applied this protocol to generate genetically engineered plants from a high yielding Indian clone RRII 105 of Hevea brasiliensis (Hb). Calli were co-cultured with Agrobacterium tumefaciens harboring a plasmid vector containing the Hb superoxide dismutase (SOD) gene and the reporter gene used was beta-glucuronidase (GUS) gene (uidA). The selectable marker gene used was neomycin phosphotransferase (nptII) and kanamycin was used as selection agent. We found that a suitable transformation protocol for Hevea consists of a 3-d co-cultivation with Agrobacterium in the presence of 20 mM acetosyringone, 15 mM betaine HCl, and 11.55 mM proline followed by selection on medium containing 300 mg/L kanamycin. Transformed calli surviving on medium containing 300 mg/L kanamycin showed a strong GUS-positive reaction. Upon subsequent subculture into

  17. The Polymorphisms of Ser49Gly and Gly389Arg in Beta-1-Adrenergic Receptor Gene in Major Depression

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    KOKUT, Süleyman; ATAY, İnci Meltem; UZ, Efkan; AKPINAR, Abdullah; DEMİRDAŞ, Arif

    2015-01-01

    Introduction It was reported that the genetic susceptibility of major depressive disorder (MDD) is related with genetic polymorphisms. The aim of this study was to investigate the possible association of the genotype and allele frequencies of Ser49Gly and Arg389Gly polymorphisms in MDD by comparing them with healthy subjects. Methods A total of 144 patients with MDD diagnosed according to Diagnostic and Statistical Manual of Mental Disorders, 4th Edition (DSM-IV) criteria and 105 healthy controls were included in the study. Polymerase chain reaction (PCR) with restriction fragment length polymorphism (RFLP) was used for genotyping. Results Of the 144 participants in the MDD group, 77 (53.5%) had homozygous wild type (AA), 57 (39.6%) had heterozygous type (AG), and 10 (6.9%) had mutant (GG) genotype for Ser49Gly, whereas 75 (52.1%) had homozygous wild type (GG), 59 (41.0%) had heterozygous (GC) type, and 10 (6.9%) had mutant homozygous (CC) genotype for Gly386Arg. There were no significant difference in the allele and genotype frequencies of the beta-1-adrenergic receptor (ADRB1) gene for Ser49Gly and Arg389Gly polymorphisms after comparing with healthy controls (p=0.626; p=0.863 and p=0.625; p=0.914). Conclusion The results of our study did not reveal a major effect of the polymorphism of Ser49Gly and Gly389Arg in the ADRB1 gene in MDD. Further studies with larger sample size are required to elucidate the role of other beta-1 adrenergic gene polymorphisms in MDD.

  18. A novel pro-Arg motif recognized by WW domains.

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    Bedford, M T; Sarbassova, D; Xu, J; Leder, P; Yaffe, M B

    2000-04-07

    WW domains mediate protein-protein interactions through binding to short proline-rich sequences. Two distinct sequence motifs, PPXY and PPLP, are recognized by different classes of WW domains, and another class binds to phospho-Ser-Pro sequences. We now describe a novel Pro-Arg sequence motif recognized by a different class of WW domains using data from oriented peptide library screening, expression cloning, and in vitro binding experiments. The prototype member of this group is the WW domain of formin-binding protein 30 (FBP30), a p53-regulated molecule whose WW domains bind to Pro-Arg-rich cellular proteins. This new Pro-Arg sequence motif re-classifies the organization of WW domains based on ligand specificity, and the Pro-Arg class now includes the WW domains of FBP21 and FE65. A structural model is presented which rationalizes the distinct motifs selected by the WW domains of YAP, Pin1, and FBP30. The Pro-Arg motif identified for WW domains often overlaps with SH3 domain motifs within protein sequences, suggesting that the same extended proline-rich sequence could form discrete SH3 or WW domain complexes to transduce distinct cellular signals.

  19. Mutation of Arg154 to Gly154 in urokinase augments its fibrin-specificity.

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    Peng, G; Ma, Z; Kuai, L; Zhu, D

    1997-04-01

    Rscu-PA and its mutant constructed by in vitro site specific mutagenesis of Arg154 in rscu-PA to Gly154 (mscu-PA) were both expressed in Escherichia coli. After in vitro denaturation and renaturation, the rscu-PA and mscu-PA were purified to homogeneity by Zn2+ selective precipitation, anti-u-PA IgG-sepharose CL 4B affinity chromatography. After activation by plasmin, the kinetic constants for the resultant mtcu-PA against synthetic substrate S2444 hydrolysis were found to be essentially identical to rtcu-PA, suggesting that no impairment had been exerted on the catalytic active site of mtcu-PA. However, both 125I-fibrin plasma-clot lysis and fibrinogenolysis showed that mtcu-PA possessed a higher fibrinolytic activity but hardly any degradation of fibrinogen in plasma compared to rtcu-PA and rscu-PA. It was concluded that the substitution of Arg154 by Gly154 in tcu-PA promoted the fibrin-specificity of urokinase.

  20. [Regional features of obesity-associated gene polymorphism (rs9939609 FTO gene and gene Trp64Arg ADRB3) in Russian population].

    Science.gov (United States)

    Baturin, A K; Sorokina, E Iu; Pogozheva, A V; Peskova, E V; Makurina, O N; Tutel'ian, V A

    2014-01-01

    Recent studies have shown a significant association with obesity polymorphisms: rs9939609 gene due to fat mass and obesity FTO in European and some Asian and African American populations Trp64Arg ADRB3 gene in several European populations. Association of variants rs9939609 and Trp64Arg obesity was studied in 1244 the inhabitants of Moscow and Sverdlovsk regions. Genotyping was performed using allele-specific amplification, detection results in real time using TaqMan-probes complementary DNA polymorphic sites. The frequency of the mutant allele of the FTO gene in the population of Moscow and Sverdlovsk region was 45.1%, with the TT genotype was detected in 30.2% of cases, AT--49.5%, AA--20.3%. Women had the presence of the mutant allele more likely than men (48.4 vs. 42.5%). People with obesity were more genotypes AA (26.3%) and AT (52.8%) compared to the surveyed with a BMI of less than 30 kg/m2 (respectively 18.1 and 50.7%). A significantly higher incidence of risk allele A was found in individuals with obesity (52.6 and 43.4%). The presence of the mutant allele of the gene ADRB3 among the population of Moscow and Sverdlovsk regions was noted in 7.4% of cases. While 15.5% of patients had a heterozygous genotype Trp64Arg ADRB3, that is consistent with international research. The frequency of the risk allele and genotype Arg64 Trp64Arg in women (9.3 and 18.5%) was significantly higher than men (6.2 and 12.2%). The presence of the mutant allele and genotype Trp64Arg ADRB3 (respectively, 9.1 and 18.1%) were significantly more marked in the examined obese compared with those with a body mass index less than 30 kg/m2 (7.4 and 14.9%), but these differences were not statistically significant. The results of these studies suggest that genetic variants of the FTO gene rs9939609 genotype and Trp64Arg ADRB3 contribute to the development of obesity among residents of Moscow and Sverdlovsk Region of Russia. The risk of obesity increases in the case of combined polymorphisms in

  1. Distributive characteristics of Ser49Gly and Gly389Arg genetic polymorphisms of β1-adrenoceptor in Chinese Han and Dai populations

    Institute of Scientific and Technical Information of China (English)

    Zhao-qian LIU; Hong-hao ZHOU; Jie LIU; Zhi-hua XIANG; Min-yu HU; Wei MO; Lian-sheng WANG; Dong-sheng OU-YANG; Nan HE; Dan WANG

    2006-01-01

    Aim: Genetic polymorphisms causing Ser49Gly and Gly389Arg mutants of β1-adrenoceptor may result in significant changes in the function of this receptor.The aim of the present study was to investigate the frequencies of the Ser49Gly and Gly389Arg mutant alleles in healthy Chinese populations and to investigate the differences between 2 Chinese ethnic groups (Han and Dai populations) with respect to the frequencies of these alleles. Methods: A total of 225 Han Chinese and 175 Dai Chinese unrelated healthy volunteers were recruited for this study.Genomic DNA was extracted from peripheral blood leukocytes by using a standard manual chloroform-phenol extraction. Fragments spanning the 2 polymorphisms were amplified by using polymerase chain reaction with template genomic DNA and relevant primers. The DNA products including the polymorphic loci were subjected to restriction endonuclease digestion with Eco0l09I and BcgI.Digested fragments were detected with an ultraviolet detector after electrophoresis (100 V for approximately 1.5 h). Results: The frequencies of the Gly49 and Arg389 alleles were, respectively, 16.2% and 76.4% in the Han population and 14.6%and 75.7% in the Dai population. Conclusion: The polymorphisms causing the Ser49Gly and Gly389Arg mutations of the β1-adrenoceptor existed in both healthy Han and Dai Chinese populations. The frequencies of the Ser49Gly and Gly389Arg mutant alleles were not significantly different in the Han and Dai populations.However, the frequency of the Gly389 variant seems to be significantly lower in these 2 populations than in an African-American population.

  2. Arg188 in rice sucrose transporter OsSUT1 is crucial for substrate transport

    Directory of Open Access Journals (Sweden)

    Sun Ye

    2012-11-01

    Full Text Available Abstract Background Plant sucrose uptake transporters (SUTs are H+/sucrose symporters related to the major facilitator superfamily (MFS. SUTs are essential for plant growth but little is known about their transport mechanism. Recent work identified several conserved, charged amino acids within transmembrane spans (TMS in SUTs that are essential for transport activity. Here we further evaluated the role of one of these positions, R188 in the fourth TMS of OsSUT1, a type II SUT. Results The OsSUT1(R188K mutant, studied by expression in plants, yeast, and Xenopus oocytes, did not transport sucrose but showed a H+ leak that was blocked by sucrose. The H+ leak was also blocked by β-phenyl glucoside which is not translocated by OsSUT1. Replacing the corresponding Arg in type I and type III SUTs, AtSUC1(R163K and LjSUT4(R169K, respectively, also resulted in loss of sucrose transport activity. Fluorination at the glucosyl 3 and 4 positions of α-phenyl glucoside greatly decreased transport by wild type OsSUT1 but did not affect the ability to block H+ leak in the R188K mutant. Conclusion OsSUT1 R188 appears to be essential for sucrose translocation but not for substrate interaction that blocks H+ leak. Therefore, we propose that an additional binding site functions in the initial recognition of substrates. The corresponding Arg in type I and III SUTs are equally important. We propose that R188 interacts with glucosyl 3-OH and 4-OH during translocation.

  3. XRCC1 Arg194Trp and Arg399Gln polymorphisms and arsenic methylation capacity are associated with urothelial carcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Chiang, Chien-I [School of Public Health, College of Public Health and Nutrition, Taipei Medical University, Taipei, Taiwan (China); Huang, Ya-Li [Department of Public Health, School of Medicine, College of Medicine, Taipei Medical University, Taipei, Taiwan (China); Chen, Wei-Jen [School of Public Health, College of Public Health and Nutrition, Taipei Medical University, Taipei, Taiwan (China); Shiue, Horng-Sheng [Department of Chinese Medicine, Chang Gung Memorial Hospital and Chang Gung University College of Medicine, Taoyuan, Taiwan (China); Huang, Chao-Yuan; Pu, Yeong-Shiau [Department of Urology, National Taiwan University Hospital, College of Medicine National Taiwan University, Taipei, Taiwan (China); Lin, Ying-Chin [Department of Family Medicine, Shung Ho Hospital, Taipei Medical University, New Taipei, Taiwan (China); Department of Health Examination, Wan Fang Hospital, Taipei Medical University, Taipei, Taiwan (China); Division of Family Medicine, School of Medicine, Taipei Medical University, Taipei, Taiwan (China); Hsueh, Yu-Mei, E-mail: ymhsueh@tmu.edu.tw [School of Public Health, College of Public Health and Nutrition, Taipei Medical University, Taipei, Taiwan (China); Department of Public Health, School of Medicine, College of Medicine, Taipei Medical University, Taipei, Taiwan (China)

    2014-09-15

    The association between DNA repair gene polymorphisms and bladder cancer has been widely studied. However, few studies have examined the correlation between urothelial carcinoma (UC) and arsenic or its metabolites. The aim of this study was to examine the association between polymorphisms of the DNA repair genes, XRCC1 Arg194Trp, XRCC1 Arg399Gln, XRCC3 Thr241Met, and XPD Lys751Gln, with urinary arsenic profiles and UC. To this end, we conducted a hospital-based case–control study with 324 UC patients and 647 age- and gender-matched non-cancer controls. Genomic DNA was used to examine the genotype of XRCC1 Arg194Trp, XRCC1 Arg399Gln, XRCC3 Thr241Met, and XPD Lys751Gln by PCR-restriction fragment length polymorphism analysis (PCR-RFLP). Urinary arsenic profiles were measured by high performance liquid chromatography (HPLC) linked with hydride generator and atomic absorption spectrometry. The XRCC1 399 Gln/Gln and 194 Arg/Trp and Trp/Trp genotypes were significantly related to UC, and the odds ratio (OR) and 95% confidence interval (95%CI) were 1.68 (1.03–2.75) and 0.66 (0.48–0.90), respectively. Participants with higher total urinary arsenic levels, a higher percentage of inorganic arsenic (InAs%) and a lower percentage of dimethylarsinic acid (DMA%) had a higher OR of UC. Participants carrying XRCC1 risk diplotypes G-C/G-C, A-C/A-C, and A-T/G-T, and who had higher total arsenic levels, higher InAs%, or lower DMA% compared to those with other XRCC1 diplotypes had a higher OR of UC. Our results suggest that the XRCC1 399 Gln/Gln and 194 Arg/Arg DNA repair genes play an important role in poor arsenic methylation capacity, thereby increasing the risk of UC in non-obvious arsenic exposure areas. - Highlights: • The XRCC1 399Gln/Gln genotype was significantly associated with increased OR of UC. • The XRCC1 194 Arg/Trp and Trp/Trp genotype had a significantly decreased OR of UC. • Combined effect of the XRCC1 genotypes and poor arsenic methylation capacity on

  4. Polymorphism Trp64Arg of beta 3 adrenoreceptor gene: allelic frequencies and influence on insulin resistance in a multicenter study of Castilla-León Polimorfismo TRP64ARG del gen receptor beta 3: frecuencia alélica e influencia en la resistencia a la insulina en un estudio multicéntrico de Castilla y León

    Directory of Open Access Journals (Sweden)

    D. A. de Luis

    2010-04-01

    Full Text Available Background and objective: The genetic variant (Trp64Arg is a missense mutation located within the beta3 adrenoreceptor (Beta3AR. The aim of our study was to investigate the influence of Trp64Arg polymorphism in the Beta3AR gene on insulin resistance in obese patients and the allelic distribution of this polymorphismin a geographic area of Spain. Design: A population of 264 obese patients was analyzed. A bioimpedance, blood pressure, an assessment of nutritional intake, and biochemical parameters were measured. The beta 3 adrenoreceptor gene polymorphism(Trp64Arg was genotyped. Results: Two hundred and twenty six patients (77 males/149 females (85.6% had the genotype Trp64/Trp64 (wild type group with and average age of 41.12 ± 13.1 years and 38 patients (16 males/22 females Trp64/Arg64 (14.4% (mutant type group with an average age of 40.5 ± 12.7 years. High frequencies of Arg64 allele were observed in Salamanca and Valladolid. In the mutant type group, HOMA (3.75 ± 2.77 vs 5.27 ± 5.4; p Introducción y objetivos: La variante genética (Trp64Arg es una mutación localizada en el adrenoreceptor Beta 3 (Beta3AR. El objetivo de nuestro trabajo es evaluar la influencia de el polimorfismo Trp64Arg del gen de Beta3AR sobre la resistencia a la insulina en pacientes obesos, así como la distribución alélica de este polimorfismo en un área geográfica de España. Diseño: Una muestra de 264 pacientes obesos fue analizada. Se realizó una bioimpedancia, evaluación nutricional y análisis bioquímico. Se genotiparon a los pacientes en función delpolimorfismos Tr64Arg del gen adrenoreceptor-beta 3. Resultados: Un total de 227 pacientes (77 varones/149 mujeres (85,6% presentaron el genotipo Trp64/Trp64 (grupo genotipo salvaje, con una media de edad de 41,12 ± 13,1 años y un total de 38 pacientes (16 varones/22 mujeres Trp64/Arg64 (14,4% (grupo genotipo mutante con una edad media de 40,5 ± 12,7 años. Se detectó una alta frecuencia alélica (Arg64

  5. Structure prediction of the EcoRV DNA methyltransferase based on mutant profiling, secondary structure analysis, comparison with known structures of methyltransferases and isolation of catalytically inactive single mutants.

    Science.gov (United States)

    Jeltsch, A; Sobotta, T; Pingoud, A

    1996-05-01

    The EcoRV DNA methyltransferase (M.EcoRV) is an alpha-adenine methyltransferase. We have used two different programs to predict the secondary structure of M.EcoRV. The resulting consensus prediction was tested by a mutant profiling analysis. 29 neutral mutations of M.EcoRV were generated by five cycles of random mutagenesis and selection for active variants to increase the reliability of the prediction and to get a secondary structure prediction for some ambiguously predicted regions. The predicted consensus secondary structure elements could be aligned to the common topology of the structures of the catalytic domains of M.HhaI and M.TaqI. In a complementary approach we have isolated nine catalytically inactive single mutants. Five of these mutants contain an amino acid exchange within the catalytic domain of M.EcoRV (Val2-Ala, Lys81Arg, Cys192Arg, Asp193Gly, Trp231Arg). The Trp231Arg mutant binds DNA similarly to wild-type M.EcoRV, but is catalytically inactive. Hence this mutant behaves like a bona fide active site mutant. According to the structure prediction, Trp231 is located in a loop at the putative active site of M.EcoRV. The other inactive mutants were insoluble. They contain amino acid exchanges within the conserved amino acid motifs X, III or IV in M.EcoRV confirming the importance of these regions.

  6. Expression of the gene for resistance to phaseolotoxin (argK depends on the activity of genes phtABC in Pseudomonas syringae pv. phaseolicola.

    Directory of Open Access Journals (Sweden)

    Selene Aguilera

    Full Text Available The bacterium Pseudomonas syringae pv. phaseolicola produces phaseolotoxin in a temperature dependent manner, being optimally produced between 18°C and 20°C, while no detectable amounts are present above 28°C. Phaseolotoxin is an effective inhibitor of ornithine carbamoyltransferase (OCTase activity from plant, mammalian and bacterial sources and causes a phenotypic requirement for arginine. To protect the cell from its own toxin, P. syringae pv. phaseolicola synthesizes a phaseolotoxin-resistant OCTase (ROCT. The ROCT is the product of the argK gene and is synthesized only under conditions leading to phaseolotoxin synthesis. The argK gene is included in a chromosomal fragment named Pht cluster, which contains genes involved in the synthesis of phaseolotoxin. The aim of the present work was to investigate the possible involvement of other genes included in the Pht cluster in the regulation of gene argK. We conducted transcriptional analyses of argK in several mutants unable to produce phaseolotoxin, transcriptional fusions and electrophoretic mobility shift assays, which allowed us to determine that genes phtABC, located within the Pht cluster, participate in the transcriptional repression of gene argK at temperatures not permissive for phaseolotoxin biosynthesis. This repression is mediated by a protein present in both toxigenic and nontoxigenic strains of P. syringae and in E. coli, and requires the coordinated participation of phtA, phtB and phtC products in order to carry out an efficient argK repression.

  7. Triple point mutation Asp10-->His, Asn101-->Asp, Arg148-->Ser in T4 phage lysozyme leads to the molten globule.

    Science.gov (United States)

    Uversky, V N; Leontiev, V V; Gudkov, A T

    1992-12-01

    The triple amino acid replacement (Asp10-->His, Asn101-->Asp, Arg148-->Ser) in T4 phage lysozyme was carried out by site-directed mutagenesis. At acid pH (2.7) the mutant is in a conformational state with the properties of the molten globule: (i) the mutant protein molecule is essentially compact; (ii) its CD spectrum in the near UV region is drastically reduced in intensity as compared with the wild type protein spectrum; (iii) the CD spectrum in the far UV region indicates the presence of pronounced secondary structure in the mutant; (iv) unlike the wild type protein the mutant protein can bind the hydrophobic fluorescent probe, ANS.

  8. XRCC1 genetic polymorphism Arg399Gln and hepatocellular carcinoma risk: a meta-analysis.

    Science.gov (United States)

    Liu, Fei; Li, Bo; Wei, Yonggang; Yan, Lvnan; Wen, Tianfu; Zhao, Jichun; Xu, Mingqing

    2011-07-01

    Studies investigating the association between X-ray repair cross-complementing group 1 (XRCC1) genetic polymorphism Arg399Gln and hepatocellular carcinoma (HCC) risk report conflicting results. The aim of this study was to quantitatively summarize the evidence for such a relationship. Two investigators independently searched the Medline, Embase, CNKI and Chinese Biomedicine Database. Summary odds ratios (ORs) and 95% confidence intervals (95% CIs) for XRCC1 polymorphism and HCC were calculated in a fixed-effects model (the Mantel-Haenszel method) and a random-effects model (the DerSimonian and Laird method) when appropriate. The pooled ORs were performed for a codominant model (Gln/Gln vs. Arg/Arg, Arg/Gln vs. Arg/Arg), a dominant model (Gln/Gln+Arg/Gln vs. Arg/Arg) and a recessive model (Gln/Gln vs. Arg/Gln+Arg/Arg). This meta-analysis included 11 case-control studies, which included 2208 HCC cases and 3265 controls. Overall, the variant genotypes (Gln/Gln and Arg/Gln) of Arg399Gln were not associated with HCC risk when compared with the wild-type Arg/Arg homozygote (Gln/Gln vs. Arg/Arg, OR=1.01, 95% CI=0.79-1.28; Arg/Gln vs. Arg/Arg, OR=1.09, 95% CI=0.81-1.45). Similarly, no associations were found in the dominant and recessive models (dominant model, OR=1.12, 95% CI=0.85-1.47; recessive model, OR=0.99, 95% CI=0.79-1.25). Limiting the analysis to the studies within Hardy-Weinberg equilibrium, the results were persistent and robust. When stratifying for ethnicity, country/region and source of controls, no evidence of a significant association was observed in any subgroup. No publication bias was found in the present study. No association is found between the XRCC1 polymorphism Arg399Gln and the risk of HCC. © 2011 John Wiley & Sons A/S.

  9. Tiotropium is noninferior to salmeterol in maintaining improved lung function in B16-Arg/Arg patients with asthma.

    Science.gov (United States)

    Bateman, Eric D; Kornmann, Oliver; Schmidt, Peter; Pivovarova, Anna; Engel, Michael; Fabbri, Leonardo M

    2011-08-01

    The efficacy and safety of inhaled long-acting β(2)-adrenergic agonists in asthmatic patients with the B16-Arg/Arg genotype has been questioned, and the use of antimuscarinics has been proposed as an alternative in patients whose symptoms are not controlled by inhaled corticosteroids (ICSs). We compared the efficacy and safety of the long-acting anticholinergic tiotropium with salmeterol and placebo added to an ICS in B16-Arg/Arg patients with asthma that was not controlled by ICSs alone. In a double-blind, double-dummy, placebo-controlled trial, after a 4-week run-in period with 50 μg of twice-daily salmeterol administered through a metered-dose inhaler, 388 asthmatic patients were randomized 1:1:1 to 16 weeks of treatment with 5 μg of Respimat tiotropium administered daily in the evening, 50 μg of salmeterol administered twice daily through a metered-dose inhaler, or placebo. Patients aged 18 to 67 years demonstrated reversibility to bronchodilators, and their symptoms were uncontrolled by regular ICSs (400-1000 μg of budesonide/equivalent). ICS regimens were maintained throughout the trial. The mean weekly morning peak expiratory flow (PEF) before randomization was 358 ± 115.7 L/min (range, 80.3-733.0 L/min). Changes in weekly PEF from the last week of the run-in period to the last week of treatment (primary end point: change in PEF) were -3.9 ± 4.87 L/min (n = 128) for tiotropium and -3.2 ± 4.64 L/min (n = 134) for salmeterol, and these were superior to placebo (-24.6 ± 4.84 L/min, n = 125, P < .05). Tiotropium was noninferior to salmeterol (estimated difference, -0.78 L/min [95% CI, -13.096 to 11.53]; P = .002; α = .025, 1-sided; noninferiority, 20 L/min). Tiotropium and salmeterol were numerically superior to placebo in some patient-reported secondary outcomes. Adverse events were comparable across treatments. Tiotropium was more effective than placebo and as effective as salmeterol in maintaining improved lung function in B16-Arg/Arg patients

  10. Structural insights on mouse L-threonine dehydrogenase: A regulatory role of Arg180 in catalysis.

    Science.gov (United States)

    He, Chao; Huang, Xianyu; Liu, Yanhong; Li, Fudong; Yang, Yang; Tao, Hongru; Han, Chuanchun; Zhao, Chen; Xiao, Yazhong; Shi, Yunyu

    2015-12-01

    Mouse L-threonine dehydrogenase (mTDH), which belongs to the short-chain dehydrogenase/reductase (SDR) superfamily and mediates threonine catabolism, plays pivotal roles in both powerful biosynthesis and signaling in mouse stem cells and has a regulatory residue Arg180. Here we determined three crystal structures of mTDH: wild-type (WT) in the apo form; in complex with NAD(+) and a substrate analog, glycerol, or with only NAD(+); as well as the R180K variant with NAD(+). This is the first description of a structure for mammalian SDR-type TDH. Structural comparison revealed the structural basis for SDR-type TDH catalysis remains strictly conserved in bacteria and mammals. Kinetic enzyme assays, and isothermal titration calorimetry (ITC) measurements indicated the R180K mutation has little effect on NAD(+) binding affinity, whereas affects the substrate's affinity for the enzyme. The crystal structure of R180K with NAD(+), biochemical and spectroscopic studies suggested that the R180K mutant should bind NAD(+) in a similar way and have a similar folding to the WT. However, the R180K variant may have difficulty adopting the closed form due to reduced interaction of residue 180 with a loop which connects a key position for mTDH switching between the closed and open forms in mTDH catalysis, and thereby exhibited a significantly decreased kcat/Km value toward the substrate, L-Thr. In sum, our results suggest that activity of GalE-like TDH can be regulated by remote interaction, such as hydrogen bonding and hydrophobic interaction around the Arg180 of mTDH.

  11. Knockin mouse with mutant Gα11 mimics human inherited hypocalcemia and is rescued by pharmacologic inhibitors

    DEFF Research Database (Denmark)

    Roszko, Kelly L; Bi, Ruiye; Gorvin, Caroline M

    2017-01-01

    in patients with autosomal-dominant hypocalcemia type 2 (ADH2), an inherited disorder of hypocalcemia, low parathyroid hormone (PTH), and hyperphosphatemia. We have generated knockin mice harboring the point mutation GNA11 c.C178T (p.Arg60Cys) identified in ADH2 patients. The mutant mice faithfully replicated...

  12. Use of Radioiodinated Peptide Arg-Arg-Leu Targeted to Neovascularization as well as Tumor Cells in Molecular Tumor Imaging

    Institute of Scientific and Technical Information of China (English)

    Xia Lu; Ping Yan; Rong-fu Wang; Meng Liu; Ming-ming Yu; Chun-li Zhang

    2012-01-01

    Objective:To explore a tumor peptide imaging agent Arginine-Arginine-Leucine (Tyr-Cys-Gly-Gly-Arg-Arg-Leu-Gly-Gly-Cys,tripeptide RRL [tRRL]) that targeted to tumor cells and tumor-derived endothelial cells (TDECs) and primarily investigate the possible relationship between tRRL and vascular endothelial growth factor receptor 2 (VEGFR-2).Methods:The tRRL sequence motif was identified as a tumor molecular marker specifically binding to TDECs.Tyrosine was conjugated to the amino terminal of RRL (Cys-Gly-Gly-Arg-Arg-Leu-Gly-Gly-Cys) for labeling with radionuclide iodine-131 (131I-tRRL).The uptake ability and molecular binding of tRRL to tumor cells and angiogenic endothelium were studied using flow cytometry and radioactivity counter in vitro.Whether VEGFR-2 is the binging site of tRRL was investigated.Biodistribution and single-photon emission computed tomography (SPECT) imaging of 131I-tRRL were used to evaluate the effectiveness of this new imaging agent to visualize varied tumor xenografts in nude mice.Results:In vitro cellular uptake experiments revealed that tRRL could not only adhere to tumor angiogenic endothelial cells but also largely accumulate in malignant tumor cells.VEGFR-2,which is highly expressed on TDECs,was probably not the solely binding ligand for tRRL targeted to tumor angiogenic endothelium.131I-tRRL mainly accumulated in tumors in vivo,not other organs at 24 h after injection.SPECT imaging with 131I-tRRL clearly visualized tumors in nude mice,especially at 24 h.Conclusion:Radioiodinated tRRL offers a noninvasive nuclear imaging method for functional molecular imaging of tumors targeted to neovascularization,and may be a promising candidate for tumor radioimmunotherapeutic carrier.

  13. Arg deficiency does not influence the course of Myelin Oligodendrocyte Glycoprotein (MOG35-55)-induced experimental autoimmune encephalomyelitis

    DEFF Research Database (Denmark)

    Jacobsen, Freja Aksel; Hulst, Camilla; Bäckström, Thomas;

    2016-01-01

    extensively studied in immune activation, roles for Arg are incompletely characterized. To investigate the role for Arg in experimental autoimmune encephalomyelitis, we studied disease development in Arg-/- mice. Methods: Arg-/- and Arg+/+ mice were generated from breeding of Arg+/- mice on the C57BL/6......Background: Inhibition of Abl kinases has an ameliorating effect on the rodent model for multiple sclerosis, experimental autoimmune encephalomyelitis, and arrests lymphocyte activation. The family of Abl kinases consists of the Abl1/Abl and Abl2/Arg tyrosine kinases. While the Abl kinase has been...... background. Mice were immunized with the myelin oligodendrocyte glycoprotein (MOG)35-55 peptide and disease development recorded. Lymphocyte phenotypes of wild type Arg+/+ and Arg-/- mice were studied by in vitro stimulation assays and flow cytometry. Results: The breeding of Arg+/+ and Arg-/- mice showed...

  14. P21 Ser31Arg polymorphism and cervical cancer risk: a meta-analysis.

    Science.gov (United States)

    Li, Ya; Liu, Fei; Tan, Shiqiao; Li, Shangwei

    2011-04-01

    Studies investigating the association between p21 genetic polymorphism Ser31Arg and cervical cancer (CC) risk report conflicting results. The aim of this study was to quantitatively summarize the evidence for p21 polymorphism and CC risk. Two investigators independently searched the MEDLINE, Embase, CNKI, and Chinese Biomedicine databases. Summary odds ratios (ORs) and 95% confidence intervals (CIs) for p21 polymorphism and CC were calculated in a fixed effects model (the Mantel-Haenszel method) and a random effects model (the DerSimonian and Laird method) when appropriate. The pooled ORs were performed for codominant model (Arg/Arg vs Ser/Ser and Arg/Ser vs Ser/Ser), dominant model (Arg/Arg + Arg/Ser vs Ser/Ser), and recessive model (Arg/Arg vs Arg/Ser + Ser/Ser). Subgroup analyses were performed by country, matched controls, and Hardy-Weinberg equilibrium in the controls and study sample size. This meta-analysis included 10 case-control studies from an Asian population, which included 1415 CC cases and 1947 controls. Overall, the variant genotypes (Arg/Arg and Arg/Ser) of Ser31Arg were not associated with CC risk, when compared with the wild-type homozygote Ser/Ser (Arg/Arg vs Ser/Ser: OR, 1.30; 95% CI, 0.81-2.08; Arg/Ser vs Ser/Ser: OR, 1.06; 95% CI, 0.72-1.55). Similarly, no associations were found in the dominant and recessive models (dominant model: OR, 1.05; 95% CI, 0.73-1.51; recessive model: OR, 1.28; 95% CI, 0.86-1.90). Stratified analyses also detected no significant association in any subgroup, except among those studies from "other" country and those studies with controls deviated from Hardy-Weinberg equilibrium. No association was found between the p21 polymorphism Arg31Ser and risk of CC among Asians. In the future, additional studies based on white and African American patients should be performed to re-evaluate the association.

  15. Polycyclic aromatic hydrocarbons (PAHs) enriching antibiotic resistance genes (ARGs) in the soils.

    Science.gov (United States)

    Chen, Baowei; He, Rong; Yuan, Ke; Chen, Enzhong; Lin, Lan; Chen, Xin; Sha, Sha; Zhong, Jianan; Lin, Li; Yang, Lihua; Yang, Ying; Wang, Xiaowei; Zou, Shichun; Luan, Tiangang

    2017-01-01

    The prevalence of antibiotic resistance genes (ARGs) in modern environment raises an emerging global health concern. In this study, soil samples were collected from three sites in petrochemical plant that represented different pollution levels of polycyclic aromatic hydrocarbons (PAHs). Metagenomic profiling of these soils demonstrated that ARGs in the PAHs-contaminated soils were approximately 15 times more abundant than those in the less-contaminated ones, with Proteobacterial being the preponderant phylum. Resistance profile of ARGs in the PAHs-polluted soils was characterized by the dominance of efflux pump-encoding ARGs associated with aromatic antibiotics (e.g., fluoroquinolones and acriflavine) that accounted for more than 70% of the total ARGs, which was significantly different from representative sources of ARG pollution due to wide use of antibiotics. Most of ARGs enriched in the PAHs-contaminated soils were not carried by plasmids, indicating the low possibilities of them being transferred between bacteria. Significant correlation was observed between the total abundance of ARGs and that of Proteobacteria in the soils. Proteobacteria selected by PAHs led to simultaneously enriching of ARGs carried by them in the soils. Our results suggested that PAHs could serve as one of selective stresses for greatly enriching of ARGs in the human-impacted environment.

  16. The Escherichia coli argW-dsdCXA genetic island is highly variable, and E. coli K1 strains commonly possess two copies of dsdCXA.

    Science.gov (United States)

    Moritz, Rebecca L; Welch, Rodney A

    2006-11-01

    The genome sequences of Escherichia coli pathotypes reveal extensive genetic variability in the argW-dsdCXA island. Interestingly, the archetype E. coli K1 neonatal meningitis strain, strain RS218, has two copies of the dsdCXA genes for d-serine utilization at the argW and leuX islands. Because the human brain contains d-serine, an epidemiological study emphasizing K1 isolates surveyed the dsdCXA copy number and function. Forty of 41 (97.5%) independent E. coli K1 isolates could utilize d-serine. Southern blot hybridization revealed physical variability within the argW-dsdC region, even among 22 E. coli O18:K1:H7 isolates. In addition, 30 of 41 K1 strains, including 21 of 22 O18:K1:H7 isolates, had two dsdCXA loci. Mutational analysis indicated that each of the dsdA genes is functional in a rifampin-resistant mutant of RS218, mutant E44. The high percentage of K1 strains that can use d-serine is in striking contrast to our previous observation that only 4 of 74 (5%) isolates in the diarrheagenic E. coli (DEC) collection have this activity. The genome sequence of diarrheagenic E. coli isolates indicates that the csrRAKB genes for sucrose utilization are often substituted for dsdC and a portion of dsdX present at the argW-dsdCXA island of extraintestinal isolates. Among DEC isolates there is a reciprocal pattern of sucrose fermentation versus d-serine utilization. The ability to use d-serine is a trait strongly selected for among E. coli K1 strains, which have the ability to infect a wide range of extraintestinal sites. Conversely, diarrheagenic E. coli pathotypes appear to have substituted sucrose for d-serine as a potential nutrient.

  17. Site-directed mutagenesis from Arg195 to His of a microalgal chloroplastidial glycerol-3-phosphate acyltransferase causes an increase in phospholipid levels in yeast

    Directory of Open Access Journals (Sweden)

    Long-Ling eOuyang

    2016-03-01

    Full Text Available To analyze the contribution of glycerol-3-phosphate acyltransferase (GPAT to the first acylation of glycerol-3-phosphate (G-3-P, the present study focused on a functional analysis of the GPAT gene from Lobosphaera incisa (designated as LiGPAT and the subcellular localization of the encoded protein LiGPAT. A full-length cDNA of LiGPAT consisting of a 1,305-bp ORF, a 1,652-bp 5′-UTR, and a 354-bp 3′-UTR, was cloned. The ORF encoded a 434-amino acid peptide, of which 63 residues at the N-terminus defined a chloroplast transit peptide. LiGPAT was exclusively localized to chloroplasts, which was shown by co-expression of LiGPAT with eGFP in Chlamydomonas reinhardtii and by immunogold labeling in L. incisa. Considering the conservation of His among the G-3-P binding sites from chloroplastidial GPATs and the substitution of His by Arg at position 195 in the LiGPAT mature protein (designated mLiGPAT, we established the heterologous expression of either mLiGPAT or its mutant (Arg195His (sdmLiGPAT in the GPAT-deficient yeast mutant gat1Δ. Lipid profile analyses of these transgenic yeasts not only validated the acylation function of LiGPAT but also indicated that the site-directed mutagenesis from Arg195 to His led to an increase in the phospholipid level in yeast. Semi-quantitative analysis of mLiGPAT and sdmLiGPAT, together with the structural superimposition of their G-3-P binding sites, indicated that the increased enzymatic activity was caused by the enlarged accessible surface of the phosphate group binding pocket when Arg195 was mutated to His. Thus, the potential of genetic manipulation of GPAT to increase the glycerolipid level in L. incisa and other microalgae would be of great interest.

  18. Effects of Arg26 and Lys27 mutation on the bioactivity of HNTX-Ⅳ

    Institute of Scientific and Technical Information of China (English)

    XIONG Xia; XU Xia; LI Dongling; CHEN Ping; LIANG Songping

    2007-01-01

    Hainantoxin-Ⅳ (HNTX-Ⅳ)was isolated from the Chinese bird spider Ornithoctorcs hainana and identified as a novel antagonist of tetrodotoxin-sensitive (TTX-S)sodium channels.As revealed by the solution structure of HNTX-Ⅳ solved by two-dimensional nuclear magnetic resonance (2D-NMR),HNTX-Ⅳ adopts an inhibitor cystine knot motif.To check the role of basic residues during HNTX-Ⅳ's interaction with TTX-S sodium channels,R26A and K27A mutants of HNTX-Ⅳ were constructed by solid-phase chemical synthesis.The synthesized peptides were purified and refolded under optimized oxidation conditions.Correct synthesis and folding were confirmed by MALDI-TOF mass spectrometry and NMR spectroscopy,respectively.Using the whole-cell patch-clamp technique,Lys27 but not Arg26 was identified as a key residue for HNTX-Ⅳ's bioactivity against TTX-S sodium channels,because R26A-HNTX-Ⅳ showed slightly reduced activity and K27A-HNTX-Ⅳ showed almost no inhibition.

  19. The BRCA1 c. 5096G>A p.Arg1699Gln (R1699Q) intermediate risk variant

    DEFF Research Database (Denmark)

    Moghadasi, Setareh; Meeks, Huong D.; Vreeswijk, Maaike Pg

    2017-01-01

    BACKGROUND: We previously showed that the BRCA1 variant c.5096G>A p.Arg1699Gln (R1699Q) was associated with an intermediate risk of breast cancer (BC) and ovarian cancer (OC). This study aimed to assess these cancer risks for R1699Q carriers in a larger cohort, including follow-up of previously...... studied families, to further define cancer risks and to propose adjusted clinical management of female BRCA1*R1699Q carriers. METHODS: Data were collected from 129 BRCA1*R1699Q families ascertained internationally by ENIGMA (Evidence-based Network for the Interpretation of Germline Mutant Alleles...... risk for BC and OC. Breast surveillance for female carriers based on mammogram annually from age 40 is advised. Bilateral salpingo-oophorectomy should be considered based on family history....

  20. [Distribution, dissemination and removal of antibiotic resistant genes (ARGs) in the aquatic environment].

    Science.gov (United States)

    Wen, Han-qing; Shi, Jun; Xun, Hao; Deng, Hui-ping

    2015-02-01

    In recent years, the intensive use of antibiotics induces the development of antibiotic resistant genes (ARGs), which is an increasingly critical problem affecting human health, and the potential toxic effects of the ARGs have drawn great attention all over the world. This review gave an overview of the occurrence, potential sources, fate and ecological risks of ARGs in the environment. What's more, the removal of ARGs by different treatment processes such as sludge digestion, constructed wetland, disinfection and advanced treatments were assessed, and the improving directions of different treatment processes were also pointed out. Additionally, the highlights in need for further research were proposed based on the current pollution status.

  1. Distribution of antibiotic resistance genes (ARGs) in anaerobic digestion and land application of swine wastewater.

    Science.gov (United States)

    Sui, Qianwen; Zhang, Junya; Chen, Meixue; Tong, Juan; Wang, Rui; Wei, Yuansong

    2016-06-01

    Swine farm and the adjacent farmland are hot spots of ARGs. However, few studies have investigated the on-site occurrence of ARGs distributed in the process of anaerobic digestion (AD) followed by land application of swine wastewater. Two typical swine farms, in southern and northern China respectively, with AD along with land application were explored on ARG distributions. ARGs were highly abundant in raw swine wastewater, AD effectively reduced the copy number of all detected ARGs (0.21-1.34 logs removal), but the relative abundance with different resistance mechanisms showed distinctive variation trends. The reduction efficiency of ARGs was improved by stable operational temperature and longer solid retention time (SRT) of AD. ARGs in soil characterized the contamination from the irrigation of the digested liquor. The total ARGs quantity in soil fell down by 1.66 logs in idle period of winter compared to application period of summer in the northern region, whereas the total amount was steady with whole-year application in south. Some persistent (sul1 and sul2) and elevated ARGs (tetG and ereA) in AD and land application need more attention.

  2. ARGE DFC - A European approach to MCFC commercialization

    Energy Technology Data Exchange (ETDEWEB)

    Kraus, P.; Huppmann, G. [MTU Friedrichshafen GmbH, Muenchen (Germany); Heiming, A. [Ruhgras AG, Dorsten (Germany)] [and others

    1996-12-31

    The largest European program for the commercialization of the molten carbonate fuel cell technology is carried out by the European Direct Fuel Cell Consortium (ARGE DFC). The consortium consists of the following companies: (1) MTU Friedrichshafen GmbH (Germany), within the DaimlerBenz Group responsible for off-road propulsion and decentralized energy systems. (2) Haldor Topsoe A/S (Denmark), a plant engineering company and specialist for catalytic processes (3) Elkraft A.m.b.A. (Denmark), one of the countries two utility companies (4) Ruhrgas AG (Germany), the largest German gas company (5) RWE AG (Germany), the largest German electrical utility company MTU acts as a consortium leader. The company shares a license and technology exchange agreement with Energy Research Corporation of Danbury, Connecticut.

  3. Characterization of Site-Directed Mutants in the lac Permease of Escherichia coli. 1. Replacement of Histidine Residues

    NARCIS (Netherlands)

    Püttner, Irene B.; Sarkar, Hemanta K.; Padan, Etana; Lolkema, Julius S.; Kaback, H. Ronald

    1989-01-01

    Wild-type lac permease from Escherichia coli and two site-directed mutant permeases containing Arg in place of His35 and His39 or His322 were purified and reconstituted into proteoliposomes. H35-39R permease is indistinguishable from wild type with regard to all modes of translocation. In contrast,

  4. TP53 Arg72Pro polymorphism is associated with esophageal cancer risk: A meta-analysis

    Institute of Scientific and Technical Information of China (English)

    De-Ke Jiang; Lei Yao; Wen-Zhang Wang; Bo Peng; Wei-Hua Ren; Xian-Mei Yang; Long Yu

    2011-01-01

    AIM: To investigate the association between TP53 Ar-g72Pro polymorphism and esophageal cancer (EC) risk using meta-analysis.METHODS: All eligible studies published before March 1, 2010 were selected by searching PubMed using key-words “p53” or “TP53”, “polymorphism” or “variation”, “esophageal” and “cancer” or “carcinoma”. Crude odds ratios (ORs) with 95% confidence intervals (CIs) were assessed for EC risk associated with TP53 Arg72Pro polymorphism using fixed- and random-effects models.RESULTS: Nine case-control studies involving 5545 subjects were included in this meta-analysis. Signifi-cantly reduced risk of EC was associated with TP53 genotypes for Arg/Arg + Arg/Pro vs Pro/Pro (OR = 0.73, 95% CI: 0.57-0.94, P = 0.014). Subgroup analy-ses according to the source of controls and the speci-mens used for determining TP53 Arg72Pro genotypes or sample size showed that significantly reduced risk was observed only in studies which have population-based controls (Arg/Arg vs Pro/Pro: OR = 0.56, 95% CI: 0.47-0.66, P < 0.001), and use white blood cells or normal tissue to assess TP53 genotypes of cases (Arg/Arg vs Pro/Pro: OR = 0.56, 95% CI: 0.47-0.65, P < 0.001) or include at least 200 subjects (Arg/Arg vs Pro/Pro: OR = 0.56, 95% CI: 0.47-0.65, P < 0.001). Analysis restricted to well-designed studies also sup-ported the significantly decreased risk of EC (Arg/Arg vs Pro/Pro: OR = 0.54, 95% CI: 0.46-0.64, P < 0.001). CONCLUSION: TP53 Arg72 carriers are significantly as-sociated with decreased EC risk. Nevertheless, more well-designed studies are needed to confirm our findings.

  5. The Polymorphism of DNA Repair Gene ERCC2/XPD Arg156Arg and Susceptibility to Breast Cancer in a Chinese Population

    DEFF Research Database (Denmark)

    Yin, J. Y.; Liang, D. H.; Vogel, Ulla Birgitte

    2009-01-01

    found between ERCC2/XPD Arg156Arg and risk of breast cancer (AA/AC versus CC: OR = 0.79, 95% CI = 0.49-1.28, P = 0.33; AA versus CC: OR = 0.89, 95% CI = 0.49-1.63, P = 0.72; AC versus CC: OR = 0.74, 95% CI = 0.44-1.24, P = 0.25). Breast cancer cases with the variant AA genotype were marginally younger...... the association between ERCC2/XPD Arg156Arg and susceptibility to breast cancer in a Chinese population, we conducted a hospital-based case-control study consisting of 129 patients with breast cancer and 205 controls matched by age, gender, and ethnicity. PCR-RFLP was used for genotyping. No associations were...... (mean age 45 years) than cases with the wild CC genotype (mean age 50 years) (P = 0.05). There were no differences in risk estimates in relation to menopause and occurrence of breast cancer. Our findings do not suggest that ERCC2/XPD Arg156Arg contributes to breast cancer susceptibility in a Chinese...

  6. Aminopeptidase activity in rat brain synaptosomes - 2-mercaptoethanol stimulation and Arg-vasopressin degradation

    Energy Technology Data Exchange (ETDEWEB)

    Simmons, W.H.; Orawski, A.T.

    1986-03-05

    Rat brain synaptic plasma membranes contain an amastatin-inhibited aminopeptidase activity which degrades Arg-vaso-pressin (AVP). The pH optimum for AVP cleavage was found to be 6.8, similar to that reported for oxytocin. The ability of other peptides and arylamides such as oxytocin, Tyr-Phe-Met-Arg-Phe-NH/sub 2/ and Arg-Arg-..beta..NA to inhibit cleavage of (/sup 3/H-Tyr/sup 2/)-AVP suggests that the enzyme may not be specific for AVP. The AVP-cleaving activity has been solubilized and partially characterized. Synaptosomes were lysed with hypotonic buffer, washed, and extracted with 1% Nonidet P-40 detergent. The solubilized protein was chromatographed by gel filtration HPLC on Superose 6. A single peak of activity was found with a M.W. = 117,000 which could hydrolyze 1mM Ala-..beta..NA, Arg-..beta..NA, Arg-Arg-..beta..NA, Phe-Met and Phe-Arg as well as slowly cleave AVP with the ultimate release of /sup 3/H-Tyr. 2-Mercaptoethanol (3.9mM) (ME) stimulated activity 3.6 to 6.6-fold for arylamide and dipeptide substrates, but 35-fold for labelled AVP, possibly owing to reduction of the AVP disulfide bond. All activities in the presence of ME were completely inhibited by 0.2mM amastatin.

  7. Transcriptome analysis of the Lactococcus lactis ArgR and AhrC regulons

    DEFF Research Database (Denmark)

    Larsen, Rasmus; van Hijum, Sacha A. F. T.; Martinussen, Jan;

    2008-01-01

    In previous studies, we have shown that direct protein-protein. interaction between the two regulators ArgR and AhrC in Lactococcus lactis is required for arginine-dependent repression of the biosynthetic argC promoter and the activation of the catabolic arcA promoter. Here, we establish the global...

  8. Characterization of five new mutants in the carboxyl-terminal domain of human apolipoprotein E: No cosegregation with severe hyperlipidemia

    Energy Technology Data Exchange (ETDEWEB)

    Maagdenberg, A.M.J.M. van den; Bruijn, I.H. de; Hofker, M.H.; Frants, R.R. (Leiden Univ. (Netherlands)); Knijff, P. de; Smelt, A.H.M.; Leuven, J.A.G.; van' t Hooft, F.; Assmann, G.; Havekes, L.M. (Univ. Hospital, Leiden (Netherlands)); Weng, Wei; Funke, H. (Westfalische Wilhelms-Universitaet, Muester (Germany))

    1993-05-01

    Assessment of the apolipoprotein E (apoE) phenotype by isoelectric focusing of both hyperlipidemic and normolipidemic individuals identified five new variants. All mutations were confined to the downstream part of the APOE gene by using denaturing gradient gel electrophoresis (DGGE). Sequence analysis revealed five new mutations causing unique amino acid substitutions in the carboxyl-terminal part of the protein containing the putative lipid-binding domain. Three hyperlipoproteinemic probands were carriers of the APOE*2(Va1236[r arrow]Glu) allele, the APOE*3(Cys112-Arg; Arg251[r arrow]Gly) allele, or the APOE*1(Arg158[r arrow]Cys; Leu252[r arrow]Glu) allele. DGGE of the region encoding the receptor-binding domain was useful for haplotyping the mutations at codons 112 and 158. Family studies failed to demonstrate cosegregation between the new mutations and severe hyperlipoproteinemia, although a number of carriers for the APOE*3(Cys112[r arrow]Arg; Arg251[r arrow]Gly) allele and the APOE*1(Arg158-Cys; Leu252[r arrow]Glu) allele expressed hypertriglyceridemia and/ or hypercholesterolemia. Two other mutant alleles, APOE*4[sup [minus

  9. Methylation of Gata3 protein at Arg-261 regulates transactivation of the Il5 gene in T helper 2 cells.

    Science.gov (United States)

    Hosokawa, Hiroyuki; Kato, Miki; Tohyama, Hiroyuki; Tamaki, Yuuki; Endo, Yusuke; Kimura, Motoko Y; Tumes, Damon John; Motohashi, Shinichiro; Matsumoto, Masaki; Nakayama, Keiichi I; Tanaka, Tomoaki; Nakayama, Toshinori

    2015-05-22

    Gata3 acts as a master regulator for T helper 2 (Th2) cell differentiation by inducing chromatin remodeling of the Th2 cytokine loci, accelerating Th2 cell proliferation, and repressing Th1 cell differentiation. Gata3 also directly transactivates the interleukin-5 (Il5) gene via additional mechanisms that have not been fully elucidated. We herein identified a mechanism whereby the methylation of Gata3 at Arg-261 regulates the transcriptional activation of the Il5 gene in Th2 cells. Although the methylation-mimicking Gata3 mutant retained the ability to induce IL-4 and repress IFNγ production, the IL-5 production was selectively impaired. We also demonstrated that heat shock protein (Hsp) 60 strongly associates with the methylation-mimicking Gata3 mutant and negatively regulates elongation of the Il5 transcript by RNA polymerase II. Thus, arginine methylation appears to play a pivotal role in the organization of Gata3 complexes and the target gene specificity of Gata3.

  10. Mallotus philippinensis Muell. Arg (Euphorbiaceae: Ethnopharmacology and Phytochemistry Review

    Directory of Open Access Journals (Sweden)

    Mayank Gangwar

    2014-01-01

    Full Text Available Mallotus philippinensis Muell. Arg (Euphorbiaceae are widely distributed perennial shrub or small tree in tropical and subtropical region in outer Himalayas regions with an altitude below 1,000 m and are reported to have wide range of pharmacological activities. Mallotus philippinensis species are known to contain different natural compounds, mainly phenols, diterpenoids, steroids, flavonoids, cardenolides, triterpenoids, coumarins, isocoumarins, and many more especially phenols; that is, bergenin, mallotophilippinens, rottlerin, and isorottlerin have been isolated, identified, and reported interesting biological activities such as antimicrobial, antioxidant, antiviral, cytotoxicity, antioxidant, anti-inflammatory, immunoregulatory activity protein inhibition against cancer cell. We have selected all the pharmacological aspects and toxicological and all its biological related studies. The present review reveals that Mallotus philippinensis is a valuable source of medicinally important natural molecules and provides convincing support for its future use in modern medicine. However, the existing knowledge is very limited about Mallotus philippinensis and its different parts like steam, leaf, and fruit. Further, more detailed safety data pertaining to the acute and subacute toxicity and cardio- and immunotoxicity also needs to be generated for crude extracts or its pure isolated compounds. This review underlines the interest to continue the study of this genus of the Euphorbiaceae.

  11. Somatic mosaicism for the COL7A1 mutation p.Gly2034Arg in the unaffected mother of a patient with dystrophic epidermolysis bullosa pruriginosa.

    Science.gov (United States)

    van den Akker, P C; Pasmooij, A M G; Meijer, R; Scheffer, H; Jonkman, M F

    2015-03-01

    Dystrophic epidermolysis bullosa (DEB) is a heritable blistering disorder caused by mutations in the type VII collagen gene, COL7A1. Although revertant mosaicism is well known in DEB, 'forward' somatic mosaicism, in which a pathogenic mutation arises on a wild-type (WT) background, extending beyond the germ cells, has not been reported. It is therefore unknown what proportion of sporadic dominant DEB (DDEB) cases result from de novo mutations or somatic mosaic parents. In the clinically unaffected mother of a patient with DDEB pruriginosa due to the p.Gly2034Arg mutation, we identified the p.Gly2034Arg mutation in a proportion of lymphocytes and skin cells (mutational load 10-25%). Our data emphasize that forward mosaicism occurs in DDEB and highlight that mutation analysis should always be performed in the parents of sporadic DDEB patients to confirm the de novo status of the mutation. Ultimately, this will reveal the frequency of true de novo mutations and somatic mosaicism in parents, which has important implications for genetic counselling. Our data indicate that the threshold of mutant type VII procollagen to develop DDEB must be higher than 10-25%, which provides a rationale for therapeutic approaches aimed at increasing the WT : mutant type VII collagen ratio.

  12. Influence of Glu/Arg, Asp/Arg, and Glu/Lys Salt Bridges on α-Helical Stability and Folding Kinetics.

    Science.gov (United States)

    Meuzelaar, Heleen; Vreede, Jocelyne; Woutersen, Sander

    2016-06-07

    Using a combination of ultraviolet circular dichroism, temperature-jump transient-infrared spectroscopy, and molecular dynamics simulations, we investigate the effect of salt bridges between different types of charged amino-acid residue pairs on α-helix folding. We determine the stability and the folding and unfolding rates of 12 alanine-based α-helical peptides, each of which has a nearly identical composition containing three pairs of positively and negatively charged residues (either Glu(-)/Arg(+), Asp(-)/Arg(+), or Glu(-)/Lys(+)). Within each set of peptides, the distance and order of the oppositely charged residues in the peptide sequence differ, such that they have different capabilities of forming salt bridges. Our results indicate that stabilizing salt bridges (in which the interacting residues are spaced and ordered such that they favor helix formation) speed up α-helix formation by up to 50% and slow down the unfolding of the α-helix, whereas salt bridges with an unfavorable geometry have the opposite effect. Comparing the peptides with different types of charge pairs, we observe that salt bridges between side chains of Glu(-) and Arg(+) are most favorable for the speed of folding, probably because of the larger conformational space of the salt-bridging Glu(-)/Arg(+) rotamer pairs compared to Asp(-)/Arg(+) and Glu(-)/Lys(+). We speculate that the observed impact of salt bridges on the folding kinetics might explain why some proteins contain salt bridges that do not stabilize the final, folded conformation.

  13. Lack of Arg972 polymorphism in the IRS1 gene in Parakand Brazilian Indians

    OpenAIRE

    Bezerra, RMN; Chadid, TT; Altemani, CM; Sales, TSI; Menezes, R.; Soares, MCP; Saad, STO; Saad, MJA

    2004-01-01

    Several polymorphisms in the insulin receptor substrate-1 (IRS1) gene have been reported in the last years. The most common IRS1 variant, a Gly --> Arg substitution at codon 972 (Arg972 IRS1), is more prevalent among subjects who have features of insulin resistance syndrome associated, or not, with type 2 diabetes in European populations. To determine whether the absence of IRS1 polymorphism is a more general characteristic of Paleo-Indian-derived populations, we examined the Arg972 IRS1 poly...

  14. Lack of Arg972 polymorphism in the IRS1 gene in Parakanã Brazilian Indians.

    Science.gov (United States)

    Bezerra, Rosângela M N; Chadid, Thiago T; Altemani, Claúdia M; Sales, Teresa S I; Menezes, Raimundo; Soares, Manoel C P; Saad, Sara T O; Saad, Mario J A

    2004-02-01

    Several polymorphisms in the insulin receptor substrate-1 (IRS1) gene have been reported in the last years. The most common IRS1 variant, a Gly --> Arg substitution at codon 972 (Arg972 IRS1), is more prevalent among subjects who have features of insulin resistance syndrome associated, or not, with type 2 diabetes in European populations. To determine whether the absence of IRS1 polymorphism is a more general characteristic of Paleo-Indian-derived populations, we examined the Arg972 IRS1 polymorphism in Parakanã Indians and found a lack of this polymorphism in the Parakanã population.

  15. WOOD ANATOMY OF Phyllanthus sellowianus Müll. Arg. (PHYLLANTHACEAE

    Directory of Open Access Journals (Sweden)

    Luciano Denardi

    2013-06-01

    Full Text Available http://dx.doi.org/10.5902/198050989294The present study aimed to describe the wood anatomy of Phyllanthus sellowianus Müll. Arg.(Phyllanthaceae, as well as the research of anatomical characters associated with the flexibility of their stems, desirable characteristic for bioengineering works. Belonging to the group of  reophyllous and with up to three  meters in height, the species occurs naturally in the south of Brazil, Uruguay and Argentina. The exsicat and wood sample from a P. sellowianus, located in the municipality of Santa Maria, Rio Grande do Sul state, Brazi, were deposited in the University Federal de Santa Maria under the numbers HDCF 5587 e 5588. Standard technique was used to set the histological lamina of wood. For macerations used the method of Jeffrey. The Wood anatomy description followed the IAWA Committee (1989. The microscopic analysis revealed the presence vassels of small diameter (< 60µm, radial multiples of 2 to 4 units, and simple perforation plates, obliquely, axial parenchyma absent, heterogeneous rays, with 1-3 cells large, and gelatinous fibers. The absence of axial parenchyma and laticifer tubes is compatible to the Phyllanthaceae group. The presence of perforated ray cells is, probably, the main anatomic component for differentiation of P. sellowianus from the other species of the genera. The abundant presence of gelatinous fibers certainly is one of the main anatomical characteristics related to the high flexibility of its stem.

  16. Listeria monocytogenes 10403S Arginine Repressor ArgR Finely Tunes Arginine Metabolism Regulation under Acidic Conditions

    Science.gov (United States)

    Cheng, Changyong; Dong, Zhimei; Han, Xiao; Sun, Jing; Wang, Hang; Jiang, Li; Yang, Yongchun; Ma, Tiantian; Chen, Zhongwei; Yu, Jing; Fang, Weihuan; Song, Houhui

    2017-01-01

    Listeria monocytogenes is able to colonize human and animal intestinal tracts and to subsequently cross the intestinal barrier, causing systemic infection. For successful establishment of infection, L. monocytogenes must survive the low pH environment of the stomach. L. monocytogenes encodes a functional ArgR, a transcriptional regulator belonging to the ArgR/AhrC arginine repressor family. We aimed at clarifying the specific functions of ArgR in arginine metabolism regulation, and more importantly, in acid tolerance of L. monocytogenes. We showed that ArgR in the presence of 10 mM arginine represses transcription and expression of the argGH and argCJBDF operons, indicating that L. monocytogenes ArgR plays the classical role of ArgR/AhrC family proteins in feedback inhibition of the arginine biosynthetic pathway. Notably, transcription and expression of arcA (encoding arginine deiminase) and sigB (encoding an alternative sigma factor B) were also markedly repressed by ArgR when bacteria were exposed to pH 5.5 in the absence of arginine. However, addition of arginine enabled ArgR to derepress the transcription and expression of these two genes. Electrophoretic mobility shift assays showed that ArgR binds to the putative ARG boxes in the promoter regions of argC, argG, arcA, and sigB. Reporter gene analysis with gfp under control of the argG promoter demonstrated that ArgR was able to activate the argG promoter. Unexpectedly, deletion of argR significantly increased bacterial survival in BHI medium adjusted to pH 3.5 with lactic acid. We conclude that this phenomenon is due to activation of arcA and sigB. Collectively, our results show that L. monocytogenes ArgR finely tunes arginine metabolism through negative transcriptional regulation of the arginine biosynthetic operons and of the catabolic arcA gene in an arginine-independent manner during lactic acid-induced acid stress. ArgR also appears to activate catabolism as well as sigB transcription by anti

  17. Arc/Arg3.1 Mediates Homeostatic Synaptic Scaling of AMPA Receptors

    National Research Council Canada - National Science Library

    Shepherd, Jason D; Rumbaugh, Gavin; Wu, Jing; Chowdhury, Shoaib; Plath, Niels; Kuhl, Dietmar; Huganir, Richard L; Worley, Paul F

    2006-01-01

    .... Here, we demonstrate that Arc/Arg3.1, an immediate-early gene (IEG) that is rapidly induced by neuronal activity associated with information encoding in the brain, mediates homeostatic synaptic scaling of AMPA type glutamate receptors (AMPARs...

  18. Association between p21 Ser31Arg polymorphism and cancer risk: a meta-analysis

    Institute of Scientific and Technical Information of China (English)

    Hongxia Ma; Ziyuan Zhou; Sheng Wei; Qingyi Wei

    2011-01-01

    P21 (CDKN1A), a key cell cycle regulatory protein that governs cell cycle progression from G1 to S phase, can regulate cell proliferation, growth arrest, and apoptosis. The Ser31Arg polymorphism is located in the highly conserved region of p21 and may encode functionally distinct proteins. Although many epidemiological studies have been conducted to evaluate the association between the p21 Ser31Arg polymorphism and cancer risk, the findings remain conflicting. This meta-analysis with 33 077 cases and 45 013 controls from 44 published case-control studies showed that the variant homozygous 31Arg/Arg genotype was associated with an increased risk of numerous types of cancers in a random-effect model (homozygote comparison: OR = 1.17, 95% CI = 0.99 to 1.37, P = 0.0002 for the heterogeneity test; recessive model comparison: OR = 1.16, 95% CI = 1.01 to 1.33, P = 0.0001 for the heterogeneity test). Stratified analysis revealed that increased cancer risk associated with the 31Arg/Arg genotype remained significant in subgroups of colorectal cancer, estrogen-related cancer, Caucasians, population-based studies, studies with matching information or a larger sample size. Heterogeneity analysis showed that tumor type contributed to substantial between-study heterogeneity (recessive model comparison: x2 = 21.83, df = 7, P = 0.003). The results from this large-sample sized meta-analysis suggest that the p21 31Arg/Arg genotype may serve as a potential marker for increased cancer risk.

  19. Meta and pooled analyses of FGFR4 Gly388Arg polymorphism as a cancer prognostic factor.

    Science.gov (United States)

    Frullanti, Elisa; Berking, Carola; Harbeck, Nadia; Jézéquel, Pascal; Haugen, Aage; Mawrin, Christian; Parise, Orlando; Sasaki, Hidefumi; Tsuchiya, Norihiko; Dragani, Tommaso A

    2011-07-01

    Fibroblast growth factor receptor 4 (FGFR4) contains a Gly388Arg functional polymorphism (rs351855) that has shown contrasting results in association studies. In this study, we assessed the association between the FGFR4 Gly388Arg polymorphism and cancer prognosis. Meta-analysis and pooled analysis of 6817 and 2537 cancer cases, respectively, were carried out by nodal status and overall survival. The study included the following types of cancer: brain, breast, colorectal, head and neck, larynx, lung, melanoma, prostate, sarcomas. A statistically significant association between the Arg388Arg genotype and nodal involvement was found in the meta-analysis (odds ratio=1.33, 95% confidence interval 1.01-1.74). In the pooled analysis, the Arg388 allele carriers showed an increased hazard of poor overall survival compared with homozygous carriers of the common Gly388 allele, even after adjusting for nodal status (hazard ratio=1.21, 95% confidence interval 1.05-1.40). These results provide evidence of a role for the FGFR4 Gly388Arg polymorphism in modulating patients' outcome in different types of cancer, thus offering to clinicians a new marker to predict predisposition to poor survival in cancer patients.

  20. Sequence diversity within the argF, fbp and recA genes of natural isolates of Neisseria meningitidis: interspecies recombination within the argF gene.

    Science.gov (United States)

    Zhou, J; Spratt, B G

    1992-08-01

    Studies of natural populations of Neisseria meningitidis using multilocus enzyme electrophoresis have shown extensive genetic variation within this species, which, it has been proposed, implies a level of sequence diversity within meningococci that is greater than that normally considered as the criterion for species limits in bacteria. To obtain a direct measure of the sequence diversity among meningococci, we obtained the nucleotide sequences of most of the argF, recA and fbp genes of eight meningococci of widely differing electrophoretic type (from the reference collection of Caugant). Sequence variation between the meningococcal strains ranged from 0-0.6% for fbp, 0-1.3% for argF, and 0-3.3% for recA. These levels of diversity are no greater than those found within Escherichia coli 'housekeeping' genes and suggest that multilocus enzyme electrophoresis may overestimate the extent of nucleotide sequence diversity within meningococci. The average sequence divergence between the Neisseria meningitidis strains and N. gonorrhoeae strain FA19 was 1.0% for fbp and 1.6% for recA. The argF gene, although very uniform among the eight meningococcal isolates, had a striking mosaic structure when compared with the gonococcal argF gene: two regions of the gene differed by greater than 13% in nucleotide sequence between meningococci and gonococci, whereas the rest of the gene differed by less than 1.7%. One of the diverged regions was shown to have been introduced from the argF gene of a commensal Neisseria species that is closely related to Neisseria cinerea. The source of the other region was unclear.

  1. AZT-related mutation Lys70Arg in reverse transcriptase of human immunodeficiency virus type 1 confers decrease in susceptibility to ddATP in in vitro RT inhibition assay.

    Science.gov (United States)

    Sharma, P L; Chatis, P A; Dogon, A L; Mayers, D L; McCutchan, F E; Page, C; Crumpacker, C S

    1996-09-15

    The genetic basis for didanosine (ddl) resistance in human immunodeficiency virus (HIV-1) has previously been shown to be commonly associated with a Leu to Val change at codon 74 in the HIV-1 RT gene. In this study sequential viral isolates were analyzed from five patients with prior zidovudine (AZT) use who received 6 to 16 months of ddl therapy. Following ddl therapy, viral isolates exhibited an increased AZT susceptibility and decreased ddl susceptibility. Sequence and nested PCR analysis of the HIV-1 RT gene revealed that two viral isolates contained the Leu to Val change at codon 74, and three other isolates with reduced susceptibility to ddl each contained changes at codons 65, 70, and 72. Site-directed mutagenesis was employed to insert specific mutations in RT gene of proviral clone pNL4-3. Analysis of virion-associated reverse transcriptase activity indicated that the Lys70Arg mutation resulted in an enzyme with 2- to 4-fold decreased susceptibility to ddATP. Statistical analysis of the inhibitory concentration for RT activity between pNL4-3 and mutant Lys70Arg viruses obtained in three independent RT inhibition assays was significant (P = 0.05) by student t test paired analysis. Drug susceptibility assays on the virus with Lys70Arg mutation showed a marginal decrease in susceptibility to ddl (1.5- to 2-fold) and about 4- to 6-fold decrease in susceptibility to AZT. Mutations Lys65Glu and Arg72Ser resulted in an impaired RT with greatly diminished functional RT activity. The AZT-associated Lys70Arg mutation results in an RT enzyme with decreased susceptibility to ddATP.

  2. Isolation, characterization and long term preservation of mutant strains of Xanthophyllomyces dendrorhous.

    Science.gov (United States)

    Baeza, Marcelo; Retamales, Patricio; Sepúlveda, Dionisia; Lodato, Patricia; Jiménez, Antonio; Cifuentes, Víctor

    2009-04-01

    The yeast Xanthophyllomyces dendrorhous is biotechnologically important due to its ability to produce the pigment astaxanthin, but is poorly understood at the genetic level. This is mainly because its preservation is difficult and many of the mutants obtained are unstable. The objectives of the present work were (i) the mutagenesis X. dendrorhous and, (ii) isolation of mutants with auxotrophic markers suitable for genetic studies of the carotenogenesis pathway and sexual cycle. Additionally, two kinds of preservation methods at the laboratory level were tested for the storage of strains. A collection of X. dendrorhous mutants affected in the production of carotenoid pigments or development of sexual structures and auxotrophic requirements were isolated by treatment with N-methyl-N'-nitro-N-nitrosoguanidine and the antibiotic nystatin. From a detailed analysis about the requirements of auxotrophic mutants the ARG7, ARG3 and PRO3 loci can be defined in this yeast. Among the methods assayed for the long-term preservation of X. dendrorhous strains, the dehydrated gelatin drop method showed the highest recovery of viable yeast after storage for 65 months. No changes in auxotrophic properties and in macro or micro morphology were observed after applying the latter method.

  3. Point mutation of Arg440 to his in cytochrome P450c17 causes severe 17{alpha}-hydroxylase deficiency

    Energy Technology Data Exchange (ETDEWEB)

    Fardella, C.E.; Hum, D.W.; Miller, W.L. [Univ. of California, San Francisco, CA (United States); Homoki, J. [Univ. of Ulm (Germany)

    1994-07-01

    Genetic disorders in the gene encoding P450c17 cause 17{alpha}-hydroxylase deficiency. The consequent defects in the synthesis of cortisol and sex steroids cause sexual infantilism and a female phenotype in both genetic sexes as well as mineralorcorticoid excess and hypertension. A 15-yr-old patient from Germany was seen for absent pubertal development and mild hypertension with hypokalemia, high concentrations of 17-deoxysteroids, and hypergonadotropic hypogonadism. Analysis of her P450c17 gene by polymerase chain reaction amplification and direct sequencing showed mutation of codon 440 from CGC (Arg) to CAC (His). Expression of a vector encoding this mutated form of P450c17 in transfected nonsteroidogenic COS-1 cells showed that the mutant P450c17 protein was produced, but it lacked both 17{alpha}-hydroxylase and 17,20-lyase activities. To date, 15 different P450c17 mutations have been described in 23 patients with 17{alpha}-hydroxylase deficiency, indicating that mutations in this gene are due to random events. 36 refs., 3 figs., 2 tabs.

  4. Insulin Receptor Substrate-1 (IRS-1 Gly927Arg: Correlation with Gestational Diabetes Mellitus in Saudi Women

    Directory of Open Access Journals (Sweden)

    Khalid Khalaf Alharbi

    2014-01-01

    Full Text Available Pregnant women with gestational diabetes mellitus (GDM and type 2 diabetes mellitus (T2DM share a common pathophysiology associated with similar risk factors. Genetic variants used to determine the risk of developing T2DM might also be associated with the prevalence of GDM. The aim of the present study was to scrutinize the relationship between the G972R polymorphism of the insulin receptor substrate-1 (IRS-1 gene with GDM in the Saudi female population. This is a case-control study that monitored 500 Saudi women. Subjects with GDM (n=200 were compared with non-GDM (n=300 controls. We opted to evaluate rs1801278 polymorphism in the IRS1 gene, which plays a critical role in the insulin-signaling pathway. Genotyping was performed with the Polymerase Chain Reaction-Restriction Fragment Length Polymorphism (PCR-RFLP method. The frequency of the rs1801278 polymorphism was significantly higher in women with GDM than in women with non-GDM (for TT + CT versus CC: P=0.02. Additionally, there was a significant increase in the frequency of the Arg-encoding mutant allele from GDM to non-GDM (for T versus C: P=0.01. Our results suggest that the rs1801278 polymorphism in the IRS-1 gene is involved in the occurrence of GDM in the Saudi population.

  5. Insulin receptor substrate-1 (IRS-1) Gly927Arg: correlation with gestational diabetes mellitus in Saudi women.

    Science.gov (United States)

    Alharbi, Khalid Khalaf; Khan, Imran Ali; Abotalib, Zeinab; Al-Hakeem, Malak Mohammed

    2014-01-01

    Pregnant women with gestational diabetes mellitus (GDM) and type 2 diabetes mellitus (T2DM) share a common pathophysiology associated with similar risk factors. Genetic variants used to determine the risk of developing T2DM might also be associated with the prevalence of GDM. The aim of the present study was to scrutinize the relationship between the G972R polymorphism of the insulin receptor substrate-1 (IRS-1) gene with GDM in the Saudi female population. This is a case-control study that monitored 500 Saudi women. Subjects with GDM (n = 200) were compared with non-GDM (n = 300) controls. We opted to evaluate rs1801278 polymorphism in the IRS1 gene, which plays a critical role in the insulin-signaling pathway. Genotyping was performed with the Polymerase Chain Reaction-Restriction Fragment Length Polymorphism (PCR-RFLP) method. The frequency of the rs1801278 polymorphism was significantly higher in women with GDM than in women with non-GDM (for TT + CT versus CC: P = 0.02). Additionally, there was a significant increase in the frequency of the Arg-encoding mutant allele from GDM to non-GDM (for T versus C: P = 0.01). Our results suggest that the rs1801278 polymorphism in the IRS-1 gene is involved in the occurrence of GDM in the Saudi population.

  6. FGFR4 Gly388Arg polymorphism and prostate cancer risk in Scottish men.

    Science.gov (United States)

    Ho, C K M; Anwar, S; Nanda, J; Habib, F K

    2010-03-01

    Fibroblast growth factor receptor 4 (FGFR4), a member of the fibroblast growth receptor family, was recently reported to be more abundantly expressed in malignant than benign prostate cells. A single nucleotide polymorphism at position 388 of the FGFR4 amino-acid sequence results in the substitution of glycine (Gly) with arginine (Arg) and higher frequency of the ArgArg genotype was previously found in prostate cancer patients. DNA was extracted from the blood drawn from 399 prostate cancer patients, 150 BPH patients and 294 healthy community controls. Polymerase chain reaction was carried out and single nucleotide polymorphisms of FGFR4 were identified by restriction enzyme digestion. No overall association is detectable between the Arg allele and increased prostate cancer risk. Subgroup analysis shows a higher incidence of the heterozygous ArgGly genotype in cancer cases than in the combined group of BPH and controls (PFGFR4 is not associated with increased risk of prostate cancer in Scottish men. This observation is in contrast with results from two previous studies conducted in the USA and Japan.

  7. Arg-Phe-amide-related peptides influence gonadotropin-releasing hormone neurons

    Institute of Scientific and Technical Information of China (English)

    Haluk Kelestimur; Emine Kacar; Aysegul Uzun; Mete Ozcan; Selim Kutlu

    2013-01-01

    The hypothalamic Arg-Phe-amide-related peptides, gonadotropin-inhibitory hormone and orthologous mammalian peptides of Arg-Phe-amide, may be important regulators of the hypothalamus-pituitary-gonadal reproductive axis. These peptides may modulate the effects of kisspeptins because they are presently recognized as the most potent activators of the hypothalamus-pituitary-gonadal axis. However, their effects on gonadotropin-releasing hormone neurons have not been investigated. In the current study, the GT1–7 cell line-expressing gonadotropin-releasing hormone was used as a model to explore the effects of Arg-Phe- amide-related peptides on kisspeptin activation. Intracellular calcium concentration was quantified using the calcium-sensitive dye, fura-2 acetoxymethyl ester. Gonadotropin-releasing hormone released into the medium was detected via enzyme-linked immunosorbent assay. Results showed that 100 nmol/L kisspeptin-10 significantly increased gonadotropin-releasing hormone levels (at 120 minutes of exposure) and intracellular calcium concentrations. Co-treatment of kisspeptin with 1 μmol/L gonadotropin-inhibitory hormone or 1 μmol/L Arg-Phe-amide-related peptide-1 significantly attenuated levels of kisspeptin-induced gonadotropin-releasing hormone but did not affect kisspeptin-induced elevations of intracellular calcium concentration. Overall, the results suggest that gonadotropin-inhibitory hormone and Arg-Phe-amide-related peptide-1 may have inhibitory effects on kisspeptin-activated gonadotropin-releasing hormone neurons independent of the calcium signaling pathway.

  8. Interaction pattern of Arg 62 in the A-pocket of differentially disease-associated HLA-B27 subtypes suggests distinct TCR binding modes.

    Directory of Open Access Journals (Sweden)

    Elisa Nurzia

    Full Text Available The single amino acid replacement Asp116His distinguishes the two subtypes HLA-B*2705 and HLA-B*2709 which are, respectively, associated and non-associated with Ankylosing Spondylitis, an autoimmune chronic inflammatory disease. The reason for this differential association is so far poorly understood and might be related to subtype-specific HLA:peptide conformations as well as to subtype/peptide-dependent dynamical properties on the nanoscale. Here, we combine functional experiments with extensive molecular dynamics simulations to investigate the molecular dynamics and function of the conserved Arg62 of the α1-helix for both B27 subtypes in complex with the self-peptides pVIPR (RRKWRRWHL and TIS (RRLPIFSRL, and the viral peptides pLMP2 (RRRWRRLTV and NPflu (SRYWAIRTR. Simulations of HLA:peptide systems suggest that peptide-stabilizing interactions of the Arg62 residue observed in crystal structures are metastable for both B27 subtypes under physiological conditions, rendering this arginine solvent-exposed and, probably, a key residue for TCR interaction more than peptide-binding. This view is supported by functional experiments with conservative (R62K and non-conservative (R62A B*2705 and B*2709 mutants that showed an overall reduction in their capability to present peptides to CD8+ T cells. Moreover, major subtype-dependent differences in the peptide recognition suggest distinct TCR binding modes for the B*2705 versus the B*2709 subtype.

  9. The Effect of a Novel c.820C>T (Arg274Trp) Mutation in the Mitofusin 2 Gene on Fibroblast Metabolism and Clinical Manifestation in a Patient

    Science.gov (United States)

    Kawalec, Maria; Kabzińska, Dagmara; Kochański, Andrzej; Krzyśko, Krystiana A.; Zabłocka, Barbara

    2017-01-01

    Charcot-Marie-Tooth disease type 2A (CMT2A) is an autosomal dominant axonal peripheral neuropathy caused by mutations in the mitofusin 2 gene (MFN2). Mitofusin 2 is a GTPase protein present in the outer mitochondrial membrane and responsible for regulation of mitochondrial network architecture via the fusion of mitochondria. As that fusion process is known to be strongly dependent on the GTPase activity of mitofusin 2, it is postulated that the MFN2 mutation within the GTPase domain may lead to impaired GTPase activity, and in turn to mitochondrial dysfunction. The work described here has therefore sought to verify the effects of MFN2 mutation within its GTPase domain on mitochondrial and endoplasmic reticulum morphology, as well as the mtDNA content in a cultured primary fibroblast obtained from a CMT2A patient harboring a de novo Arg274Trp mutation. In fact, all the parameters studied were affected significantly by the presence of the mutant MFN2 protein. However, using the stable model for mitofusin 2 obtained by us, we were next able to determine that the Arg274Trp mutation does not impact directly upon GTP binding. Such results were also confirmed for GTP-hydrolysis activity of MFN2 protein in patient fibroblast. We therefore suggest that the biological malfunctions observable with the disease are not consequences of impaired GTPase activity, but rather reflect an impaired contribution of the GTPase domain to other MFN2 activities involving that region, for example protein-protein interactions. PMID:28076385

  10. Dominant-negative mutation p.Arg324Thr in KCNA1 impairs Kv1.1 channel function in episodic ataxia.

    Science.gov (United States)

    Tristán-Clavijo, Enriqueta; Scholl, Francisco G; Macaya, Alfons; Iglesias, Gemma; Rojas, Ana M; Lucas, Miguel; Castellano, Antonio; Martinez-Mir, Amalia

    2016-11-01

    Episodic ataxia type 1 is a rare autosomal dominant neurological disorder caused by mutations in the KCNA1 gene that encodes the α subunit of voltage-gated potassium channel Kv1.1. The functional consequences of identified mutations on channel function do not fully correlate with the clinical phenotype of patients. A clinical and genetic study was performed in a family with 5 patients with episodic ataxia type 1, with concurrent epilepsy in 1 of them. Protein expression, modeling, and electrophysiological analyses were performed to study Kv1.1 function. Whole-genome linkage and candidate gene analyses revealed the novel heterozygous mutation p.Arg324Thr in the KCNA1 gene. The encoded mutant Kv1.1 channel displays reduced currents and altered activation and inactivation. Taken together, we provide genetic and functional evidence that mutation p.Arg324Thr in the KCNA1 gene is pathogenic and results in episodic ataxia type 1 through a dominant-negative effect. © 2016 International Parkinson and Movement Disorder Society. © 2016 International Parkinson and Movement Disorder Society.

  11. Connexin mutants and cataracts

    Directory of Open Access Journals (Sweden)

    Eric C Beyer

    2013-04-01

    Full Text Available The lens is a multicellular, but avascular tissue that must stay transparent to allow normal transmission of light and focusing of it on the retina. Damage to lens cells and/or proteins can cause cataracts, opacities that disrupt these processes. The normal survival of the lens is facilitated by an extensive network of gap junctions formed predominantly of connexin46 and connexin50. Mutations of the genes that encode these connexins (GJA3 and GJA8 have been identified and linked to inheritance of cataracts in human families and mouse lines. In vitro expression studies of several of these mutants have shown that they exhibit abnormalities that may lead to disease. Many of the mutants reduce or modify intercellular communication due to channel alterations (including loss of function or altered gating or due to impaired cellular trafficking which reduces the number of gap junction channels within the plasma membrane. However, the abnormalities detected in studies of other mutants suggest that they cause cataracts through other mechanisms including gain of hemichannel function (leading to cell injury and death and formation of cytoplasmic accumulations (that may act as light scattering particles. These observations and the anticipated results of ongoing studies should elucidate the mechanisms of cataract development due to mutations of lens connexins and abnormalities of other lens proteins. They may also contribute to our understanding of the mechanisms of disease due to connexin mutations in other tissues.

  12. Polymorphism of FGFR4 Gly388Arg does not confer an increased risk to breast cancer development.

    Science.gov (United States)

    Naidu, R; Har, Y C; Taib, N A

    2009-01-01

    The genotype analysis of the Gly and Arg allele at codon 388 of fibroblast growth factor receptor-4 (FGFR4) gene was evaluated using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method in a hospital-based Malaysian population. Peripheral blood samples were collected from 387 breast cancer patients and 252 normal and healthy women who had no history of any malignancy. The aim of the present study was to evaluate the association between the FGFR4 Gly388Arg polymorphism and breast cancer risk as well as clinicopathological parameters of the patients. The Gly/Gly, Gly/Arg, Arg/Arg, and Arg allele genotypes were detected in 46.3%, 44.4%, 9.3%, and 53.7% of breast cancer cases, respectively. The distribution of genotype (p = 0.204) and allele (p = 0.086) frequencies of FGFR4 polymorphism were not significantly different between the breast cancer cases and normal individuals. Women who were Arg/ Arg homozygotes (OR = 1.714, 95% CI 0.896-3.278), Gly/Arg heterozygotes (OR = 1.205, 95% CI 0.863-1.683), carriers of Arg allele genotype (OR = 1.269, 95% CI 0.921-1.750), or Arg allele (OR = 1.246, 95% CI 0.970-1.602) were not associated with breast cancer risk. The Arg allele genotype was significantly associated with lymph node metastases (p = 0.001) but not with other clinicopathological parameters. Our findings suggest that the polymorphic variant at codon 388 of FGFR4 gene does not confer increased risk to breast cancer development but it may be a potential genetic marker for tumor prognosis.

  13. Paralysis periodica paramyotonica caused by SCN4A Arg1448Cys mutation.

    Science.gov (United States)

    Hsu, Wei-Chih; Huang, Yung-Chuan; Wang, Chung-Wei; Hsueh, Chia-Hsiang; Lai, Ling-Ping; Yeh, Jiann-Horng

    2006-06-01

    Paralysis periodica paramyotonica is an overlapping disease that shares the features of paramyotonia characteristic of paramyotonia congenita (PC) and periodic paralysis characteristic of hyperkalemic periodic paralysis. We report the case of a 23-year-old man with paralysis periodica paramyotonica. His father and a younger brother also exhibited a similar phenotype. A SCN4A Arg1448Cys mutation was detected in this family. The affected family members exhibited marked shifts in compound muscle action potential amplitudes on exercise test, and muscle weakness could be induced by potassium loading and cold exposure. This case demonstrates that SCN4A Arg1448Cys can produce paralysis periodica paramyotonica. Other genetic or environmental factors may modulate the manifestation of SCN4A Arg1448Cys mutation.

  14. Paralysis Periodica Paramyotonica Caused by SCN4A Arg1448Cys Mutation

    Directory of Open Access Journals (Sweden)

    Wei-Chih Hsu

    2006-01-01

    Full Text Available Paralysis periodica paramyotonica is an overlapping disease that shares the features of paramyotonia characteristic of paramyotonia congenita (PC and periodic paralysis characteristic of hyperkalemic periodic paralysis. We report the case of a 23-year-old man with paralysis periodica paramyotonica. His father and a younger brother also exhibited a similar phenotype. A SCN4A Arg1448Cys mutation was detected in this family. The affected family members exhibited marked shifts in compound muscle action potential amplitudes on exercise test, and muscle weakness could be induced by potassium loading and cold exposure. This case demonstrates that SCN4A Arg1448Cys can produce paralysis periodica paramyotonica. Other genetic or environmental factors may modulate the manifestation of SCN4A Arg1448Cys mutation.

  15. Structures of the N47A and E109Q mutant proteins of pyruvoyl-dependent arginine decarboxylase from Methanococcus jannaschii

    Energy Technology Data Exchange (ETDEWEB)

    Soriano, Erika V. [Department of Chemistry and Chemical Biology, Cornell University, Ithaca, NY 14850-1301 (United States); McCloskey, Diane E. [Department of Cellular and Molecular Physiology, Pennsylvania State University, College of Medicine, Hershey, PA 17033 (United States); Kinsland, Cynthia [Department of Chemistry and Chemical Biology, Cornell University, Ithaca, NY 14850-1301 (United States); Pegg, Anthony E. [Department of Cellular and Molecular Physiology, Pennsylvania State University, College of Medicine, Hershey, PA 17033 (United States); Ealick, Steven E., E-mail: see3@cornell.edu [Department of Chemistry and Chemical Biology, Cornell University, Ithaca, NY 14850-1301 (United States)

    2008-04-01

    The crystal structures of two arginine decarboxylase mutant proteins provide insights into the mechanisms of pyruvoyl-group formation and the decarboxylation reaction. Pyruvoyl-dependent arginine decarboxylase (PvlArgDC) catalyzes the first step of the polyamine-biosynthetic pathway in plants and some archaebacteria. The pyruvoyl group of PvlArgDC is generated by an internal autoserinolysis reaction at an absolutely conserved serine residue in the proenzyme, resulting in two polypeptide chains. Based on the native structure of PvlArgDC from Methanococcus jannaschii, the conserved residues Asn47 and Glu109 were proposed to be involved in the decarboxylation and autoprocessing reactions. N47A and E109Q mutant proteins were prepared and the three-dimensional structure of each protein was determined at 2.0 Å resolution. The N47A and E109Q mutant proteins showed reduced decarboxylation activity compared with the wild-type PvlArgDC. These residues may also be important for the autoprocessing reaction, which utilizes a mechanism similar to that of the decarboxylation reaction.

  16. Structural and functional analysis of aldolase B mutants related to hereditary fructose intolerance.

    Science.gov (United States)

    Esposito, Gabriella; Vitagliano, Luigi; Santamaria, Rita; Viola, Antonietta; Zagari, Adriana; Salvatore, Francesco

    2002-11-06

    Hereditary fructose intolerance (HFI) is a recessively inherited disorder of carbohydrate metabolism caused by impaired function of human liver aldolase (B isoform). 25 enzyme-impairing mutations have been identified in the aldolase B gene. We have studied the HFI-related mutant recombinant proteins W147R, A149P, A174D, L256P, N334K and delta6ex6 in relation to aldolase B function and structure using kinetic assays and molecular graphics analysis. We found that these mutations affect aldolase B function by decreasing substrate affinity, maximal velocity and/or enzyme stability. Finally, the functional and structural analyses of the non-natural mutant Q354E provide insight into the catalytic role of Arg(303), whose natural mutants are associated to HFI.

  17. Mapping and characterization of the hisD-unlinked nif mutants in Klebsiella pneumoniae.

    Science.gov (United States)

    Xue, Z; Jiang, Q; Shen, S

    1980-02-01

    Two hisD-unlinked genes NifC5 and NifC7 are mapped in the chromosome of K. pneumoniae. The sequence of NifC5 and NifC7 is suggested as NifC5--gltB--NifC7--argG. The P1 infected E. coli lysate can transduce the mutant C-7 to be Nif+ transductant, yet fails to transduce the hisD-linked nif mutants to be Nif+ ones. This indicates that the gene encoding C-7 is not the structural gene of nitrogen fixation and is present in E. coli. It is actually a gene specifying the glutamate synthetase. SDS electrophoresis shows the marked low content of nitrogen reductase and immunoelectrophoretic test reveals the reduced amount of both nitrogenase and nitrogen reductase in the mutant cells.

  18. Secretos de Mutantes

    OpenAIRE

    Marín, Martha; Muñoz, Germán; Serrano, Rafael

    2017-01-01

    Apartándose de enfoques que consideran las culturas juveniles como ‘desviaciones sociales', ‘tribus urbanas' o ‘nuevos movimientos políticos', Secretos de mutantes bucea en culturas juveniles urbanas como la Skinhead, el Punk, el Metal, el Hardcore, el Grunge y el Hip Hop, explorándolas desde un punto de vista inédito: su dimensión de creación, para percibir los cruciales y casi desconocidos procesos que sus miembros llevan a cabo en estos vastos universos de experimentación. Esta obra se nut...

  19. A pedigree analysis of two homozygous mutant Gitelman syndrome cases.

    Science.gov (United States)

    Luo, Jiewei; Yang, Xiao; Liang, Jixing; Li, Weihua

    2015-01-01

    Gitelman syndrome (GS) is a salt-wasting tubulointerstitial disease of autosomal recessive inheritance (OMIM613395) caused by genic mutation of SLC12A3, which codes thiazide-sensitive Na-Cl cotransporter (NCCT) gene. The gene mutation of the majority of GS patients is compound heterozygous. This study analyzes two cases of GS gene mutation and the clinical phenotype. Twenty patients of two GS pedigrees underwent direct sequence alignment of 26 exons of SLC12A3 to spot and locate mutant site. Proband A of Pedigree I had three mutant sites: Arg928Cys, a homozygote, missense mutation, and two homozygous silent mutations, Ala122Ala and Thr465Thr, and 8 members of Pedigree I carried Arg928Cy heterozygous mutation. Proband B of Pedigree II had a homozygote, Ser710X, and a termination codon was spotted, which would inevitably be translated into abridged and defective protein, and 7 members had Ser710X heterozygous mutation. The heterozygous mutation carriers of the two pedigrees often have stimulus-controlled hypokalemia after strenuous exercise. The parents of Proband A are cousins, a case of intermarriage. Both probands show hypokalemia, hypochloraemia, hypocalcinuria, hyperreninemia, and hyperaldosteronemia; Proband A has normal serum magnesium and increased urinary sodium excretion, while Proband B has hypomagnesemia and increased urinary magnesium ion excretion. Both probands have normal or lower blood pressure, weakness and numbness of lower extremities, muscular soreness, and occasional palpitations and chest discomfort. Proband A wearies easily and Proband B has occasional joint numbness and pain. These two homozygous mutations are responsible for the morbidity of two GS families and they show heterogenicity of clinical phenotype.

  20. An association between TRP64ARG polymorphism of the B3 adrenoreceptor gene and some metabolic disturbances

    Directory of Open Access Journals (Sweden)

    Abilova Samai S

    2011-10-01

    Full Text Available Abstract Backgrounds B3 adrenoreceptors (ADRB3 are abundant in adipose tissue and play the role in its metabolism and lipolysis. Some variants of the ADRB3 gene may predispose subjects for the development obesity and metabolic abnormalities in the setting of modern sedentary lifestyle. ADRB3 gene polymorphism association with metabolic disturbances has never been studied before in the ethnic Kyrgyz population. Aim To study an association between Trp64Arg polymorphism of the ADRB3 and metabolic syndrome (MS components in an ethnic Kyrgyz group. Materials and methods 213 Ethnic Kyrgyz volunteers over the age of 30 were enrolled in the study. The assessment plan for each individual comprised of general physical and anthropometric exams as well as laboratory tests (glucose, lipid panel, insulin and genotyping by Trp64Arg polymorphism of the ADRB3. MS diagnosis was consistent with modified ATP III criteria (2005. Logistic regression analysis was performed to test the potential independent association between Arg64 allele with obesity, abdominal obesity (AO and arterial hypertension (AH. Results Trp64Arg polymorphism of the ADRB3 was assessed in 213 individuals (145 men, 68 women aged 30-73 (mean age 50.7 ± 7.6. Arg64 allele frequency was 0.239; ADRB3 genotype distribution among participants was: Trp64 homozygotes 54.5%, Trp64Arg 43.2% and Arg64 homozygotes 2.3%. There was an association between Trp64Arg и Arg64Arg genotypes and higher BMI, WC and obesity frequency (p Conclusion Arg64 allele of the ADRB3 gene in the studied group has an association with MS components such as obesity, AO and decreased HDL-C level.

  1. A novel distinctive cerebrovascular phenotype is associated with heterozygous Arg179 ACTA2 mutations

    NARCIS (Netherlands)

    Munot, Pinki; Saunders, Dawn E.; Milewicz, Dianna M.; Regalado, Ellen S.; Ostergaard, John R.; Braun, Kees P.; Kerr, Timothy; Lichtenbelt, Klaske D.; Philip, Sunny; Rittey, Christopher; Jacques, Thomas S.; Cox, Timothy C.; Ganesan, Vijeya

    2012-01-01

    Mutations in the ACTA2 gene lead to diffuse and diverse vascular diseases; the Arg179His mutation is associated with an early onset severe phenotype due to global smooth muscle dysfunction. Cerebrovascular disease associated with ACTA2 mutations has been likened to moyamoya disease, but appears to h

  2. An ARGS-aggrecan assay for analysis in blood and synovial fluid

    DEFF Research Database (Denmark)

    Larsson, S; Lohmander, Stefan; Struglics, A

    2014-01-01

    OBJECTIVE: To validate a modified ligand-binding assay for the detection of aggrecanase generated aggrecan fragments with the ARGS neoepitope in synovial fluid (SF) and blood, and to verify the identity of aggrecan fragments found in blood. DESIGN: An enzyme-linked immunosorbent assay (ELISA) on ...

  3. [Effects of Thermophilic Composting on Antibiotic Resistance Genes (ARGs) of Swine Manure Source].

    Science.gov (United States)

    Zheng, Ning-guo; Huang, Nan; Wang, Wei-wei; Yu, Man; Chen, Xiao-yang; Yao, Yan-lai; Wang, Wei-ping; Hong, Chun-lai

    2016-05-15

    To investigate the effects of thermophilic composting process on antibiotic resistance genes (ARGs) of swine manure source at a field scale, the abundance of four erythromycin resistance genes (ermA, ermB, ermC and ermF), three β-lactam resistance genes (blaTEM, blaCTX and blaSHV) and two quinolone resistance genes (qnrA and qnrS) were quantified by quantitative PCR ( qPCR) during the composting process. The results suggested that the erm genes' copy numbers were significantly higher than those of the bla and qnr genes in the early stage of composting (P composting process, bla and qnr genes were at low levels, while erm genes were still at high levels. Even through ermF was proliferated comparing with the initial copies. These results indicated that thermophilic composting process could not effectively remove all ARGs. For some ARGs, compost may be a good bioreactor resulting in their proliferation. Application of composting products on farmland may cause transference of ARGs.

  4. The role of Arg445 and Asp498 in the human thyroid hormone transporter MCT8

    NARCIS (Netherlands)

    S. Groeneweg (Stefan); E.C.H. Friesema (Edith); S. Kersseboom (Simone); W. Klootwijk (Willem); W.E. Visser (Wil Edward); R.P. Peeters (Robin); T.J. Visser (Theo)

    2014-01-01

    textabstractMonocarboxylate transporter 8 (MCT8) facilitates cellular influx and efflux of the thyroid hormones (THs) T4 and T3. Mutations in MCT8 lead to severe psychomotor retardation. Here, we studied the importance of 2 highly conserved residues (Arg445 in transmembrane domain 8 and Asp498 in tr

  5. Involvement of the FGFR4 Arg388 allele in head and neck squamous cell carcinoma.

    Science.gov (United States)

    Streit, Sylvia; Bange, Johannes; Fichtner, Alexander; Ihrler, Stephan; Issing, Wolfgang; Ullrich, Axel

    2004-08-20

    Fibroblast growth factor receptors (FGFRs) have been implicated in various forms of human hyperproliferative disorders such as cancers of the cervix and bladder. We investigated the expression pattern of FGFR4 and the clinical significance of the recently identified Gly/Arg polymorphism (388) in head and neck squamous cell carcinomas (HNSCCs) of the oral cavity and the oropharynx. Sections from 104 paraffin-embedded tumors were analyzed by a restriction fragment length polymorphism-based method to determine the FGFR4 genotypes. Protein expression was investigated immunohistochemically and graded into a low, intermediate, or high degree of staining. FGFR4 expression was scored as high in 17, as intermediate in 59 and as low in 28 cases. The FGFR4 Arg388 allele was found in 59 tumors, 46 of them having heterozygous and 13 homozygous genotypes. High expression of the FGFR4 Arg388 allele was significantly associated with reduced overall survival (p = 0.032) and with an advanced tumor stage (p = 0.023), whereas expression of the FGFR4 Gly388 had no impact on disease progression. Our findings indicate that high expression of FGFR4 in connection with the Arg388 allele is associated with poor clinical outcome and support the significance of FGFR4 as a diagnostic marker and a target for therapeutic intervention in human HNSCC.

  6. Hemoglobin Moabit: alpha 86 (F7) Leu leads to Arg: a new unstable abnormal hemoglobin.

    Science.gov (United States)

    Knuth, A; Pribilla, W; Marti, H R; Winterhalter, K H

    1979-01-01

    A new alpha chain abnormal hemoglobin variant was found in a Turkish patient with a mild Heinz body hemolytic anemia and splenomegaly. The substitution alpha 86 Leu leads to Arg, which is next to the heme binding proximal histidine, is responsible for a marked instability of the molecule. The oxygen affinity of the erythrocytes was found to be slightly decreased.

  7. The architecture of ArgR-DNA complexes at the genome-scale in> Escherichia coli

    DEFF Research Database (Denmark)

    Cho, Suhyung; Cho, Yoo-Bok; Kang, Taek Jin;

    2015-01-01

    DNA-binding motifs that are recognized by transcription factors (TFs) have been well studied; however, challenges remain in determining the in vivo architecture of TF-DNA complexes on a genome-scale. Here, we determined the in vivo architecture of Escherichia coli arginine repressor (ArgR)-DNA co...

  8. Molecular Interactions Between the Active Sites of RGD (Arg-Gly-Asp with its Receptor (Integrine

    Directory of Open Access Journals (Sweden)

    E. Jauregui

    2000-03-01

    Full Text Available A study of the molecular interactions between the active sites of RGD (Arg-Gly-Asp with it Receptor using simultaions is reported. Our calculations indicate that the guanidine-carboxylate complex is energetically favourd with respect to the guanidine-methyl tetrazole complex.

  9. A dynamic Asp-Arg interaction is essential for catalysis in microsomal prostaglandin E2 synthase.

    Science.gov (United States)

    Brock, Joseph S; Hamberg, Mats; Balagunaseelan, Navisraj; Goodman, Michael; Morgenstern, Ralf; Strandback, Emilia; Samuelsson, Bengt; Rinaldo-Matthis, Agnes; Haeggström, Jesper Z

    2016-01-26

    Microsomal prostaglandin E2 synthase type 1 (mPGES-1) is responsible for the formation of the potent lipid mediator prostaglandin E2 under proinflammatory conditions, and this enzyme has received considerable attention as a drug target. Recently, a high-resolution crystal structure of human mPGES-1 was presented, with Ser-127 being proposed as the hydrogen-bond donor stabilizing thiolate anion formation within the cofactor, glutathione (GSH). We have combined site-directed mutagenesis and activity assays with a structural dynamics analysis to probe the functional roles of such putative catalytic residues. We found that Ser-127 is not required for activity, whereas an interaction between Arg-126 and Asp-49 is essential for catalysis. We postulate that both residues, in addition to a crystallographic water, serve critical roles within the enzymatic mechanism. After characterizing the size or charge conservative mutations Arg-126-Gln, Asp-49-Asn, and Arg-126-Lys, we inferred that a crystallographic water acts as a general base during GSH thiolate formation, stabilized by interaction with Arg-126, which is itself modulated by its respective interaction with Asp-49. We subsequently found hidden conformational ensembles within the crystal structure that correlate well with our biochemical data. The resulting contact signaling network connects Asp-49 to distal residues involved in GSH binding and is ligand dependent. Our work has broad implications for development of efficient mPGES-1 inhibitors, potential anti-inflammatory and anticancer agents.

  10. The architecture of ArgR-DNA complexes at the genome-scale in Escherichia coli

    DEFF Research Database (Denmark)

    Cho, Suhyung; Cho, Yoo-Bok; Kang, Taek Jin

    2015-01-01

    R)-DNA complexes using high-throughput sequencing of exonuclease-treated chromatin-immunoprecipitated DNA (ChIP-exo). The ChIP-exo has a unique peak-pair pattern indicating 5′ and 3′ ends of ArgR-binding region. We identified 62 ArgR-binding loci, which were classified into three groups, comprising single, double...

  11. Interaction between ArgR and AhrC controls regulation of arginine metabolism in Lactococcus lactis

    NARCIS (Netherlands)

    Larsen, R; Kok, J; Kuipers, OP

    2005-01-01

    The expression of arginine metabolism in Lactococcus lactis is controlled by the two homologous transcriptional regulators ArgR and AhrC. Genome sequence analyses have shown that the occurrence of multiple homologues of the ArgR family of transcriptional regulators is a common feature of many low-G

  12. Characterization of a Chinese hamster ovary cell mutant having a mutation in elongation factor-2.

    Directory of Open Access Journals (Sweden)

    Pradeep K Gupta

    Full Text Available Retroviral insertional mutagenesis provides an effective forward genetic method for identifying genes involved in essential cellular pathways. A Chinese hamster ovary cell line mutant resistant to several bacterial ADP-ribosylating was obtained by this approach. The toxins used catalyze ADP-ribosylation of eukaryotic elongation factor 2 (eEF-2, block protein synthesis, and cause cell death. Strikingly, in the CHO PR328 mutant cells, the eEF-2 substrate of these ADP-ribosylating toxins was found to be modified, but the cells remained viable. A systematic study of these cells revealed the presence of a structural mutation in one allele of the eEF-2 gene. This mutation, Gly717Arg, is close to His715, the residue that is modified to become diphthamide. This Arg substitution prevents diphthamide biosynthesis at His715, rendering the mutated eEF-2 non-responsive to ADP-ribosylating toxins, while having no apparent effect on protein synthesis. Thus, CHO PR328 cells are heterozygous, having wild type and mutant eEF-2 alleles, with the latter allowing the cells to survive even in the presence of ADP-ribosylating toxins. Here, we report the comprehensive characterization of these cells.

  13. ECB deacylase mutants

    Science.gov (United States)

    Arnold, Frances H.; Shao, Zhixin; Zhao, Huimin; Giver, Lorraine J.

    2002-01-01

    A method for in vitro mutagenesis and recombination of polynucleotide sequences based on polymerase-catalyzed extension of primer oligonucleotides is disclosed. The method involves priming template polynucleotide(s) with random-sequences or defined-sequence primers to generate a pool of short DNA fragments with a low level of point mutations. The DNA fragments are subjected to denaturization followed by annealing and further enzyme-catalyzed DNA polymerization. This procedure is repeated a sufficient number of times to produce full-length genes which comprise mutants of the original template polynucleotides. These genes can be further amplified by the polymerase chain reaction and cloned into a vector for expression of the encoded proteins.

  14. Decreased uv mutagenesis in cdc8, a DNA replication mutant of Saccharomyces cerevisiae

    Energy Technology Data Exchange (ETDEWEB)

    Prakash, L.; Hinkle, D.; Prakash, S.

    1978-01-01

    A DNA replication mutant of yeast, cdc8, was found to decrease uv-induced reversion of lys2-1, arg4-17, tryl and ural. This effect was observed with all three alleles of cdc8 tested. Survival curves obtained following uv irradiation in cdc8 rad double mutants show that cdc8 is epistatic to rad6, as well as to rad1; cdc8 rad51 double mutants seem to be more sensitive than the single mutants. Since uv-induced reversion in cdc8 rad1 and cdc8 rad51 double mutants is like that of the cdc8 single mutants, we conclude that CDC8 plays a direct role in error-prone repair. To test whether CDC8 codes for a DNA polymerase, we have purified both DNA polymerase I and DNA polymerase II from cdc8 and CDC+ cells. The purified DNA polymerases from cdc8 were no more heat labile than those from CDC+, suggesting that CDC8 is not a structural gene for either enzyme.

  15. The LMNA mutation p.Arg321Ter associated with dilated cardiomyopathy leads to reduced expression and a skewed ratio of lamin A and lamin C proteins

    Energy Technology Data Exchange (ETDEWEB)

    Al-Saaidi, Rasha [Research Unit for Molecular Medicine, Aarhus University and Aarhus University Hospital, Aarhus (Denmark); Rasmussen, Torsten B. [Department of Cardiology, Aarhus University Hospital, Aarhus (Denmark); Palmfeldt, Johan [Research Unit for Molecular Medicine, Aarhus University and Aarhus University Hospital, Aarhus (Denmark); Nissen, Peter H. [Department of Clinical Biochemistry, Aarhus University Hospital, Aarhus (Denmark); Beqqali, Abdelaziz [Heart Failure Research Center, Academic Medical Center, Amsterdam (Netherlands); Hansen, Jakob [Department of Forensic Medicine, Bioanalytical Unit, University of Aarhus (Denmark); Pinto, Yigal M. [Heart Failure Research Center, Academic Medical Center, Amsterdam (Netherlands); Boesen, Thomas [Department of Molecular Biology and Genetics, University of Aarhus (Denmark); Mogensen, Jens [Department of Cardiology, Odense University Hospital, Odense (Denmark); Bross, Peter, E-mail: peter.bross@ki.au.dk [Research Unit for Molecular Medicine, Aarhus University and Aarhus University Hospital, Aarhus (Denmark)

    2013-11-15

    Dilated cardiomyopathy (DCM) is a disease of the heart muscle characterized by cardiac chamber enlargement and reduced systolic function of the left ventricle. Mutations in the LMNA gene represent the most frequent known genetic cause of DCM associated with disease of the conduction systems. The LMNA gene generates two major transcripts encoding the nuclear lamina major components lamin A and lamin C by alternative splicing. Both haploinsuffiency and dominant negative effects have been proposed as disease mechanism for premature termination codon (PTC) mutations in LMNA. These mechanisms however are still not clearly established. In this study, we used a representative LMNA nonsense mutation, p.Arg321Ter, to shed light on the molecular disease mechanisms. Cultured fibroblasts from three DCM patients carrying this mutation were analyzed. Quantitative reverse transcriptase PCR and sequencing of these PCR products indicated that transcripts from the mutant allele were degraded by the nonsense-mediated mRNA decay (NMD) mechanism. The fact that no truncated mutant protein was detectable in western blot (WB) analysis strengthens the notion that the mutant transcript is efficiently degraded. Furthermore, WB analysis showed that the expression of lamin C protein was reduced by the expected approximately 50%. Clearly decreased lamin A and lamin C levels were also observed by immunofluorescence microscopy analysis. However, results from both WB and nano-liquid chromatography/mass spectrometry demonstrated that the levels of lamin A protein were more reduced suggesting an effect on expression of lamin A from the wild type allele. PCR analysis of the ratio of lamin A to lamin C transcripts showed unchanged relative amounts of lamin A transcript suggesting that the effect on the wild type allele was operative at the protein level. Immunofluorescence microscopy analysis showed no abnormal nuclear morphology of patient fibroblast cells. Based on these data, we propose that

  16. Demonstration of the functional role of conserved Glu-Arg residues in the Staphylococcus aureus ferrichrome transporter.

    Science.gov (United States)

    Vinés, Enrique D; Speziali, Craig D; Heinrichs, David E

    2014-02-01

    The features that govern the interaction of ligand binding proteins with membrane permeases of cognate ABC transporters are largely unknown. Using sequence alignments and structural modeling based on the structure of the Escherichia coli BtuCD vitamin B12 transporter, we identified six conserved basic residues in the permease, comprised of FhuB and FhuG proteins, in the ferrichrome transporter of Staphylococcus aureus. Using alanine-scanning mutagenesis we demonstrate that two of these residues, FhuB Arg-71 and FhuG Arg-61, play a more dominant role in transporter function than FhuB Arg-74 and Arg-311, and FhuG Arg-64 and Lys-306. Moreover, we show that at positions 71 and 61 in FhuB and FhuG, respectively, arginine cannot be substituted for lysine without loss of transporter function. Previously, our laboratory demonstrated the importance of conserved acidic residues in the ferrichrome binding protein, FhuD2. Taken together, these results support the hypothesis that Glu-Arg salt bridges are critical for the interaction of the ligand binding protein with the transmembrane domains FhuB and FhuG. This hypothesis was further studied by "charge swapping" experiments whereby we constructed a S. aureus strain expressing FhuD2 with conserved residues Glu-97 and Glu-231 replaced by Arg and FhuB and FhuG with conserved basic residues Arg-71 and Arg-61, respectively, replaced by Glu. A strain containing this combination of substitutions restored partial function to the ferrichrome transporter. The results provide a direct demonstration of the functional importance of conserved basic residues on the extracellular surface of the ferrichrome permease in the Gram-positive bacterium S. aureus.

  17. Three new species of the genus Arge (Hymenoptera: Symphyta: Argidae from South Korea with key to species of the subfamily Arginae

    Directory of Open Access Journals (Sweden)

    Jin-Kyung Choi

    2016-06-01

    Full Text Available Three new species, Arge koreana Wei & Lee sp. nov. from South Korea, Arge pseudorejecta Wei & Lee sp. nov., and Arge shengi Wei & Lee sp. nov. from South Korea and China are described. Keys to known genera of Argidae and known species of Arginae from South Korea are provided.

  18. Hexane soluble extract of Mallotus philippensis (Lam.) Muell. Arg. root possesses anti-leukaemic activity

    OpenAIRE

    Khan, Musa; Qureshi, Rizwana Aleem; Hussain, Masroor; Mehmood, Khalid; Khan, Rahmat Ali

    2013-01-01

    Background Mallotus philippensis (Lam.) Muell. Arg. is a well known medicinal plant of Asia and Australia. Various compounds from different aerial parts of the plant have been reported possessing potent pharmacological, antiviral, antibacterial and cytotoxic activities. We were interested to determine the effects of some root extracts from M. philippensis on human promyelocytic leukemia HL-60 cell proliferation, cell cycle regulators and apoptosis in order to investigate its anti-leukemic pot...

  19. Mammalian neuronal actions of FMRFamide and the structurally related opioid Met-enkephalin-Arg6-Phe7.

    Science.gov (United States)

    Gayton, R J

    1982-07-15

    Since the enkephalins were first isolated a number of opioid peptides have been discovered, including a heptapeptide with the sequence Tyr-Gly-Gly-Phe-Met-Arg-Phe (Met-enkephalin-Arg6-Phe7). The heptapeptide was first isolated from chromaffin granules in bovine adrenal medulla, but using immunochemical techniques it has now been identified in human, rat and bovine brains. The C-terminal tetrapeptide of this molecule (Phe-Met-Arg-Phe) occurs in amidated form as the molluscan peptide FMRFamide. Antisera raised against FMRFamide have revealed immunoreactive material in the brains of several vertebrate species, including the rat where it occurs in nerve cell bodies and terminals. I now report that ionophoretically applied FMRFamide has an excitatory effect on rat medullary neurones which is unaffected by the opiate antagonist naloxone. In contrast, Met-enkephalin-Arg6-Phe7 and leucine-enkephalin (Leu-enkephalin) have predominantly depressant effects, which suggests that FMRFamide acts at a separate receptor.

  20. Comparative studies on the discrepant fragmentation mechanisms of the GLy-Asp-Gly-Arg and Arg-Gly-Asp-Gly: evidence for the mobile proton model.

    Science.gov (United States)

    2014-01-01

    The fragmentation mechanisms of singly protonated Gly-Asp-Gly-Arg (GDGRI and Arg-Gly-Asp-Gly (RGDGJ were investigated by mass spectrometry and theoretical methods. Both protonated molecules are fragmented mainly at the Asp-Gly amide bond C-terminal to Asp, as supported by quantum chemical calculations. Charge distributions of C and N atoms (Qc + QN) on the amide bonds were collected when the ionizing proton was fixed at different nitrogen atoms along the backbone for each peptide. Compared with the neutral molecules, the total charges of C and N atoms (Qc + QN] for the singly charged peptides tended to be negative when the proton was located at the backbone nitrogen atoms. A relatively larger value of QC + QN corresponds to a higher trend of fragmentation, which is consistent with the experimental relative abundances data that the predominant ions are y2 for [GDGR + H]+ and b3 for [RGDG + H]+. Also, the anhydride mechanism driven by the C-terminal COOH for [RGDG + H]+ was explored by a quantum-mechanical/molecular-mechanical method. Calculations indicate that the protonated peptide can be cleaved through an unusual charge-directed pathway by forming a salt bridge at the C-termini. The formation of the anhydride linkage is much more feasible since this process needs very little energy and is exother- mic, though the subsequent nucleophilic attack on the Asp carbonyl carbon is more difficult. The combined experimental and theoretical methods substantiate the mobile proton model, which opens a way to analyze quantitatively the discrepant fragmentation of dissociated peptides in peptide/protein identification.

  1. A Strong Promoter Provided with the Gene Encoding Arginyl-tRNA Synthetase(argS) from Escherichia coli.

    Science.gov (United States)

    Liu, Mo-Fang; Li, Tong; Yin, Zhao-Bao; Xu, Min-Gang; Wang, En-Duo; Wang, Yin-Lai

    2000-01-01

    Previous studies showed that the gene argS encoding the arginyl-tRNA synthetase(ArgRS) from Escherichia coli(E.coli), was overexpressed 1 000 folds in the E.coli transformant TG1/pUC-argS, while the gene leuS, encoding the leucyl-tRNA synthetase(LeuRS) from E.coli, was only overproduced 35-fold in the same case. To investigate why the expression of these two aminoacyl-tRNA synthetase genes is so different, a fused gene (termed parg-leuS) was constructed by replacement of the 5' flanking region of leuS to 5' flanking region of argS. In the E.coli transformant TG1/pUC-parg-leuS, the activity of LeuRS was only improved 8.5-fold, which was much lower than that of the transformant harboring the recombinant plasmid pUC18-leuS or pKK-leuS. However, by RNA dot hybridization the amount of mRNA produced in the transcription of parg-leuS was about 5 times than that of the wild type leuS, and was similar to that of pKK-leuS, suggesting that the promoter of argS is very strong. Analysis of the secondary structure around the initiation codon among three mRNAs showed that the secondary structure of the mRNA from parg-leuS was the strongest of the three mRNAs. From the results, it could be deduced that expression of the fused gene parg-leuS might be controlled at the translational level and the strong secondary structure of this mRNA may hinder translation initiation and result in a low translation efficiency.

  2. Knockin mouse with mutant Gα11 mimics human inherited hypocalcemia and is rescued by pharmacologic inhibitors

    Science.gov (United States)

    Roszko, Kelly L.; Bi, Ruiye; Gorvin, Caroline M.; Xiong, Xiao-Feng; Inoue, Asuka; Thakker, Rajesh V.; Strømgaard, Kristian; Gardella, Thomas

    2017-01-01

    Heterotrimeric G proteins play critical roles in transducing extracellular signals generated by 7-transmembrane domain receptors. Somatic gain-of-function mutations in G protein α subunits are associated with a variety of diseases. Recently, we identified gain-of-function mutations in Gα11 in patients with autosomal-dominant hypocalcemia type 2 (ADH2), an inherited disorder of hypocalcemia, low parathyroid hormone (PTH), and hyperphosphatemia. We have generated knockin mice harboring the point mutation GNA11 c.C178T (p.Arg60Cys) identified in ADH2 patients. The mutant mice faithfully replicated human ADH2. They also exhibited low bone mineral density and increased skin pigmentation. Treatment with NPS 2143, a negative allosteric modulator of the calcium-sensing receptor (CASR), increased PTH and calcium concentrations in WT and mutant mice, suggesting that the gain-of-function effect of GNA11R6OC is partly dependent on coupling to the CASR. Treatment with the Gα11/q-specific inhibitor YM-254890 increased blood calcium in heterozygous but not in homozygous GNA11R60C mice, consistent with published crystal structure data showing that Arg60 forms a critical contact with YM-254890. This animal model of ADH2 provides insights into molecular mechanism of this G protein–related disease and potential paths toward new lines of therapy. PMID:28194446

  3. Xaa-Arg-Gly triplets in the collagen triple helix are dominant binding sites for the molecular chaperone HSP47.

    Science.gov (United States)

    Koide, Takaki; Takahara, Yoshifumi; Asada, Shinichi; Nagata, Kazuhiro

    2002-02-22

    HSP47 is an essential procollagen-specific molecular chaperone that resides in the endoplasmic reticulum of procollagen-producing cells. Recent advances have revealed that HSP47 recognizes the (Pro-Pro-Gly)(n) sequence but not (Pro-Hyp-Gly)(n) and that HSP47 recognizes the triple-helical conformation. In this study, to better understand the substrate recognition by HSP47, we synthesized various collagen model peptides and examined their interaction with HSP47 in vitro. We found that the Pro-Arg-Gly triplet forms an HSP47-binding site. The HSP47 binding was observed only when Arg residues were incorporated in the Yaa positions of the Xaa-Yaa-Gly triplets. Amino acids in the Xaa position did not largely affect the interaction. The recognition of the Arg residue by HSP47 was specific to its side-chain structure because replacement of the Arg residue by other basic amino acids decreased the affinity to HSP47. The significance of Arg residues in HSP47 binding was further confirmed by using residue-specific chemical modification of types I and III collagen. Our results demonstrate that Xaa-Arg-Gly sequences in the triple-helical procollagen molecule are dominant binding sites for HSP47 and enable us to predict HSP47-binding sites in homotrimeric procollagen molecules.

  4. A novel distinctive cerebrovascular phenotype is associated with heterozygous Arg179 ACTA2 mutations

    Science.gov (United States)

    Munot, Pinki; Saunders, Dawn E.; Milewicz, Dianna M.; Regalado, Ellen S.; Ostergaard, John R.; Braun, Kees P.; Kerr, Timothy; Lichtenbelt, Klaske D.; Philip, Sunny; Rittey, Christopher; Jacques, Thomas S.; Cox, Timothy C.

    2012-01-01

    Mutations in the ACTA2 gene lead to diffuse and diverse vascular diseases; the Arg179His mutation is associated with an early onset severe phenotype due to global smooth muscle dysfunction. Cerebrovascular disease associated with ACTA2 mutations has been likened to moyamoya disease, but appears to have distinctive features. This study involved the analysis of neuroimaging of 13 patients with heterozygous missense mutations in ACTA2 disrupting Arg179. All patients had persistent ductus arteriosus and congenital mydriasis, and variable presentation of pulmonary hypertension, bladder and gastrointestinal problems associated with this mutation. Distinctive cerebrovascular features were dilatation of proximal internal carotid artery, occlusive disease of terminal internal carotid artery, an abnormally straight course of intracranial arteries, and absent basal ‘moyamoya’ collaterals. Patterns of brain injury supported both large and small vessel disease. Key differences from moyamoya disease were more widespread arteriopathy, the combination of arterial ectasia and stenosis and, importantly, absence of the typical basal ‘moyamoya’ collaterals. Evaluation of previously published cases suggests some of these features are also seen in the ACTA2 mutations disrupting Arg258. The observation that transition from dilated to normal/stenotic arterial calibre coincides with where the internal carotid artery changes from an elastic to muscular artery supports the hypothesis that abnormal smooth muscle cell proliferation caused by ACTA2 mutations is modulated by arterial wall components. Patients with persistent ductus arteriosus or congenital mydriasis with a label of ‘moyamoya’ should be re-evaluated to ensure the distinctive neuroimaging features of an ACTA2 mutation have not been overlooked. This diagnosis has prognostic and genetic implications, and mandates surveillance of other organ systems, in particular the aorta, to prevent life-threatening aortic dissection

  5. The Evolution of Regional Disparities in the Rural Areas of Muscelele Argeşului

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    CĂTĂLINA CÂRSTEA

    2012-01-01

    Full Text Available Regional disparities are clearly a significant challenge to the new European construction. Disparities are often sources of instability but are also sources of antagonism and unrest. In the case of Muscelele Argeşului, in the past, it was a unitary area in terms of economic and social development, the two Middle Ages royal capitals keeping the balance of this area. After the 50s, the situation changed radically and the uneven development of the towns and villages began to widen. Some villages were collectivized; others were industrialized, while a third category was simply omitted from the development plans. As a consequence, development disparities evolved and became common during the 80s. After 1992, around the towns of Curtea de Argeş and Câmpulung, two relatively more developed regions emerged, while regions in the north and central part of the area were disadvantaged. Beginning with 2002, we are witnessing a reduction of disparities because the underdeveloped areas are increasing and incorporate previously developed areas. The area is starting to become a huge underdeveloped area and some of the main factors that led to this situation are the decay of the two poles of regional development and the closing of the coal mines. Today, the entire area is experiencing a rebirth and the entrepreneurial activities are playing an important role. The huge number of small hotels and accommodation establishments constructed in the west part of the area in the last five years, coupled with several investment projects in Curtea de Argeş, has led to the increased regional importance of the town to the expense of Câmpulung. In the meantime, former mining centres that where the engines of growth for the region became rather underdeveloped. The main cause of this is the fact that the region does not have perspectives and lacks the much needed entrepreneurial activities.

  6. Tyrosyl-DNA phosphodiesterase I catalytic mutants reveal an alternative nucleophile that can catalyze substrate cleavage.

    Science.gov (United States)

    Comeaux, Evan Q; Cuya, Selma M; Kojima, Kyoko; Jafari, Nauzanene; Wanzeck, Keith C; Mobley, James A; Bjornsti, Mary-Ann; van Waardenburg, Robert C A M

    2015-03-01

    Tyrosyl-DNA phosphodiesterase I (Tdp1) catalyzes the repair of 3'-DNA adducts, such as the 3'-phosphotyrosyl linkage of DNA topoisomerase I to DNA. Tdp1 contains two conserved catalytic histidines: a nucleophilic His (His(nuc)) that attacks DNA adducts to form a covalent 3'-phosphohistidyl intermediate and a general acid/base His (His(gab)), which resolves the Tdp1-DNA linkage. A His(nuc) to Ala mutant protein is reportedly inactive, whereas the autosomal recessive neurodegenerative disease SCAN1 has been attributed to the enhanced stability of the Tdp1-DNA intermediate induced by mutation of His(gab) to Arg. However, here we report that expression of the yeast His(nuc)Ala (H182A) mutant actually induced topoisomerase I-dependent cytotoxicity and further enhanced the cytotoxicity of Tdp1 His(gab) mutants, including H432N and the SCAN1-related H432R. Moreover, the His(nuc)Ala mutant was catalytically active in vitro, albeit at levels 85-fold less than that observed with wild type Tdp1. In contrast, the His(nuc)Phe mutant was catalytically inactive and suppressed His(gab) mutant-induced toxicity. These data suggest that the activity of another nucleophile when His(nuc) is replaced with residues containing a small side chain (Ala, Asn, and Gln), but not with a bulky side chain. Indeed, genetic, biochemical, and mass spectrometry analyses show that a highly conserved His, immediately N-terminal to His(nuc), can act as a nucleophile to catalyze the formation of a covalent Tdp1-DNA intermediate. These findings suggest that the flexibility of Tdp1 active site residues may impair the resolution of mutant Tdp1 covalent phosphohistidyl intermediates and provide the rationale for developing chemotherapeutics that stabilize the covalent Tdp1-DNA intermediate.

  7. Structural and functional studies of hemoglobin Moabit (alpha 86(F7) Leu-->Arg.

    Science.gov (United States)

    Gulbis, B; Tshilolo, L; Wajcman, H; Riou, J; Galactéros, F; Promé, D; Kister, J; Papassotiriou, I; Van Laethem, Y; Vertongen, F

    1997-06-01

    An abnormal hemoglobin fraction was detected on high performance liquid chromatography profile performed for the measurement of glycated hemoglobin in a 55-year-old caucasian patient. The structural and functional studies were performed by standard techniques. Separation of hemoglobins by alkaline electrophoresis and by IEF revealed a slightly more rapid fraction than does Hb S. By acid electrophoresis, no abnormal Hb fraction could be observed. Separation of globin chains by electrophoresis demonstrated an alpha-chain variant and by chromatography, a fraction which eluted between beta and gamma globin chains. Tryptic digests and amino acid analysis have demonstrated a previously described substitution of Leu-->Arg alpha 86(F7).

  8. Rational design and characterization of D-Phe-Pro-D-Arg-derived direct thrombin inhibitors.

    Directory of Open Access Journals (Sweden)

    Ana C Figueiredo

    Full Text Available The tremendous social and economic impact of thrombotic disorders, together with the considerable risks associated to the currently available therapies, prompt for the development of more efficient and safer anticoagulants. Novel peptide-based thrombin inhibitors were identified using in silico structure-based design and further validated in vitro. The best candidate compounds contained both L- and D-amino acids, with the general sequence D-Phe(P3-Pro(P2-D-Arg(P1-P1'-CONH₂. The P1' position was scanned with L- and D-isomers of natural or unnatural amino acids, covering the major chemical classes. The most potent non-covalent and proteolysis-resistant inhibitors contain small hydrophobic or polar amino acids (Gly, Ala, Ser, Cys, Thr at the P1' position. The lead tetrapeptide, D-Phe-Pro-D-Arg-D-Thr-CONH₂, competitively inhibits α-thrombin's cleavage of the S2238 chromogenic substrate with a K(i of 0.92 µM. In order to understand the molecular details of their inhibitory action, the three-dimensional structure of three peptides (with P1' L-isoleucine (fPrI, L-cysteine (fPrC or D-threonine (fPrt in complex with human α-thrombin were determined by X-ray crystallography. All the inhibitors bind in a substrate-like orientation to the active site of the enzyme. The contacts established between the D-Arg residue in position P1 and thrombin are similar to those observed for the L-isomer in other substrates and inhibitors. However, fPrC and fPrt disrupt the active site His57-Ser195 hydrogen bond, while the combination of a P1 D-Arg and a bulkier P1' residue in fPrI induce an unfavorable geometry for the nucleophilic attack of the scissile bond by the catalytic serine. The experimental models explain the observed relative potency of the inhibitors, as well as their stability to proteolysis. Moreover, the newly identified direct thrombin inhibitors provide a novel pharmacophore platform for developing antithrombotic agents by exploring the

  9. Estudo anatômico do lenho de Sapium haematospermum Müll. Arg. (Euphorbiaceae)

    OpenAIRE

    Siegloch, Anelise Marta; Marchiori, José Newton Cardoso; Universidade Federal de Santa Maria; Santos, Sidinei Rodrigues dos

    2014-01-01

    No presente estudo é descrito o lenho de Sapium haematospermum Müll. Arg., com base em material procedente de São Francisco de Assis, Rio Grande do Sul. Foram observadas as seguintes características anatômicas, comuns em Euphorbioideae e gênero Sapium: anéis de crescimento pouco conspícuos; poros de diâmetro médio, pouco numerosos e em curtos múltiplos radiais; placas de perfuração simples; pontoações intervasculares grandes; parênquima apotraqueal difuso-em-agregados; e raios uni e bisseriad...

  10. Biologia reprodutiva de Psychotria poeppigiana Mull. Arg. (Rubiaceae em mata de galeria Reproductive biology of Psychotria poeppigiana Mull. Arg. (Rubiaceae in gallery forest

    Directory of Open Access Journals (Sweden)

    Christiano Peres Coelho

    2004-09-01

    Full Text Available Psychotria L. (Rubiaceae é o gênero com mais espécies heterostílicas dentre as Angiospermas. O objetivo deste trabalho é descrever a biologia floral, fenologia, o sistema reprodutivo e os polinizadores de Psychotria poeppigiana Müll. Arg. As inflorescências são capitadas terminais, com brácteas vermelhas e flores amarelas apresentando dois morfos distintos: brevistiladas e longistiladas, caracterizadas por flores hermafroditas com diferentes comprimentos de estiletes e posicionamento oposto das anteras (hercogamia recíproca. Encontrou-se diferenças no tamanho das anteras, dos grãos de pólen e da superfície estigmática entre os morfotipos. Polinizações controladas mostraram que os morfotipos apresentam auto-incompatibilidade e intramorfo-incompatibilidade ao nível do estigma e do estilete. Os visitantes são pequenas vespas, abelhas, borboletas e beija-flores, que visitam as flores com maior freqüência no início da manhã. De acordo com a freqüência e eficiência no comportamento de transportar os grãos de pólen entre os morfos florais, o principal polinizador foi o beija-flor Thalurania furcata. O néctar é produzido em pequena quantidade (máximo de 8µl e durante a abertura das flores, que ocorre entre 5 e 7h. A espécie forma agrupamentos devido à existência de reprodução vegetativa.Psychotria L. (Rubiaceae is the genus with the most heterostylous species in the angiosperms. The objective of this study is to describe floral biology, phenology, reproductive system, and pollinators of Psychotria poeppigiana Müll. Arg. The terminal inflorescences are capitate with red bracts and yellow flowers, and presented two different morphs: pin and thrum, characterized by hermaphroditic flowers with different lengths of styles and opposed position of the anthers (reciprocal herkogamy. Dimorphism was also observed in the size of anthers, pollen grains and stigmatic surfaces. Controlled pollination revealed self

  11. Chapa aglomerada de cimento-madeira de Hevea brasiliensis Müll. Arg. Cement-bonded particleboard of Hevea brasiliensis Müll. Arg.

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    Esmeralda Yoshico Arakaki Okino

    2004-06-01

    Full Text Available Chapas de partículas de cimento-madeira foram confeccionadas com a madeira de quatro clones de Hevea brasiliensis Müll. Arg. (seringueira: IAN 717, IAN 873, GT 711 e AVROS 1301. Confeccionaram-se as chapas na proporção de 1:4:1 (madeira:cimento:água por peso e nas dimensões de 450 x 450 x 13 mm e densidade nominal de 1,4 g/cm³, com a adição de 4% de cloreto de cálcio di-hidratado (CaCl2.2H2O como acelerador. Foram testadas partículas fervidas e não-fervidas dos quatro clones, totalizando oito tratamentos, sendo em cada um destes, com quatro repetições, avaliadas as propriedades mecânicas e físicas das chapas, segundo a norma ASTM D 1037 - 96a. De forma geral, os melhores resultados de propriedades físicas e mecânicas foram obtidos nas chapas com partículas do clone AVROS 1301. No teste de hidratação do cimento, a madeira de seringueira in natura foi classificada como de "inibição extrema", porém com a adição de CaCl2 o foi como de "baixa inibição". Essa madeira se mostrou tecnicamente viável à produção de chapas de cimento-madeira, independentemente do clone.Cement-bonded particleboards of rubberwood were manufactured with four clones of Hevea brasiliensis Müll. Arg. (rubberwood: IAN 717, IAN 873, GT 711 and AVROS 1301. Boards of 450 x 450 x 13 mm were manufactured in a ratio of 1:4:1 (wood/cement/water, weight basis, with 1.4 g/cm³ density and 4% calcium chloride dihydrated - CaCl2.2H2O as accelerator. The particles of four clones were tested in treated and untreated conditions, totaling eight treatments. In each treatment with four replicates, the physical and mechanical properties were evaluated according to ASTM D 1037 - 96a standard. Overall, the best mechanical and physical results were obtained with the cement-bonded particleboard made with particles from clone AVROS 1301. Rubberwood has shown to be "highly inhibitory" in the hydration test, however when CaCl2 was added the inhibition index decreased and

  12. THE INCIDENCE OF HIV INFECTIONS IN ARGES POPULATION IN 2012 - 2013

    Directory of Open Access Journals (Sweden)

    Ionica Deliu

    2015-12-01

    Full Text Available Human immunodeficiency virus (HIV from Retroviridae family was isolated in early 80´s but his presence in Africa is much older than we previously thought, before the pandemic spread because of the social changes of 70´s (Cernescu, 1998. The incidence of HIV infections is a constant concern for public health specialists, with inferences for whole population. In this paper we presented the incidence of HIV infections in 2012/2013 period in Arges County, Romania. Studies were made in Laboratories of Public Health Department Arges. Immunoenzymatic methods were used for anti-HIV antibodies detection (Genscreen ULTRA HIV Ag-Ab kit from BIO RAD. The results of the immunoenzymatic tests were presented in order to age and gender of the tested people. New cases of HIV infection were 1.73% in 2012 and 1.10% in 2013 from all investigated samples. The positive samples were frequent in male population, and for 25 -34 years and 35-44 years age groups. Prophylaxis is the best weapon in fight against HIV/AIDS. But the properly way to stop the spread of HIV infections may be the development of vaccine or drugs to stop progression to AIDS (Al-Jabri, 2007.

  13. Ablation of Arg-tRNA-protein transferases results in defective neural tube development.

    Science.gov (United States)

    Kim, Eunkyoung; Kim, Seonmu; Lee, Jung Hoon; Kwon, Yong Tae; Lee, Min Jae

    2016-08-01

    The arginylation branch of the N-end rule pathway is a ubiquitin-mediated proteolytic system in which post-translational conjugation of Arg by ATE1-encoded Arg-tRNA-protein transferase to N-terminal Asp, Glu, or oxidized Cys residues generates essential degradation signals. Here, we characterized the ATE1-/- mice and identified the essential role of N-terminal arginylation in neural tube development. ATE1-null mice showed severe intracerebral hemorrhages and cystic space near the neural tubes. Expression of ATE1 was prominent in the developing brain and spinal cord, and this pattern overlapped with the migration path of neural stem cells. The ATE1-/- brain showed defective G-protein signaling. Finally, we observed reduced mitosis in ATE1-/- neuroepithelium and a significantly higher nitric oxide concentration in the ATE1-/- brain. Our results strongly suggest that the crucial role of ATE1 in neural tube development is directly related to proper turn-over of the RGS4 protein, which participate in the oxygen-sensing mechanism in the cells. [BMB Reports 2016; 49(8): 443-448].

  14. Apolipoprotein E4 domain interaction accelerates diet-induced atherosclerosis in hypomorphic Arg-61 Apoe mice

    Science.gov (United States)

    Eberlé, Delphine; Kim, Roy Y.; Luk, Fu Sang; de Mochel, Nabora Soledad Reyes; Gaudreault, Nathalie; Olivas, Victor R.; Kumar, Nikit; Posada, Jessica M.; Birkeland, Andrew C.; Rapp, Joseph H.; Raffai, Robert L.

    2012-01-01

    Objective Apolipoprotein (apo) E4 is an established risk factor for atherosclerosis, but the structural components underlying this association remain unclear. ApoE4 is characterized by two biophysical properties: domain interaction and molten globule state. Substituting Arg-61 for Thr-61 in mouse apoE introduces domain interaction without molten globule state, allowing us to delineate potential pro-atherogenic effects of domain interaction in vivo. Methods and Results We studied atherosclerosis susceptibility of hypomorphic Apoe mice expressing either Thr-61 or Arg-61 apoE (ApoeTh/h or ApoeRh/h mice). On a chow diet, both mouse models were normo-lipidemic with similar levels of plasma apoE and lipoproteins. However, on a high cholesterol diet, ApoeRh/h mice displayed increased levels of total plasma cholesterol and VLDL as well as larger atherosclerotic plaques in the aortic root, arch and descending aorta compared to ApoeTh/h mice. In addition, evidence of cellular dysfunction was identified in peritoneal ApoeRh/h macrophages which released lower amounts of apoE in culture medium and displayed increased expression of MHC class II molecules. Conclusions These data indicate that domain interaction mediates pro-atherogenic effects of apoE4 in part by modulating lipoprotein metabolism and macrophage biology. Pharmaceutical targeting of domain interaction could lead to new treatments for atherosclerosis in apoE4 individuals. PMID:22441102

  15. VIDRARU RESERVOIR, ROMANIA. ENVIRONMENTAL IMPACT OF THE HYDROTEHNICAL CONSTRUCTIONS ON THE UPPER COURSE OF ARGES RIVER

    Directory of Open Access Journals (Sweden)

    Laura Ana MITITELU

    2010-12-01

    Full Text Available Having an important hydrographic system, with a significant discharge potential and being located in a place that has all the forms of relief, the basin Arges is, at present, one of the most complex hydroelectric facilities from all the rivers with reservoirs in the country. Vidraru reservoir is the biggest of its 11 reservoirs. The information (data about the management of the water in Walachia dates from the year 1576, and the oldest writing about protection against floods is known as the “Ipsilantis canal”, which stated that the big waters of Dambovita river were deviated at Lunguletu in the riverbed of Ciorogarla rivulet and dates from 1774.The effects caused by the hydrotehnical constructions on the environment are numerous and profound, both positive and negative. In this essay, the analysis of the environmental impact of the hydrotehnical facilities on Arges River is made from two perspectives. The first method of analysis is the Water Directive 2000/60 and the second method is basd on a SWOT analysis, a method taken from the economy, but very efficient in establishing the current state, and also the perpective of this environemental impact.

  16. Arg-Tyr-Asp (RYD) and Arg-Cys-Asp (RCD) motifs in dendroaspin promote selective inhibition of beta1 and beta3 integrins.

    Science.gov (United States)

    Wattam, B; Shang, D; Rahman, S; Egglezou, S; Scully, M; Kakkar, V; Lu, X

    2001-05-15

    Arg-Gly-Asp (RGD) is a unique minimal integrin-binding sequence that is found within several glycoprotein ligands. This sequence has also been found in snake-venom anti-platelet proteins, including the disintegrins and dendroaspin, a natural variant of short-chain neurotoxins isolated from the venom of Dendroaspis jamesonii. In the present study, the motifs RYD and RCD were introduced into the dendroaspin scaffold to replace RGD. Both motifs in dendroaspin caused inhibition of ADP-induced platelet aggregation with IC(50) values of 200 and 300 nM respectively, similar to that of the wild-type RGD motif (170 nM). In comparison with wild-type dendroaspin, both RYD- and RCD-containing dendroaspins were more selective in the inhibition of the adhesion of K562 cells to laminin rather than to fibrinogen and fibronectin, even though they were 10-30-fold less potent at inhibiting K562 cell (containing alpha(5)beta(1) integrin) adhesion to laminin compared with wild-type. Interestingly, the RYD motif produced a similar IC(50) value to the RGD motif at inhibiting A375-SM cell (beta(3) integrin) adhesion to collagen, whereas the RCD motif was approx. 2-6-fold less potent compared with either RGD or RYD. These findings show that the selectivity of dendroaspin binding to beta(1) and beta(3) integrins can be modulated by the introduction of alternative cell recognition sequences.

  17. The Topology of the l-Arginine Exporter ArgO Conforms to an Nin-Cout Configuration in Escherichia coli: Requirement for the Cytoplasmic N-Terminal Domain, Functional Helical Interactions, and an Aspartate Pair for ArgO Function.

    Science.gov (United States)

    Pathania, Amit; Gupta, Arvind Kumar; Dubey, Swati; Gopal, Balasubramanian; Sardesai, Abhijit A

    2016-12-01

    ArgO and LysE are members of the LysE family of exporter proteins and ordinarily mediate the export of l-arginine (Arg) in Escherichia coli and l-lysine (Lys) and Arg in Corynebacterium glutamicum, respectively. Under certain conditions, ArgO also mediates Lys export. To delineate the arrangement of ArgO in the cytoplasmic membrane of E. coli, we have employed a combination of cysteine accessibility in situ, alkaline phosphatase fusion reporters, and protein modeling to arrive at a topological model of ArgO. Our studies indicate that ArgO assumes an Nin-Cout configuration, potentially forming a five-transmembrane helix bundle flanked by a cytoplasmic N-terminal domain (NTD) comprising roughly its first 38 to 43 amino acyl residues and a short periplasmic C-terminal region (CTR). Mutagenesis studies indicate that the CTR, but not the NTD, is dispensable for ArgO function in vivo and that a pair of conserved aspartate residues, located near the opposing edges of the cytoplasmic membrane, may play a pivotal role in facilitating transmembrane Arg flux. Additional studies on amino acid substitutions that impair ArgO function in vivo and their derivatives bearing compensatory amino acid alterations indicate a role for intramolecular interactions in the Arg export mechanism, and some interactions are corroborated by normal-mode analyses. Lastly, our studies suggest that ArgO may exist as a monomer in vivo, thus highlighting the requirement for intramolecular interactions in ArgO, as opposed to interactions across multiple ArgO monomers, in the formation of an Arg-translocating conduit.

  18. Role of the salt bridge between Arg176 and Glu126 in the thermal stability of the Bacillus amyloliquefaciens α-amylase (BAA).

    Science.gov (United States)

    Zonouzi, Roseata; Khajeh, Khosro; Monajjemi, Majid; Ghaemi, Naser

    2013-01-01

    In the Bacillus amyloliquefaciens α-amylase (BAA), the loop (residues 176-185; region I) that is the part of the calcium-binding site (CaI, II) has two more amino acid residues than the α-amylase from Bacillus licheniformis (BLA). Arg176 in this region makes an ionic interaction with Glu126 from region II (residues 118-130), but this interaction is lost in BLA owing to substitution of R176Q and E126V. The goal of the present work was to quantitatively estimate the effect of ionic interaction on the overall stability of the enzyme. To clarify the functional and structural significance of the corresponding salt bridge, Glu126 was deleted (ΔE126) and converted to Val (E126V), Asp (E126D), and Lys (E126K) by site-directed mutagenesis. Kinetic constants, thermodynamic parameters, and structural changes were examined for the wild-type and mutated forms using UV-visible, atomic absoption, and fluorescence emission spectroscopies. Wild type exhibited higher k(cat) and K(m) but lower catalytic efficiency than the mutant enzymes. A decreased thermostability and an increased flexibility were also found in all of the mutant enzymes when compared with the wild type. Additionally, the calcium content of the wild type was more than ΔE126. Thus, it may be suggested that ionic interaction could decrease the mobility of the discussed region, prevent the diffusion of cations, and improve the thermostability of the whole enzyme. Based on these observations, the contribution of loop destabilization may be compensated by the formation of a salt bridge that has been used as an evolutionary mechanism or structural adaptation by the mesophilic enzyme.

  19. Co-Expression of Wild-Type P2X7R with Gln460Arg Variant Alters Receptor Function.

    Directory of Open Access Journals (Sweden)

    Fernando Aprile-Garcia

    Full Text Available The P2X7 receptor is a member of the P2X family of ligand-gated ion channels. A single-nucleotide polymorphism leading to a glutamine (Gln by arginine (Arg substitution at codon 460 of the purinergic P2X7 receptor (P2X7R has been associated with mood disorders. No change in function (loss or gain has been described for this SNP so far. Here we show that although the P2X7R-Gln460Arg variant per se is not compromised in its function, co-expression of wild-type P2X7R with P2X7R-Gln460Arg impairs receptor function with respect to calcium influx, channel currents and intracellular signaling in vitro. Moreover, co-immunoprecipitation and FRET studies show that the P2X7R-Gln460Arg variant physically interacts with P2X7R-WT. Specific silencing of either the normal or polymorphic variant rescues the heterozygous loss of function phenotype and restores normal function. The described loss of function due to co-expression, unique for mutations in the P2RX7 gene so far, explains the mechanism by which the P2X7R-Gln460Arg variant affects the normal function of the channel and may represent a mechanism of action for other mutations.

  20. FGFR4 Arg388 allele correlates with tumour thickness and FGFR4 protein expression with survival of melanoma patients.

    Science.gov (United States)

    Streit, S; Mestel, D S; Schmidt, M; Ullrich, A; Berking, C

    2006-06-19

    A single nucleotide polymorphism in the gene for FGFR4 (-Arg388) has been associated with progression in various types of human cancer. Although fibroblast growth factors (FGFs) belong to the most important growth factors in melanoma, expression of FGF receptor subtype 4 has not been investigated yet. In this study, the protein expression of this receptor was analysed in 137 melanoma tissues of different progression stages by immunohistochemistry. FGFR4 protein was expressed in 45% of the specimens and correlated with pTNM tumour stages (UICC, P = 0.023 and AJCC, P = 0.046), presence of microulceration (P = 0.009), tumour vascularity (P = 0.001), metastases (P = 0.025), number of primary tumours (P = 0.022), overall survival (P = 0.047) and disease-free survival (P = 0.024). Furthermore, FGFR4 Arg388 polymorphism was analysed in 185 melanoma patients by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). The Arg388 allele was detected in 45% of the melanoma patients and was significantly associated with tumour thickness (by Clark's level of invasion (P = 0.004) and by Breslow in mm (P = 0.02)) and the tumour subtype nodular melanoma (P = 0.002). However, there was no correlation of the FGFR4 Arg388 allele with overall and disease-free survival. In conclusion, the Arg388 genotype and the protein expression of FGFR4 may be potential markers for progression of melanoma.

  1. Comparative MD Simulations Indicate a Dual Role for Arg1323.50 in Dopamine-Dependent D2R Activation.

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    Ralf C Kling

    Full Text Available Residue Arg3.50 belongs to the highly conserved DRY-motif of class A GPCRs, which is located at the bottom of TM3. On the one hand, Arg3.50 has been reported to help stabilize the inactive state of GPCRs, but on the other hand has also been shown to be crucial for stabilizing active receptor conformations and mediating receptor-G protein coupling. The combined results of these studies suggest that the exact function of Arg3.50 is likely to be receptor-dependent and must be characterized independently for every GPCR. Consequently, we now present comparative molecular-dynamics simulations that use our recently described inactive-state and Gα-bound active-state homology models of the dopamine D2 receptor (D2R, which are either bound to dopamine or ligand-free, performed to identify the function of Arg1323.50 in D2R. Our results are consistent with a dynamic model of D2R activation in which Arg1323.50 adopts a dual role, both by stabilizing the inactive-state receptor conformation and enhancing dopamine-dependent D2R-G protein coupling.

  2. Evidence of an association between the Arg72 allele of the peptide YY and increased risk of type 2 diabetes

    DEFF Research Database (Denmark)

    Torekov, Signe S; Larsen, Lesli H; Glümer, Charlotte

    2005-01-01

    tolerance test (OGTT) (P = 0.03), an increased area under the curve for the post-OGTT plasma glucose level (P = 0.03), and a lower insulinogenic index (P = 0.01). In conclusion, the common Arg allele of the PYY Arg72Thr variant modestly associates with type 2 diabetes and with type 2 diabetes......We tested the hypothesis that variants in the gene encoding the prepropeptide YY (PYY) associate with type 2 diabetes and/or obesity. Mutation analyses of DNA from 84 patients with obesity and familial type 2 diabetes identified two polymorphisms, IVS3 + 68C>T and Arg72Thr, and one rare variant......, +151C>A of PYY. The common allele of the Arg72Thr variant associated with type 2 diabetes with an allele frequency of the Arg allele of 0.667 (95% CI 0.658-0.677) among 4,639 glucose-tolerant subjects and 0.692 (0.674-0.710) among 1,326 patients with type 2 diabetes (P = 0.005, odds ratio 1.19 [95% CI...

  3. Leptin receptor gene Gln223Arg polymorphism is not associated with obesity and metabolic syndrome in Turkish children.

    Science.gov (United States)

    Komşu-Ornek, Zuhal; Demirel, Fatma; Dursun, Ahmet; Ermiş, Bahri; Pişkin, Etem; Bideci, Aysun

    2012-01-01

    The aim of the study was to investigate the relationship between leptin receptor gene (LEPR) Gln223Arg polymorphism and obesity in Turkish children. Ninety-two obese and 99 lean children (between 5-15 years) were included in the study. Twenty-three of the obese children were diagnosed with metabolic syndrome. Blood samples were collected for morning fasting blood glucose, insulin, leptin, and lipid level measurements. LEPR Gln223Arg polymorphism was analyzed by restriction fragment length polymorphism. Significant differences were observed in anthropometric measurements, fasting blood glucose, insulin, leptin, and lipid levels between obese and lean children. Serum leptin levels were markedly higher in obese children. No significant association was noted between Gln223Arg polymorphism and serum leptin, insulin and lipid levels. There were no differences in the genotype frequencies or allele distribution for Gln223Arg polymorphism among obese, obese with metabolic syndrome and lean children. Our findings suggest that there is no association between Gln223Arg polymorphism and obesity in Turkish children.

  4. Nif- Hup- mutants of Rhizobium japonicum.

    OpenAIRE

    Moshiri, F; Stults, L; Novak, P.; Maier, R J

    1983-01-01

    Two H2 uptake-negative (Hup-) Rhizobium japonicum mutants were obtained that also lacked symbiotic N2 fixation (acetylene reduction) activity. One of the mutants formed green nodules and was deficient in heme. Hydrogen oxidation activity in this mutant could be restored by the addition of heme plus ATP to crude extracts. Bacteroid extracts from the other mutant strain lacked hydrogenase activity and activity for both of the nitrogenase component proteins. Hup+ revertants of the mutant strains...

  5. ARG098, a novel anti-human Fas antibody, suppresses synovial hyperplasia and prevents cartilage destruction in a severe combined immunodeficient-HuRAg mouse model

    Directory of Open Access Journals (Sweden)

    Matsubara Tsukasa

    2010-09-01

    Full Text Available Abstract Background The anti-human Fas/APO-1/CD95 (Fas mouse/human chimeric monoclonal IgM antibody ARG098 (ARG098 targets the human Fas molecule. The cytotoxic effects of ARG098 on cells isolated from RA patients, on normal cells in vitro, and on RA synovial tissue and cartilage in vivo using implanted rheumatoid tissues in an SCID mouse model (SCID-HuRAg were investigated to examine the potential of ARG098 as a therapy for RA. Methods ARG098 binding to each cell was analyzed by cytometry. The effects of ARG098 on several cells were assessed by a cell viability assay in vitro. Effects on the RA synovium, lymphocytes, and cartilage were assessed in vivo using the SCID-HuRAg mouse model. Results ARG098 bound to cell surface Fas molecules, and induced apoptosis in Fas-expressing RA synoviocytes and infiltrating lymphocytes in the RA synovium in a dose-dependent manner. However, ARG098 did not affect the cell viability of peripheral blood mononuclear cells of RA patients or normal chondrocytes. ARG098 also induced apoptosis in RA synoviocytes and infiltrating lymphocytes in the RA synovium in vivo. The destruction of cartilage due to synovial invasion was inhibited by ARG098 injection in the modified SCID-HuRAg mouse model. Conclusions ARG098 treatment suppressed RA synovial hyperplasia through the induction of apoptosis and prevented cartilage destruction in vivo. These results suggest that ARG098 might become a new therapy for RA.

  6. Wild Accessions and Mutant Resources

    DEFF Research Database (Denmark)

    Kawaguchi, Masayoshi; Sandal, Niels Nørgaard

    2014-01-01

    Lotus japonicus, Lotus burttii, and Lotus filicaulis are species of Lotus genus that are utilized for molecular genetic analysis such as the construction of a linkage map and QTL analysis. Among them, a number of mutants have been isolated from two wild accessions: L. japonicus Gifu B-129...

  7. Hydroimidazolone modification of the conserved Arg12 in small heat shock proteins: studies on the structure and chaperone function using mutant mimics.

    Directory of Open Access Journals (Sweden)

    Ram H Nagaraj

    Full Text Available Methylglyoxal (MGO is an α-dicarbonyl compound present ubiquitously in the human body. MGO reacts with arginine residues in proteins and forms adducts such as hydroimidazolone and argpyrimidine in vivo. Previously, we showed that MGO-mediated modification of αA-crystallin increased its chaperone function. We identified MGO-modified arginine residues in αA-crystallin and found that replacing such arginine residues with alanine residues mimicked the effects of MGO on the chaperone function. Arginine 12 (R12 is a conserved amino acid residue in Hsp27 as well as αA- and αB-crystallin. When treated with MGO at or near physiological concentrations (2-10 µM, R12 was modified to hydroimidazolone in all three small heat shock proteins. In this study, we determined the effect of arginine substitution with alanine at position 12 (R12A to mimic MGO modification on the structure and chaperone function of these proteins. Among the three proteins, the R12A mutation improved the chaperone function of only αA-crystallin. This enhancement in the chaperone function was accompanied by subtle changes in the tertiary structure, which increased the thermodynamic stability of αA-crystallin. This mutation induced the exposure of additional client protein binding sites on αA-crystallin. Altogether, our data suggest that MGO-modification of the conserved R12 in αA-crystallin to hydroimidazolone may play an important role in reducing protein aggregation in the lens during aging and cataract formation.

  8. Effects of wild-type (Trp72) and mutant (Arg72) apolipoprotein(a) kringle IV-10 on the proliferation of human arterial smooth muscle cells

    Institute of Scientific and Technical Information of China (English)

    喻红; 洪嘉玲; 汪炳华; 彭芳芳; 李小明; 何春燕

    2003-01-01

    Objective To assess the atherogenicity of lipoprotein(a), the effect of the heterogeneity of lysine binding of apolipoprotein(a) [apo(a)], a plasminogen-like glycoprotein component on the proliferation of human arterial smooth muscle cells (SMCs).Results Apo(a) wt-kringle IV-10 that has lysine binding properties possessed a growth-stimulating activity to SMCs on a dose-dependence manner by stimulating cells in the G1/G0 phase of cell cycle to S and G2/M phase, and reduced significantly the amounts of endogenous active TGF-β1 in culture when compared with the control medium and the GST group (2.4±0.5 vs 8.6±1.6 and 9.1±1.7 ng/ml, P<0.01). The growth-stimulating effect of apo(a) mut-kringle IV-10 deficient in lysine binding was negligible. Conclusions Apo(a) induces SMCs growth by inhibiting the activation of latent TGF-β1, an activity that may involve the ability of apo(a) kringle IV-10 to bind lysine. The mitogenic effect of apo(a) wt-kringle IV-10 on SMCs might play an active role in the atherogenic function of lipoprotein(a).

  9. Specific function of the Met-Tyr-Trp adduct radical and residues Arg-418 and Asp-137 in the atypical catalase reaction of catalase-peroxidase KatG.

    Science.gov (United States)

    Zhao, Xiangbo; Khajo, Abdelahad; Jarrett, Sanchez; Suarez, Javier; Levitsky, Yan; Burger, Richard M; Jarzecki, Andrzej A; Magliozzo, Richard S

    2012-10-26

    Catalase activity of the dual-function heme enzyme catalase-peroxidase (KatG) depends on several structural elements, including a unique adduct formed from covalently linked side chains of three conserved amino acids (Met-255, Tyr-229, and Trp-107, Mycobacterium tuberculosis KatG numbering) (MYW). Mutagenesis, electron paramagnetic resonance, and optical stopped-flow experiments, along with calculations using density functional theory (DFT) methods revealed the basis of the requirement for a radical on the MYW-adduct, for oxyferrous heme, and for conserved residues Arg-418 and Asp-137 in the rapid catalase reaction. The participation of an oxyferrous heme intermediate (dioxyheme) throughout the pH range of catalase activity is suggested from our finding that carbon monoxide inhibits the activity at both acidic and alkaline pH. In the presence of H(2)O(2), the MYW-adduct radical is formed normally in KatG[D137S] but this mutant is defective in forming dioxyheme and lacks catalase activity. KatG[R418L] is also catalase deficient but exhibits normal formation of the adduct radical and dioxyheme. Both mutants exhibit a coincidence between MYW-adduct radical persistence and H(2)O(2) consumption as a function of time, and enhanced subunit oligomerization during turnover, suggesting that the two mutations disrupting catalase turnover allow increased migration of the MYW-adduct radical to protein surface residues. DFT calculations showed that an interaction between the side chain of residue Arg-418 and Tyr-229 in the MYW-adduct radical favors reaction of the radical with the adjacent dioxyheme intermediate present throughout turnover in WT KatG. Release of molecular oxygen and regeneration of resting enzyme are thereby catalyzed in the last step of a proposed catalase reaction.

  10. Antithrombotic Protective Effects of Arg-Pro-Gly-Pro Peptide during Emotional Stress Provoked by Forced Swimming Test in Rats.

    Science.gov (United States)

    Grigor'eva, M E; Lyapina, L A

    2017-01-01

    Blood coagulation was enhanced and all factors (total, enzyme, and non-enzyme) of the fibrinolytic system were suppressed in rats in 60 min after forced swimming test. Argininecontaining tetrapeptide glyproline Arg-Pro-Gly-Pro administered prior to this test activated fibrinolysis and prevented hypercoagulation. Administration of this peptide in 5 min after swimming test also enhanced anticoagulant, fibrinolytic, and antithrombotic activity of the blood. Therefore, glyproline Arg-Pro-Gly-Pro exerted both preventive and curative effects on the hemostasis system and prevented enhancement of blood coagulation provoked by emotional stress modeled by forced swimming test.

  11. Neovascularization and functional recovery after intracerebral hemorrhage is conditioned by the Tp53 Arg72Pro single-nucleotide polymorphism.

    Science.gov (United States)

    Rodríguez, Cristina; Sobrino, Tomás; Agulla, Jesús; Bobo-Jiménez, Verónica; Ramos-Araque, María E; Duarte, Juan J; Gómez-Sánchez, José C; Bolaños, Juan P; Castillo, José; Almeida, Ángeles

    2017-01-01

    Intracerebral hemorrhage (ICH) is a devastating subtype of stroke that lacks effective therapy and reliable prognosis. Neovascularization following ICH is an essential compensatory response that mediates brain repair and modulates the clinical outcome of stroke patients. However, the mechanism that dictates this process is unknown. Bone marrow-derived endothelial progenitor cells (EPCs) promote endothelial repair and contribute to ischemia-induced neovascularization. The human Tp53 gene harbors a common single-nucleotide polymorphism (SNP) at codon 72, which yields an arginine-to-proline amino-acidic substitution (Arg72Pro) that modulates the apoptotic activity of the p53 protein. Previously, we found that this SNP controls neuronal susceptibility to ischemia-induced apoptosis in vitro. Here, we evaluated the impact of the Tp53 Arg72Pro SNP on vascular repair and functional recovery after ICH. We first analyzed EPC mobilization and functional outcome based on the modified Rankin scale scores in a hospital-based cohort of 78 patients with non-traumatic ICH. Patients harboring the Pro allele of the Tp53 Arg72Pro SNP showed higher levels of circulating EPC-containing CD34(+) cells, EPC-mobilizing cytokines - vascular endothelial growth factor and stromal cell-derived factor-1α - and good functional outcome following ICH, when compared with the homozygous Arg allele patients, which is compatible with increased neovascularization. To assess directly whether Tp53 Arg72Pro SNP regulated neovascularization after ICH, we used the humanized Tp53 Arg72Pro knock-in mice, which were subjected to the collagenase-induced ICH. The brain endothelial cells of the Pro allele-carrying mice were highly resistant to ICH-mediated apoptosis, which facilitated cytokine-mediated EPC mobilization, cerebrovascular repair and functional recovery. However, these processes were not observed in the Arg allele-carrying mice. These results reveal that the Tp53 Arg72Pro SNP determines

  12. Isolation of pyroGlu-Gly-Arg-Phe-NH2 (Antho-RFamide), a neuropeptide from sea anemones

    DEFF Research Database (Denmark)

    Grimmelikhuijzen, C J; Ebbesen, Ditte Graff

    1986-01-01

    elegantissima. Three different methods established that the structure of the Anthopleura RFamide peptide (Antho-RFamide) is pyroGlu-Gly-Arg-Phe-NH2. Comparison of synthetic and natural Antho-RFamide and their enzymatic breakdown products on six different HPLC columns confirmed the structure of the sea anemone......A radioimmunoassay has been developed for peptides containing the carboxyl-terminal sequence Arg-Phe-NH2 (RFamide). Using this radioimmunoassay and applying cation-exchange chromatography and HPLC, we have isolated an RFamide peptide from acetic acid extracts of the sea anemone Anthopleura...

  13. As formações em bolas argênticas do sistema nervoso simpático

    Directory of Open Access Journals (Sweden)

    José Fernandez

    1957-09-01

    Full Text Available Foram estudados, histològicamente, gânglios simpáticos provenientes de 40 pacientes portadores de enfermidades variadas e 40 gânglios tidos como normais, para pesquisar as formações cm bolas argênticas. As impregnações foram feitas pelos métodos de Cajal, Arteta, Castro, Gros, Bielschowsky e Agduhr. Foi verificado que a presença de bolas argênticas é mais constante nas doenças circulatórias periféricas; nestas afecções, foi verificada, também, a presença de um pigmento corável pelos tricrômios de Gomori e de Goldner e que, possìvelmente, tem o mesmo significado que os descritos por Fedorow 12 corados pelo vermelho neutro; tais pigmentos foram relacionados com as formações em bolas argênticas, levando em consideração os trabalhos experimentais dos autores da escola russa. A presença das formações em bolas argênticas, tanto nos casos normais como nos patológicos, é atribuída a uma hiperfunção, por excitação, do simpático, por serem mais numerosas nos casos de transtornos periféricos. Foi observado que os neurônios em íntimo contacto com as formações em bolas argênticas, apresentam sempre alterações que vão desde o grau mais leve (tumefação do soma celular, retificação dos prolongamentos, neurofibrilas mais fortemente impregnadas, até o mais grave (hiperplasia e hipertrofia dos dendritos, degeneração retrátil, degeneração pigmentar. Foi notado que as terminações das fibras pré-ganglionares e as próprias fibras apresentam, muitas vêzes, alterações (hipertrofia, sinuosidade e grande poder de impregnação; as bolas argênticas foram observadas tanto no final dos dendritos como no das fibras pré-ganglionares. Foram encontradas formações de bolas argênticas em rosário, concluindo-se que elas têm a mesma significação que as alterações claviformes. Conclui-se, finalmente, que as bolas argênticas são produtos patológicos.

  14. A ergologia na Argélia The ergology in Algeria

    Directory of Open Access Journals (Sweden)

    Fyad Abderrahmane

    2011-01-01

    Full Text Available O depoimento revela aspectos das relações de trabalho, focalizando principalmente nos direitos do trabalho de proteção aos riscos e à saúde. Além disso, apresenta elementos históricos da colaboração entre professores-pesquisadores da Universidade de Oran, na Argélia, e professores-pesquisadores do Departamento de Ergologia da Universidade de Provence, na França, para a implementação de projetos e formação de médicos do trabalho.The testimony reveals aspects of labor relations, focusing primarily on labor rights protection and health risks. Maneover, it presents historical elements of collaboration between teachers and researchers at the Univesity of Oran, Algeria, and research faculty of the Departement of Ergology University of Provence, France for project implementation and training of occupational physicians.

  15. Arg-Phe-amide-like peptides in the primitive nervous systems of coelenterates

    DEFF Research Database (Denmark)

    Grimmelikhuijzen, C J; Ebbesen, Ditte Graff

    1985-01-01

    By using immunocytochemistry and radioimmunoassays, several substances resembling vertebrate or invertebrate neuropeptides have been found in the nervous systems of coelenterates. The most abundant neuropeptides were those related to the molluscan neuropeptide Phe-Met-Arg-Phe-amide (FMRFamide). O...... with a RFamide antiserum and [J-125]-YFMRFamide as tracer, the RFamide-like peptide from sea anemones was isolated. After cation-exchange chromatography, gelfiltration and HPLC, this peptide was obtained in a pure form.......). Of antisera against different fragments of FMRFamide, those against RFamide were superior in recognizing the coelenterate peptide. Incubation of whole mounts with these RFamide antisera visualized the coelenterate nervous system in such a detail as has previously not been possible. By using a radioimmunoassay...

  16. Hemoglobin Willamette (β51Pro → Arg): Case Report and Literature Review

    Science.gov (United States)

    de Sousa Dias, Matheus Martins; Távora, Saymon Medeiros; de Galiza Neto, Gentil Claudino; de Souza, Jacqueline Holanda; da Silva, Herivaldo Ferreira

    2017-01-01

    We report a case of hemoglobin (Hb) Willamette (β51 Pro → Arg) in the Hematology Department of a tertiary hospital in Fortaleza, Northeast of Brazil. A literature review of the cases described in health sciences databases using as a descriptor Hb Willamette was performed, revealing 12 reported cases, of which only one presented with anemia. Herein, we describe a case of a female 29 years old, with hemoglobinopathy Willamette presenting clinically with anemia, having the lowest hemoglobin rate of the published cases. The relatives of the patient were evaluated andthe patient’s mother corresponded to the first description of the association between Hb Willamette and HbC. Among the hemoglobinopathies, hemoglobin Willamette is an extremely rare disease; therefore it is important to analyze its clinical and laboratory manifestations for accurate diagnosis and assessment of potential interactions with other genetic variants. PMID:28286631

  17. Hemoglobin Willamette (b51 Pro → Arg: case report and literature review

    Directory of Open Access Journals (Sweden)

    Orivaldo Alves Barbosa

    2017-03-01

    Full Text Available We report a case of hemoglobin (Hb Willamette (β51 Pro → Arg in the Hematology Department of a tertiary hospital in Fortaleza, Northeast of Brazil. A literature review of the cases described in health sciences databases using as a descriptor Hb Willamette was performed, revealing 12 reported cases, of which only one presented with anemia. Herein, we describe a case of a female 29 years old, with hemoglobinopathy Willamette presenting clinically with anemia, having the lowest hemoglobin rate of the published cases. The relatives of the patient were evaluated andthe patient’s mother corresponded to the first description of the association between Hb Willamette and HbC. Among the hemoglobinopathies, hemoglobin Willamette is an extremely rare disease; therefore it is important to analyze its clinical and laboratory manifestations for accurate diagnosis and assessment of potential interactions with other genetic variants.

  18. DIVERSITY OF THRIPS FAUNA (INSECTA: THYSANOPTERA IN PLUM ORCHARD FROM MORĂREŞTI-ARGES

    Directory of Open Access Journals (Sweden)

    Daniela Bărbuceanu

    2012-12-01

    Full Text Available The present study, conducted in the plums orchard of Morăreşti-Argeş at two plum varieties, has revealed a poor biodiversity. All of 10 species of thrips are polyphagous and belong to different trophic links: 8 species of phytophagous and 2 species, Aeolothrips intermedius and Haplothrips kurdjumovi are zoophagous. The structural parameters values indicate Haplothrips minutus as plum characteristic species, with the highest values of relative abundance and frequency in samples. However, this species has insignificant damaging in plum orchards. The low values of the structural indicators of the other species express their attachment to their characteristic trophic substratum, i.e. the herbaceous layer, so they only accidentally get on the plum branches, through anemochory. The Shannon-Weaver diversity index and equitability have low values, a situation which is typical of agro-ecosystems.

  19. Hyperactive Arg39Lys mutated mnemiopsin: implication of positively charged residue in chromophore binding cavity.

    Science.gov (United States)

    Mahdavi, Atiyeh; Sajedi, Reza H; Hosseinkhani, Saman; Taghdir, Majid

    2015-04-01

    Mnemiopsin, a Ca(2+)-regulated photoprotein isolated from Mnemiopsis leidyi, belongs to the family of ctenophore photoproteins. These proteins emit blue light from a chromophore, which is tightly but non-covalently bound in their central hydrophobic core that contains 21 conserved residues. In an effort to investigate the role of Arg39 (the sole charged residue in coelenterazine binding cavity of ctenophore photoproteins) in bioluminescence properties of these photoproteins, three mutated forms of mnemiopsin 1 (R39E, R39K and R39M) were constructed and characterized. The results indicate that while the luminescence activity of R39K mutated mnemiopsin has increased about nine fold compared to the wild type, R39M and R39E mutated mnemiopsins have entirely lost their activities. The most distinguished properties of R39K mutated photoprotein are its high activity, slow rate of luminescence decay and broad pH profile compared to the wild type. The complete loss of bioluminescence activity in mutated photoproteins with negatively charged and aliphatic residues (R39E and R39M, respectively) shows that the presence of a positively charged residue at this position is necessary. The results of spectroscopic studies, including CD, intrinsic and extrinsic fluorescence measurements and acrylamide quenching studies show that, while the substitutions lead to structural rigidity in R39E and R39M mutated mnemiopsins, structural flexibility is obvious in R39K mutated mnemiopsin. The presence of a more localized positive charge on ε-amino group of Lys compared to guanidinium group of Arg residue in close proximity to the choromophre might affect its fixation in the binding cavity and result in increased bioluminescence activity in this mutated photoprotein. It appears that the polarity and flexibility of positively charged residue at this position finely tunes the luminescence properties of ctenophore photoproteins.

  20. ARG (juegos de realidad alternativa. Contribuciones, limitaciones y potencialidades para la docencia universitaria

    Directory of Open Access Journals (Sweden)

    Teresa Piñeiro Otero

    2015-01-01

    Full Text Available La ludificación de la educación ha representado una oportunidad para fomentar la interacción, la motivación y la participación del alumnado. Los ARG (las siglas inglesas de juegos de realidad alternativa ofrecen una nueva herramienta altamente inmersiva que puede implementarse en el logro de los objetivos docentes. Uno de sus puntos fuertes consiste en la suma de esfuerzos y recursos (la llamada inteligencia colectiva aplicada a la resolución de problemas. A esto se añade su combinación de plataformas en los entornos online y offline, lo que favorece el «realismo» de la experiencia. En este sentido, el presente trabajo pretende condensar las potencialidades, limitaciones y retos de los ARG al servicio de la educación universitaria. Basándose, a nivel metodológico, en la elaboración de un corpus teórico relevante y adecuado, analiza el potencial educativo de esta herramienta que, en ámbitos como el marketing o la comunicación corporativa ya ha despegado con éxito, pero que en el área educativa todavía no había sido abordada en profundidad. Recopila, además, ejemplos satisfactorios que se han desarrollado en diversas disciplinas académicas en otros países y que no resultan fácilmente localizables. Se concluye que, dados los antecedentes, potencialidades y análisis expuesto, debe valorarse la posibilidad de incorporar los juegos de realidad alternativa a la práctica de la docencia universitaria en el marco de una estrategia educativa que determine sus objetivos y sistema de evaluación más adecuado.

  1. The chimeric peptide [Lys(-2)-Arg(-1)]-sarafotoxin-S6b, composed of the endothelin pro-sequence and sarafotoxin, retains the salt-bridge staple between Arg(-1) and Asp8 previously observed in [Lys(-2)-Arg(-1)]-endothelin. Implications of this salt-bridge in the contractile activity and the oxidative folding reaction.

    Science.gov (United States)

    Aumelas, A; Chiche, L; Kubo, S; Chino, N; Watanabe, T X; Kobayashi, Y

    1999-12-01

    The chimeric peptide [Lys(-2)-Arg(-1)]-sarafotoxin-S6b (KR-SRTb) designed from the Lys-2-Arg-1 dipeptide of the endothelin pro-sequence and the sarafotoxin-S6b sequence was synthesized. Its contractile activity was found to be decreased markedly when compared with that of the parent SRTb. In contrast, the extension by the Lys-Arg dipeptide was found to increase the formation of the native disulfide isomer (82/18 versus 96/4) when the reaction was carried out in the presence of redox reagents. The solution structure of KR-SRTb was determined by NMR as a function of pH. In the carboxylic acid state, the structure consists of the cystine-stabilized alpha-helical motif, with the alpha-helical part spanning residues 9-15, and of an unstructured C-terminal tail. In the carboxylate state, the structure is characterized by a salt-bridge between Arg(-1) and Asp8, which we identified previously in the [Lys(-2)-Arg(-1)]-endothelin-1 peptide (KR-ET-1). The fact that this salt-bridge is commonly observed in KR-SRTb and KR-ET-1, despite the 33% sequence difference between the corresponding parental peptides, highlights the remarkable adaptability of the Lys-Arg extension for the formation of a special salt-bridge. As a consequence, this salt-bridge, which does not depend on either the 4-7 sequence of the loop or the C-terminal sequence, appears to be particularly well suited to improve the stability of the cystine-stabilized alpha-helical motif. Therefore, because of its high yield in the native disulfide arrangement and its high permissiveness for sequence mutation in the 4-7 loop, such a stabilized cystine-stabilized alpha-helical motif could be a valuable scaffold for the presentation of a library of constrained short peptides.

  2. Effects of endogenous nitric oxide induced by 5-fluorouracil and L-Arg on liver carcinoma in nude mice

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    AIM: To study the effects of endogeous nitric oxide induced by 5-fluorouracil (5-FU) and L-arginine (L-Arg)on the human liver carcinoma model in nude mice.METHODS: The human liver carcinoma model in nude mice was established with BEL-7402 cells and normal saline (NS), 5-FU and 5-FU + L-Arg injected intraperitoneally. The tumor size was measured. The necrotic degree and range were observed under microscope. The apoptosis of cancer cell was detected by turmina deoxynucleotidyl transferanse mediated dUTP nick end labeling (TUNEL) method. Immunohistochemical method was performed to determine the expression of iNOS, P16, BAX. The chemical colorimetry was used to test the activity and nitrate reductase method was adopted to test the concentration of nitric oxide (NO) in the tumor tissue. The BI2000 pathological image analyzer was used to analyze the result of immunohistochemistry.RESULTS: 5-FU combined with L-Arg could inhibit the tumor growth apparently. In NS, 5-FU and 5-FU+L-Arg groups, the changes of tumor volumes were 257.978 ± 59.0, 172.232 ± 66.0 and 91.523 ± 26.7 mm3,respectively (P < 0.05 5-FU vs 5-FU + L-Arg group;P < 0.05 NS vs 5-FU + L-Arg group; P < 0.05, NS vs 5-FU group).The necrotic range and apoptosis index were significantly increased after the drug injection. The necrotic range was biggest in 5-FU + L-Arg group (x2 = 15.963, P < 0.05).The apoptosis indexes were as follows: NS, 17.4% ± 6.19%; 5-FU, 31.3% ± 12.3%; and 5-FU + L-Arg, 46% ± 15.24% (P < 0.05, 5-FU vs 5-FU + L-Arg; P < 0.05, NS vs 5-FU + L-Arg; P < 0.05, NS vs 5-FU). The expression and activity of iNOS were increased in the tumor tissue.The concentration of NO was also increased. F of optical density of iNOS, iNOS activity and NO concentration are 31.693, 21.949, and 33.909, respectively, P < 0.05. The concentration of NO was related to the expression of P16 and BAX. The correlation coefficient was 0.764 and 0.554.CONCLUSION: 5-FU combined with L-Arg can inhibit the

  3. An "instant gene bank" method for heterologous gene cloning: complementation of two Aspergillus nidulans mutants with Gaeumannomyces graminis DNA.

    Science.gov (United States)

    Bowyer, P; Osbourn, A E; Daniels, M J

    1994-02-01

    We present a novel technique for gene cloning by complementation of mutations in Aspergillus nidulans with DNA from a heterologous organism, Gaeumannomyces graminis. This technique bypasses the time-consuming and difficult construction of gene libraries, making it both rapid and simple. The method relies on recombination between a fungal replicating vector pHELP1 and linear G. graminis genomic DNA during co-transformation. We were able to complement two out of seven A. nidulans mutants tested and to rescue transforming DNA from both in Escherichia coli. Complementation of the A. nidulans argB mutation resulted from integration of 8-10 kb segments of G. graminis DNA into pHELP1. The complementation of the A. nidulans pyrG mutation resulted from a complex rearrangement. Complementing DNA was shown to originate from G. graminis, and was capable of retransforming the original mutants to give the expected phenotype.

  4. Crystal structure of native and a mutant of Lampyris turkestanicus luciferase implicate in bioluminescence color shift.

    Science.gov (United States)

    Kheirabadi, Mitra; Sharafian, Zohreh; Naderi-Manesh, Hossein; Heineman, Udo; Gohlke, Ulrich; Hosseinkhani, Saman

    2013-12-01

    Firefly bioluminescence reaction in the presence of Mg(2+), ATP and molecular oxygen is carried out by luciferase. The luciferase structure alterations or modifications of assay conditions determine the bioluminescence color of firefly luciferase. Among different beetle luciferases, Phrixothrix hirtus railroad worm emits either yellow or red bioluminescence color. Sequence alignment analysis shows that the red-emitter luciferase from Phrixothrix hirtus has an additional arginine residue at 353 that is absent in other firefly luciferases. It was reported that insertion of Arg in an important flexible loop350-359 showed changes in bioluminescence color from green to red and the optimum temperature activity was also increased. To explain the color tuning mechanism of firefly luciferase, the structure of native and a mutant (E354R/356R/H431Y) of Lampyris turkestanicus luciferase is determined at 2.7Å and 2.2Å resolutions, respectively. The comparison of structure of both types of Lampyris turkestanicus luciferases reveals that the conformation of this flexible loop is significantly changed by addition of two Arg in this region. Moreover, its surface accessibility is affected considerably and some ionic bonds are made by addition of two positive charge residues. Furthermore, we noticed that the hydrogen bonding pattern of His431 with the flexible loop is changed by replacing this residue with Tyr at this position. Juxtaposition of a flexible loop (residues 351-359) in firefly luciferase and corresponding ionic and hydrogen bonds are essential for color emission.

  5. Properties of normal and mutant recombinant human ketohexokinases and implications for the pathogenesis of essential fructosuria.

    Science.gov (United States)

    Asipu, Aruna; Hayward, Bruce E; O'Reilly, John; Bonthron, David T

    2003-09-01

    Alternative splicing of the ketohexokinase (fructokinase) gene generates a "central" predominantly hepatic isoform (ketohexokinase-C) and a more widely distributed ketohexokinase-A. Only the abundant hepatic isoform is known to possess activity, and no function is defined for the lower levels of ketohexokinase-A in peripheral tissues. Hepatic ketohexokinase deficiency causes the benign disorder essential fructosuria. The molecular basis of this has been defined in one family (compound heterozygosity for mutations Gly40Arg and Ala43Thr). Here we show that both ketohexokinase isoforms are indeed active. Ketohexokinase-A has much poorer substrate affinity than ketohexokinase-C for fructose but is considerably more thermostable. The Gly40Arg mutation seems null, rendering both ketohexokinase-A and ketohexokinase-C inactive and largely insoluble. The Ala43Thr mutant retains activity, but this mutation decreases the thermal stability of both ketohexokinase-A and ketohexokinase-C. At physiologic temperature, this results in significant loss of ketohexokinase-C activity but not of ketohexokinase-A. Affected individuals who carry both mutations therefore probably have a selective deficiency of hepatic ketohexokinase, with peripheral ketohexokinase-A being preserved. These findings raise the possibility that ketohexokinase-A serves an unknown physiologic function that remains intact in essential fructosuria. Further mutation analysis in this rare disorder could illuminate the question of whether ketohexokinase-A activity is, unlike that of ketohexokinase-C, physiologically indispensable.

  6. Zona Incerta: o ARG como meio de construção colaborativa de conteúdo

    Directory of Open Access Journals (Sweden)

    Gabriel Costa Correia

    2011-05-01

    Full Text Available O presente artigo discorrerá sobre um estudo de caso realizado sobre o Alternate Reality Game (ARG Zona Incerta, realizado por membros da equipe da revista Superinteressante, publicação da Editora Abril, como estratégia de marketing alternativa para o produto Guaraná Antarctica, da empresa de bebidas AMBEV.

  7. Arg9 facilitates the translocation and downstream signal inhibition of an anti-HER2 single chain antibody

    Directory of Open Access Journals (Sweden)

    Hu Yi

    2012-07-01

    Full Text Available Abstract Background HER2 plays a critical role in the pathogenesis of many cancers and is linked to poor prognosis or cancer metastases. Monoclonal antibodies, such as Herceptin against HER2-overexpressing cancers, have showed satisfactory clinical therapeutic effect. However, they have difficulty to surmount obstacles to enter cells or blood–brain barrier. Results In this study, a cell-penetrating peptide Arg9 was linked to the C-terminus of anti-HER2 single chain antibody (MIL5scFv. Flow cytometry, confocal microscopy and electron microscopy analysis all revealed that Arg9 peptide facilitated the penetration of MIL5scFv into HER2-negative cell line NIH3T3 and orientate in mitochondria. More interestingly, Western blot assay showed the potential enhanced bioactivity of MIL5scFv-Arg9 in HER2+ cell line SKOV3, indicating that Arg9 could help large molecules (e.g. antibody to penetrate into cells and therefore enhance its anti-neoplastic function. Conclusions Our work represented an attractive by preliminary strategy to enhance the therapeutic effect of existing antibodies by entering cells easier, or more desirable, surmounting the physical barriers, especially in hard-to-reach cancers such as brain metastases cases.

  8. Functional assignment of Glu386 and Arg388 in the active site of l-galactono-¿-lactone dehydrogenase

    NARCIS (Netherlands)

    Leferink, N.G.H.; Jose, M.D.F.; Berg, van den W.A.M.; Berkel, van W.J.H.

    2009-01-01

    The flavoenzyme l-galactono-¿-lactone dehydrogenase (GALDH) catalyzes the terminal step of vitamin C biosynthesis in plants. Little is known about the catalytic mechanism of GALDH and related aldonolactone oxidoreductases. Here we identified an essential Glu–Arg pair in the active site of GALDH from

  9. ArgR and AhrC are both required for regulation of arginine metabolism in Lactococcus lactis

    NARCIS (Netherlands)

    Larsen, R; Buist, G; Kuipers, OP; Kok, J

    2004-01-01

    The DNA binding proteins ArgR and AhrC are essential for regulation of arginine metabolism in Escherichia Coli and Bacillus subtilis, respectively. A unique property of these regulators is that they form hexameric protein complexes, mediating repression of arginine biosynthetic pathways as well as a

  10. High constitutive activity of a virus-encoded seven transmembrane receptor in the absence of the conserved DRY motif (Asp-Arg-Tyr) in transmembrane helix 3

    DEFF Research Database (Denmark)

    Rosenkilde, Mette M; Kledal, Thomas N; Schwartz, Thue W

    2005-01-01

    The highly conserved Arg in the so-called DRY motif (Asp-Arg-Tyr) at the intracellular end of transmembrane helix 3 is in general considered as an essential residue for G protein coupling in rhodopsin-like seven transmembrane (7TM) receptors. In the open reading frame 74 (ORF74) receptor encoded ...

  11. Effects of Arc/Arg3.1 gene deletion on rhythmic synchronization of hippocampal CA1 neurons during locomotor activity and sleep.

    NARCIS (Netherlands)

    Malkki, H.A.I.; Mertens, P.E.C.; Lankelma, J.V.; Vinck, M.; van Schalkwijk, F.J.; van Mourik-Donga, L.B.; Battaglia, F.P.; Mahlke, C.; Kuhl, D.; Pennartz, C.M.A.

    2016-01-01

    The activity-regulated cytoskeletal-associated protein/activity regulated gene (Arc/Arg3.1) is crucial for long-term synaptic plasticity and memory formation. However, the neurophysiological substrates of memory deficits occurring in the absence of Arc/Arg3.1 are unknown. We compared hippocampal CA1

  12. The association between changes in synovial fluid levels of ARGS-aggrecan fragments, progression of radiographic osteoarthritis and self-reported outcomes: a cohort study

    DEFF Research Database (Denmark)

    Larsson, S; Englund, M; Struglics, A;

    2012-01-01

    OBJECTIVE: To investigate whether change in concentrations over time of aggrecanase generated ARGS-aggrecan in synovial fluid (SF ARGS) associates with progression of radiographic knee osteoarthritis (OA) and patient-reported outcome in subjects with previous meniscectomy. METHODS: We studied 141...

  13. Effects of Arc/Arg3.1 gene deletion on rhythmic synchronization of hippocampal CA1 neurons during locomotor activity and sleep.

    NARCIS (Netherlands)

    Malkki, H.A.I.; Mertens, P.E.C.; Lankelma, J.V.; Vinck, M.; van Schalkwijk, F.J.; van Mourik-Donga, L.B.; Battaglia, F.P.; Mahlke, C.; Kuhl, D.; Pennartz, C.M.A.

    2016-01-01

    The activity-regulated cytoskeletal-associated protein/activity regulated gene (Arc/Arg3.1) is crucial for long-term synaptic plasticity and memory formation. However, the neurophysiological substrates of memory deficits occurring in the absence of Arc/Arg3.1 are unknown. We compared hippocampal CA1

  14. Lys-[Leu8,des-Arg9]-bradykinin blocks lipopolysaccharide-induced SHR aorta hyperpolarization by inhibition of Ca(++)- and ATP-dependent K+ channels.

    Science.gov (United States)

    Farias, Nelson C; Feres, Teresa; Paiva, Antonio C M; Paiva, Therezinha B

    2004-09-13

    The mediators involved in the hyperpolarizing effects of lipopolysaccharide and of the bradykinin B1 receptor agonist des-Arg9-bradykinin on the rat aorta were investigated by comparing the responses of aortic rings of spontaneously hypertensive and normotensive Wistar rats. Endothelized rings from hypertensive rats were hyperpolarized by des-Arg9-bradykinin and lipopolysaccharide, whereas de-endothelized rings responded to lipopolysaccharide but not to des-Arg9-bradykinin. In endothelized preparations, the responses to des-Arg9-bradykinin were inhibited by Nomega-nitro-L-arginine and iberiotoxin. De-endothelized ring responses to lipopolysaccharide were inhibited by iberiotoxin, glibenclamide and B1 antagonist Lys-[Leu8,des-Arg9]-bradykinin. This antagonist also inhibited hyperpolarization by des-Arg9-bradykinin and by the á2-adrenoceptor agonist, brimonidine. Our results indicate that Ca(2+)-sensitive K+ channels are the final mediators of the responses to des-Arg9-bradykinin, whereas both Ca(2+)- and ATP-sensitive K+ channels mediate the responses to lipopolysaccharide. The inhibitory effects of Lys-[Leu8,des-Arg9]-bradykinin is due to a direct action on Ca(2+)- and ATP-sensitive potassium channels.

  15. Isolation of the neuropeptide less than Glu-Trp-Leu-Lys-Gly-Arg-Phe-NH2 (Pol-RFamide II) from the hydromedusa Polyorchis penicillatus

    DEFF Research Database (Denmark)

    Grimmelikhuijzen, C J; Rinehart, K L; Spencer, A N

    1992-01-01

    Using a radioimmunoassay for the sequence Arg-Phe-NH2 (RFamide), we have isolated the peptide less than Glu-Trp-Leu-Lys-Gly-Arg-Phe-NH2 (Pol-RFamide II) from acetic acid extracts of the hydromedusa Polyorchis penicillatus. This peptide is a neuropeptide and constitutes a peptide family together w...

  16. Bacterial mutants for enhanced succinate production

    NARCIS (Netherlands)

    Baart, G.J.E.; Beauprez, J.J.R.; Foulquie, M.M.R.; Heijnen, J.J.; Maertens, J.

    2010-01-01

    The present invention relates to a method for obtaining enhanced metabolite production in micro-organisms, and to mutants and/or transformants obtained with said method. More particularly, it relates to bacterial mutants and/or transformants for enhanced succinate production, especially mutants and/

  17. Problem-Solving Test: Tryptophan Operon Mutants

    Science.gov (United States)

    Szeberenyi, Jozsef

    2010-01-01

    This paper presents a problem-solving test that deals with the regulation of the "trp" operon of "Escherichia coli." Two mutants of this operon are described: in mutant A, the operator region of the operon carries a point mutation so that it is unable to carry out its function; mutant B expresses a "trp" repressor protein unable to bind…

  18. Lamin A/C mutants disturb sumo1 localization and sumoylation in vitro and in vivo.

    Directory of Open Access Journals (Sweden)

    Émilie Boudreau

    Full Text Available A-type lamins A and C are nuclear intermediate filament proteins in which mutations have been implicated in multiple disease phenotypes commonly known as laminopathies. A few studies have implicated sumoylation in the regulation of A-type lamins. Sumoylation is a post-translational protein modification that regulates a wide range of cellular processes through the attachment of small ubiquitin-related modifier (sumo to various substrates. Here we showed that laminopathy mutants result in the mislocalization of sumo1 both in vitro (C2C12 cells overexpressing mutant lamins A and C and in vivo (primary myoblasts and myopathic muscle tissue from the Lmna(H222P/H222P mouse model. In C2C12 cells, we showed that the trapping of sumo1 in p.Asp192Gly, p.Gln353Lys, and p.Arg386Lys aggregates of lamin A/C correlated with an increased steady-state level of sumoylation. However, lamin A and C did not appear to be modified by sumo1. Our results suggest that mutant lamin A/C alters the dynamics of sumo1 and thus misregulation of sumoylation may be contributing to disease progression in laminopathies.

  19. Novel NCC mutants and functional analysis in a new cohort of patients with Gitelman syndrome.

    Science.gov (United States)

    Glaudemans, Bob; Yntema, Helger G; San-Cristobal, Pedro; Schoots, Jeroen; Pfundt, Rolph; Kamsteeg, Erik-J; Bindels, René J; Knoers, Nine V A M; Hoenderop, Joost G; Hoefsloot, Lies H

    2012-03-01

    Gitelman syndrome (GS) is an autosomal recessive disorder characterized by hypokalemic metabolic alkalosis in conjunction with significant hypomagnesemia and hypocalciuria. The GS phenotype is caused by mutations in the solute carrier family 12, member 3 (SLC12A3) gene that encodes the thiazide-sensitive NaCl cotransporter (NCC). We analyzed DNA samples of 163 patients with a clinical suspicion of GS by direct sequencing of all 26 exons of the SLC12A3 gene. In total, 114 different mutations were identified, 31 of which have not been reported before. These novel variants include 3 deletions, 18 missense, 6 splice site and 4 nonsense mutations. We selected seven missense mutations to investigate their effect on NCC activity and plasma membrane localization by using the Xenopus laevis oocyte expression system. The Thr392Ile mutant did not display transport activity (probably class 2 mutation), while the Asn442Ser and Gln1030Arg NCC mutants showed decreased plasma membrane localization and consequently function, likely due to impaired trafficking (class 3 mutation). Even though the NaCl uptake was hampered for NCC mutants Glu121Asp, Pro751Leu, Ser475Cys and Tyr489His, the transporters reached the plasma membrane (class 4 mutation), suggesting an effect on NCC regulation or ion affinity. The present study shows the identification of 38 novel mutations in the SLC12A3 gene and provides insight into the mechanisms that regulate NCC.

  20. Analysis of Escherichia coli nicotinate mononucleotide adenylyltransferase mutants in vivo and in vitro

    Directory of Open Access Journals (Sweden)

    Rydén-Aulin Monica

    2005-09-01

    Full Text Available Abstract Background Adenylation of nicotinate mononucleotide to nicotinate adenine dinucleotide is the penultimate step in NAD+ synthesis. In Escherichia coli, the enzyme nicotinate mononucleotide adenylyltransferase is encoded by the nadD gene. We have earlier made an initial characterization in vivo of two mutant enzymes, NadD72 and NadD74. Strains with either mutation have decreased intracellular levels of NAD+, especially for one of the alleles, nadD72. Results In this study these two mutant proteins have been further characterized together with ten new mutant variants. Of the, in total, twelve mutations four are in a conserved motif in the C-terminus and eight are in the active site. We have tested the activity of the enzymes in vitro and their effect on the growth phenotype in vivo. There is a very good correlation between the two data sets. Conclusion The mutations in the C-terminus did not reveal any function for the conserved motif. On the other hand, our data has lead us to assign amino acid residues His-19, Arg-46 and Asp-109 to the active site. We have also shown that the nadD gene is essential for growth in E. coli.

  1. Dual receptor-targeting ⁹⁹mTc-labeled Arg-Gly-Asp-conjugated Alpha-Melanocyte stimulating hormone hybrid peptides for human melanoma imaging.

    Science.gov (United States)

    Xu, Jingli; Yang, Jianquan; Miao, Yubin

    2015-04-01

    The aim of this study was to examine whether the substitution of the Lys linker with the aminooctanoic acid (Aoc) and polyethylene glycol (PEG) linker could substantially decrease the non-specific renal uptake of (99m)Tc-labeled Arg-Gly-Asp-conjugated α-melanocyte stimulating hormone (α-MSH) hybrid peptides. The RGD motif {Arg-Gly-Asp-DTyr-Asp} was coupled to [Cys(3,4,10), D-Phe(7), Arg(11)]α-MSH₃₋₁₃ via the Aoc or PEG₂ linker to generate RGD-Aoc-(Arg(11))CCMSH and RGD-PEG-(Arg(11))CCMSH. The biodistribution results of (99m)Tc-RGD-Aoc-(Arg(11))CCMSH and (99m)Tc-RGD-PEG₂-(Arg(11))CCMSH were examined in M21 human melanoma-xenografted nude mice. The substitution of Lys linker with Aoc and PEG₂ linker significantly reduced the renal uptake of (99m)Tc-RGD-Aoc-(Arg(11))CCMSH and (99m)Tc-RGD-PEG₂-(Arg(11))CCMSH by 58% and 63% at 2h post-injection. The renal uptake of (99m)Tc-RGD-Aoc-(Arg(11))CCMSH and (99m)Tc-RGD-PEG₂-(Arg(11))CCMSH was 27.93 ± 3.98 and 22.01 ± 9.89% ID/g at 2 h post-injection. (99m)Tc-RGD-Aoc-(Arg(11))CCMSH displayed higher tumor uptake than (99m)Tc-RGD-PEG₂-(Arg(11))CCMSH (2.35 ± 0.12 vs. 1.71 ± 0.25% ID/g at 2 h post-injection). The M21 human melanoma lesions could be clearly visualized by SPECT/CT using (99m)Tc-RGD-Aoc-(Arg(11))CCMSH as an imaging probe. The favorable effect of Aoc and PEG₂ linker in reducing the renal uptake provided a new insight into the design of novel dual receptor-targeting radiolabeled peptides. Copyright © 2014 Elsevier Inc. All rights reserved.

  2. Missense mutations in the gene encoding prothrombin corresponding to Arg596 cause antithrombin resistance and thrombomodulin resistance.

    Science.gov (United States)

    Takagi, Yuki; Murata, Moe; Kozuka, Toshihiro; Nakata, Yukiko; Hasebe, Ryo; Tamura, Shogo; Takagi, Akira; Matsushita, Tadashi; Saito, Hidehiko; Kojima, Tetsuhito

    2016-11-30

    Antithrombin (AT) and thrombomodulin (TM) play important roles in the process of natural anticoagulation in vivo. Recently, we reported that the prothrombin Yukuhashi mutation (p.Arg596Leu) was associated with AT and TM resistance-related thrombophilia. To assess the AT and TM resistances associated with other missense mutations by single base substitution in the Arg596 codon, we generated recombinant variants (596Gln, 596Trp, 596Gly, and 596Pro) and investigated the effects on AT and TM anticoagulant functions. All variants except 596Pro were secreted in amounts comparable to that of the wild-type but exhibited variable procoagulant activities. After a 30-minute inactivation by AT, the relative residual activity of wild-type thrombin decreased to 15 ± 4.0 %, in contrast to values of all variants were maintained at above 80 %. The thrombin-AT complex formation, as determined by enzyme-linked immunosorbent assay, was reduced with all tested variants in the presence and absence of heparin. In the presence of soluble TM (sTM), the relative fibrinogen clotting activity of wild-type thrombin decreased to 16 ± 0.12 %, whereas that of tested variants was 37 %-56 %. In a surface plasmon resonance assay, missense Arg596 mutations reduced thrombin-TM affinity to an extent similar to the reduction of fibrinogen clotting inhibition. In the presence of sTM or cultured endothelial-like cells, APC generation was enhanced differently by variant thrombins in a thrombin-TM affinity-dependent manner. These data indicate that prothrombin Arg596 missense mutations lead to AT and TM resistance in the variant thrombins and suggest that prothrombin Arg596 is important for AT- and TM-mediated anticoagulation.

  3. Association analysis of the beta-3 adrenergic receptor Trp64Arg (rs4994) polymorphism with urate and gout.

    Science.gov (United States)

    Fatima, Tahzeeb; Altaf, Sara; Phipps-Green, Amanda; Topless, Ruth; Flynn, Tanya J; Stamp, Lisa K; Dalbeth, Nicola; Merriman, Tony R

    2016-02-01

    The Arg64 allele of variant rs4994 (Trp64Arg) in the β3-adrenergic receptor gene has been associated with increased serum urate and risk of gout. Our objective was to investigate the relationship of rs4994 with serum urate and gout in New Zealand European, Māori and Pacific subjects. A total of 1730 clinically ascertained gout cases and 2145 controls were genotyped for rs4994 by Taqman(®). Māori and Pacific subjects were subdivided into Eastern Polynesian (EP) and Western Polynesian (WP) sample sets. Publicly available genotype data from the Atherosclerosis Risk in Communities Study and the Framingham Heart Study were utilized for serum urate association analysis. Multivariate logistic and linear regression adjusted for potential confounders was carried out using R version 2.15.2. No significant association of the minor Arg64 (G) allele of rs4994 with gout was found in the combined Polynesian cohorts (OR = 0.98, P = 0.88), although there was evidence, after adjustment for renal disease, for association in both the WP (OR = 0.53, P = 0.03) and the lower Polynesian ancestry EP sample sets (OR = 1.86, P = 0.05). There was no evidence for association with gout in the European sample set (OR = 1.11, P = 0.57). However, the Arg64 allele was positively associated with urate in the WP data set (β = 0.036, P = 0.004, P Corrected = 0.032). Association of the Arg64 variant with increased urate in the WP sample set was consistent with the previous literature, although the protective effect of this variant with gout in WP was inconsistent. This association provides an etiological link between metabolic syndrome components and urate homeostasis.

  4. Investigation of KIF6 Trp719Arg in a case-control study of myocardial infarction: a Costa Rican population.

    Directory of Open Access Journals (Sweden)

    Lance A Bare

    Full Text Available BACKGROUND AND METHODOLOGY: The 719Arg allele of KIF6 (rs20455 was associated with coronary events in Caucasian participants of five prospective studies. We investigated whether this KIF6 variant was associated with non-fatal myocardial infarction (MI in a case-control study of an admixed population from the Central Valley of Costa Rica. Genotypes of the KIF6 variant were determined for 4,134 men and women. Cases (1,987 had survived a first MI; controls (2,147 had no history of MI and were matched to cases by age, sex, and area of residence. We tested the association between the KIF6 719Arg allele and non-fatal MI by conditional logistic regression and adjusted for admixture of founder populations. PRINCIPAL FINDINGS: Compared with the reference Trp/Trp homozygotes, KIF6 719Arg carriers were not at significantly higher risk for non-fatal MI in this study after adjustment for traditional risk factors or admixture (OR= 1.12; 95%CI, 0.98-1.28. Heterozygotes of the KIF6 Trp719Arg variant were at increased risk of non-fatal MI: the adjusted odds ratio was 1.16 (95% confidence interval, 1.01-1.34, but this association would not be significant after a multiple testing correction. CONCLUSIONS/SIGNIFICANCE: We found that carriers of the KIF6 719Arg allele were not at increased risk of non-fatal MI in a case-control study of Costa Ricans living in the Central Valley of Costa Rica.

  5. Analysis of local structure of Arg10 domain in apo-α- lactalbumin with a polarity-sensitive arginine-specific fluorescent probe

    Institute of Scientific and Technical Information of China (English)

    2009-01-01

    The polarity-sensitive fluorescent probe, 3-(4-chloro-6-p-glyoxalphenoxy-1,3,5-triazinylamino)-7- (dimethylamino)-2-methylphenazine, was used to analyze the local structure of apo-α-lactalbumin by detecting the polarity and conformational changes of the arginine residue (Arg10) domain. The polarity of the Arg10 domain in both native and heat-denatured apo-α-lactalbumin was determined, which corresponds to a dielectric constant of 16, and the hydrophobic core near the Arg10 was found to be conservative for heating. Meanwhile, the effect of Ca2+ binding on the conformational changes of the Arg10 domain was studied, revealing that the hydrophobic core near the Arg10 is insensitive to the binding of Ca2+.

  6. Association of the beta3-adrenergic receptor Trp64Arg polymorphism with common metabolic traits: studies of 7605 middle-aged white people

    DEFF Research Database (Denmark)

    Gjesing, A. P.; Andersen, G; Borch-Johnsen, K;

    2008-01-01

    of type 2 diabetes and obesity in a relatively large, homogenous study population. METHODS: The Trp64Arg polymorphism was genotyped in 7605 Danish subjects using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Association was examined in case-control studies of obesity (1529...... cases and 6049 controls) and type 2 diabetes (1373 cases and 4742 controls) and quantitative trait analyses among 5822 individuals. Furthermore, the association of Trp64Arg with type 2 diabetes was examined in a meta-analysis. RESULTS: The Trp64Arg polymorphism was not associated with obesity. However...... (p=0.02). After sex stratification these associations were only present among women. Furthermore, the Arg-allele was borderline associated with type 2 diabetes in a meta-analysis of the present and 26 previous studies (p=0.06, OR1.27 (CI: 0.99-1.63)) (n=18891). CONCLUSION/INTERPRETATION: Trp64Arg...

  7. Technetium-99m-labeled Arg-Gly-Asp-conjugated alpha-melanocyte stimulating hormone hybrid peptides for human melanoma imaging

    Energy Technology Data Exchange (ETDEWEB)

    Yang Jianquan; Guo Haixun [College of Pharmacy, University of New Mexico, Albuquerque, NM 87131 (United States); Miao Yubin, E-mail: ymiao@salud.unm.ed [College of Pharmacy, University of New Mexico, Albuquerque, NM 87131 (United States); Cancer Research and Treatment Center, University of New Mexico, Albuquerque, NM 87131 (United States); Department of Dermatology, University of New Mexico, Albuquerque, NM 87131 (United States)

    2010-11-15

    Introduction: The purpose of this study was to examine whether {sup 99m}Tc-labeled Arg-Gly-Asp (RGD)-conjugated alpha-melanocyte stimulating hormone ({alpha}-MSH) hybrid peptide targeting both melanocortin-1 (MC1) and {alpha}{sub v{beta}3} integrin receptors was superior in melanoma targeting to {sup 99m}Tc-labeled {alpha}-MSH or RGD peptide targeting only the MC1 or {alpha}{sub v{beta}3} integrin receptor. Methods: RGD-Lys-(Arg{sup 11})CCMSH, RAD-Lys-(Arg{sup 11})CCMSH and RGD-Lys-(Arg{sup 11})CCMSHscramble were designed to target both MC1 and {alpha}{sub v{beta}3} integrin receptors, MC1 receptor only and {alpha}{sub v{beta}3} integrin receptor only, respectively. The MC1 or {alpha}{sub v{beta}3} integrin receptor binding affinities of three peptides were determined in M21 human melanoma cells. The melanoma targeting properties of {sup 99m}Tc-labeled RGD-Lys-(Arg{sup 11})CCMSH, RAD-Lys-(Arg{sup 11})CCMSH and RGD-Lys-(Arg{sup 11})CCMSHscramble were determined in M21 human melanoma-xenografted nude mice. Meanwhile, the melanoma uptake of {sup 99m}Tc-RGD-Lys-(Arg{sup 11})CCMSH was blocked with various non-radiolabeled peptides in M21 melanoma xenografts. Results: RGD-Lys-(Arg{sup 11})CCMSH displayed 2.0 and 403 nM binding affinities to both MC1 and {alpha}{sub v{beta}3} integrin receptors, whereas RAD-Lys-(Arg{sup 11})CCMSH or RGD-Lys-(Arg{sup 11})CCMSHscramble lost their {alpha}{sub v{beta}3} integrin receptor binding affinity by greater than 248-fold or MC1 receptor binding affinity by more than 100-fold, respectively. The melanoma uptake of {sup 99m}Tc-RGD-Lys-(Arg{sup 11})CCMSH was 2.49 and 2.24 times (P < .05) the melanoma uptakes of {sup 99m}Tc-RAD-Lys-(Arg{sup 11})CCMSH and {sup 99m}Tc-RGD-Lys-(Arg{sup 11})CCMSHscramble at 2 h post-injection, respectively. Either RGD or (Arg{sup 11})CCMSH peptide co-injection could block 42% and 57% of the tumor uptake of {sup 99m}Tc-RGD-Lys-(Arg{sup 11})CCMSH, whereas the coinjection of RGD+(Arg{sup 11})CCMSH peptide mixture

  8. Conformational control inhibition of the BCR-ABL1 tyrosine kinase, including the gatekeeper T315I mutant, by the switch-control inhibitor DCC-2036

    Science.gov (United States)

    Chan, Wayne W.; Wise, Scott C.; Kaufman, Michael D.; Ahn, Yu Mi; Ensinger, Carol L.; Haack, Torsten; Hood, Molly M.; Jones, Jennifer; Lord, John W.; Lu, Wei Ping; Miller, David; Patt, William C.; Smith, Bryan D.; Petillo, Peter A.; Rutkoski, Thomas J.; Telikepalli, Hanumaiah; Vogeti, Lakshminarayana; Yao, Tony; Chun, Lawrence; Clark, Robin; Evangelista, Peter; Gavrilescu, L. Cristina; Lazarides, Katherine; Zaleskas, Virginia M.; Stewart, Lance J.; Van Etten, Richard A.; Flynn, Daniel L.

    2011-01-01

    Summary Acquired resistance to ABL1 tyrosine kinase inhibitors (TKIs) through ABL1 kinase domain mutations, particularly the gatekeeper mutant T315I, is a significant problem for chronic myeloid leukemia (CML) patients. Using structure-based drug design, we developed compounds that bind to residues (Arg386/Glu282) ABL1 uses to switch between inactive and active conformations. The lead “switch-control” inhibitor, DCC-2036, potently inhibits both unphosphorylated and phosphorylated ABL1 by inducing a type II inactive conformation, and retains efficacy against the majority of clinically relevant CML resistance mutants, including T315I. DCC-2036 inhibits BCR-ABL1T315I-expressing cell lines, prolongs survival in mouse models of T315I-mutant CML and B-lymphoblastic leukemia, and inhibits primary patient leukemia cells expressing T315I in vitro and in vivo, supporting its clinical development in TKI-resistant Ph+ leukemia. PMID:21481795

  9. Contribution of Arg288 of Escherichia coli elongation factor Tu to translational functionality

    DEFF Research Database (Denmark)

    Rattenborg, Thomas; Nautrup-Pedersen, Gitte; Clark, Brian F. C.

    1997-01-01

    -type factor with regard to the association and dissociation of guanine nucleotides, and the intrinsic GTPase activities are unchanged. Furthermore, the mutants were as efficient as the wild-type factor in carrying out protein synthesis in vitro in the presence of an excess of aminoacyl-tRNA. However...

  10. ArgR is an essential local transcriptional regulator of the arcABC operon in Streptococcus suis and is crucial for biological fitness in an acidic environment.

    Science.gov (United States)

    Fulde, Marcus; Willenborg, Joerg; de Greeff, Astrid; Benga, Laurentiu; Smith, Hilde E; Valentin-Weigand, Peter; Goethe, Ralph

    2011-02-01

    Streptococcus suis is one of the most important pathogens in pigs and can also cause severe infections in humans. Despite its clinical relevance, very little is known about the factors that contribute to its virulence. Recently, we identified a new putative virulence factor in S. suis, the arginine deiminase system (ADS), an arginine catabolic enzyme system encoded by the arcABC operon, which enables S. suis to survive in an acidic environment. In this study, we focused on ArgR, an ADS-associated regulator belonging to the ArgR/AhrC arginine repressor family. Using an argR knockout strain we were able to show that ArgR is essential for arcABC operon expression and necessary for the biological fitness of S. suis. By cDNA expression microarray analyses and quantitative real-time RT-PCR we found that the arcABC operon is the only gene cluster regulated by ArgR, which is in contrast to the situation in many other bacteria. Reporter gene analysis with gfp under the control of the arcABC promoter demonstrated that ArgR is able to activate the arcABC promoter. Electrophoretic mobility shift assays with fragments of the arcABC promoter and recombinant ArgR, and chromatin immunoprecipitation with antibodies directed against ArgR, revealed that ArgR interacts with the arcABC promoter in vitro and in vivo by binding to a region from -147 to -72 bp upstream of the transcriptional start point. Overall, our results show that in S. suis, ArgR is an essential, system-specific transcriptional regulator of the ADS that interacts directly with the arcABC promoter in vivo.

  11. Effect of the selective pressure of sub-lethal level of heavy metals on the fate and distribution of ARGs in the catchment scale.

    Science.gov (United States)

    Xu, Yan; Xu, Jian; Mao, Daqing; Luo, Yi

    2017-01-01

    Our previous study demonstrated that high levels of antibiotic resistance genes (ARGs) in the Haihe River were directly attributed to the excessive use of antibiotics in animal agriculture. The antibiotic residues of the Xiangjiang River determined in this study were much lower than those of the Haihe River, but the relative abundance of 16 detected ARGs (sul1, sul2 and sul3, qepA, qnrA, qnrB, qnrD and qnrS, tetA, tetB, tetW, tetM, tetQ and tetO, ermB and ermC), were as high as the Haihe River particularly in the downstream of the Xiangjiang River which is close to the extensive metal mining. The ARGs discharged from the pharmaceutical wastewater treatment plant (PWWTP) are a major source of ARGs in the upstream of the Xiangjiang River. In the downstream, selective stress of heavy metals rather than source release had a significant influence on the distinct distribution pattern of ARGs. Some heavy metals showed a positive correlation with certain ARG subtypes. Additionally, there is a positive correlation between individual ARG subtypes and heavy metal resistance genes, suggesting that heavy metals may co select the ARGs on the same plasmid of antibiotic resistant bacteria. The co-selection mechanism between specific metal and antibiotic resistance was further confirmed by these isolations encoding the resistance genotypes to antibiotics and metals. To our knowledge, this is the first study on the fate and distribution of ARGs under the selective pressure exerted by heavy metals in the catchment scale. These results are beneficial to understand the fate, and to discern the contributors of ARGs from either the source release or the selective pressure by sub-lethal levels of environmental stressors during their transport on a river catchment scale. Copyright © 2016 Elsevier Ltd. All rights reserved.

  12. Impaired 8-Hydroxyguanine Repair Activity of MUTYH Variant p.Arg109Trp Found in a Japanese Patient with Early-Onset Colorectal Cancer

    Directory of Open Access Journals (Sweden)

    Kazuya Shinmura

    2014-01-01

    Full Text Available Purpose. The biallelic inactivation of the 8-hydroxyguanine repair gene MUTYH leads to MUTYH-associated polyposis (MAP, which is characterized by colorectal multiple polyps and carcinoma(s. However, only limited information regarding MAP in the Japanese population is presently available. Since early-onset colorectal cancer (CRC is a characteristic of MAP and might be caused by the inactivation of another 8-hydroxyguanine repair gene, OGG1, we investigated whether germline MUTYH and OGG1 mutations are involved in early-onset CRC in Japanese patients. Methods. Thirty-four Japanese patients with early-onset CRC were examined for germline MUTYH and OGG1 mutations using sequencing. Results. Biallelic pathogenic mutations were not found in any of the patients; however, a heterozygous p.Arg19*  MUTYH variant and a heterozygous p.Arg109Trp MUTYH variant were detected in one patient each. The p.Arg19* and p.Arg109Trp corresponded to p.Arg5* and p.Arg81Trp, respectively, in the type 2 nuclear-form protein. The defective DNA repair activity of p.Arg5* is apparent, while that of p.Arg81Trp has been demonstrated using DNA cleavage and supF forward mutation assays. Conclusion. These results suggest that biallelic MUTYH or OGG1 pathogenic mutations are rare in Japanese patients with early-onset CRC; however, the p.Arg19* and p.Arg109Trp MUTYH variants are associated with functional impairments.

  13. Nif- Hup- mutants of Rhizobium japonicum.

    Science.gov (United States)

    Moshiri, F; Stults, L; Novak, P; Maier, R J

    1983-01-01

    Two H2 uptake-negative (Hup-) Rhizobium japonicum mutants were obtained that also lacked symbiotic N2 fixation (acetylene reduction) activity. One of the mutants formed green nodules and was deficient in heme. Hydrogen oxidation activity in this mutant could be restored by the addition of heme plus ATP to crude extracts. Bacteroid extracts from the other mutant strain lacked hydrogenase activity and activity for both of the nitrogenase component proteins. Hup+ revertants of the mutant strains regained both H2 uptake ability and nitrogenase activity. Images PMID:6874648

  14. Identification of a Long Rice Spikelet Mutant

    Institute of Scientific and Technical Information of China (English)

    WU Xian-jun; WANG Bin; HAN Zan-ping; XIE Zhao-hui; MOU Chun-hong; WANG Xu-dong

    2004-01-01

    A spontaneously occurring rice (Oryza sativa L. ) mutant, characterized by homeotic conversion in glumes and stamens, was found in the progeny of a cross. The mutant showed long glumes and glumaceous lodicules and morphological transformation of stamens into pistils. Mutant florets consisted of 1 to 3 completely developed pistils, some pistilloid stamens with filaments, but tipped by bulged tissue and 0 to 3 stigmas. It seens that the mutant phenotype of the homeotic conversions in glumes and stamens is similar to that of the B loss-of-function mutants in Arabidopsis and Antirrhinum. The mutant is controlled by a single recessive gene as a segregation ratio of 3:1 (wild type to mutant plants) was observed in the F2 generation.

  15. Angiotensin I-converting enzyme Gln1069Arg mutation impairs trafficking to the cell surface resulting in selective denaturation of the C-domain.

    Directory of Open Access Journals (Sweden)

    Sergei M Danilov

    Full Text Available BACKGROUND: Angiotensin-converting enzyme (ACE; Kininase II; CD143 hydrolyzes small peptides such as angiotensin I, bradykinin, substance P, LH-RH and several others and thus plays a key role in blood pressure regulation and vascular remodeling. Complete absence of ACE in humans leads to renal tubular dysgenesis (RTD, a severe disorder of renal tubule development characterized by persistent fetal anuria and perinatal death. METHODOLOGY/PRINCIPAL FINDINGS: Patient with RTD in Lisbon, Portugal, maintained by peritoneal dialysis since birth, was found to have a homozygous substitution of Arg for Glu at position 1069 in the C-terminal domain of ACE (Q1069R resulting in absence of plasma ACE activity; both parents and a brother who are heterozygous carriers of this mutation had exactly half-normal plasma ACE activity compared to healthy individuals. We hypothesized that the Q1069R substitution impaired ACE trafficking to the cell surface and led to accumulation of catalytically inactive ACE in the cell cytoplasm. CHO cells expressing wild-type (WT vs. Q1069R-ACE demonstrated the mutant accumulates intracellularly and also that it is significantly degraded by intracellular proteases. Q1069R-ACE retained catalytic and immunological characteristics of WT-ACE N domain whereas it had 10-20% of the nativity of the WT-ACE C domain. A combination of chemical (sodium butyrate or pharmacological (ACE inhibitor chaperones with proteasome inhibitors (MG 132 or bortezomib significantly restored trafficking of Q1069R-ACE to the cell surface and increased ACE activity in the cell culture media 4-fold. CONCLUSIONS/SIGNIFICANCE: Homozygous Q1069R substitution results in an ACE trafficking and processing defect which can be rescued, at least in cell culture, by a combination of chaperones and proteasome inhibitors. Further studies are required to determine whether similar treatment of individuals with this ACE mutation would provide therapeutic benefits such as

  16. Three novel and the common Arg677Ter RP1 protein truncating mutations causing autosomal dominant retinitis pigmentosa in a Spanish population

    Directory of Open Access Journals (Sweden)

    Antiñolo Guillermo

    2006-04-01

    Full Text Available Abstract Background Retinitis pigmentosa (RP, a clinically and genetically heterogeneous group of retinal degeneration disorders affecting the photoreceptor cells, is one of the leading causes of genetic blindness. Mutations in the photoreceptor-specific gene RP1 account for 3–10% of cases of autosomal dominant RP (adRP. Most of these mutations are clustered in a 500 bp region of exon 4 of RP1. Methods Denaturing gradient gel electrophoresis (DGGE analysis and direct genomic sequencing were used to evaluate the 5' coding region of exon 4 of the RP1 gene for mutations in 150 unrelated index adRP patients. Ophthalmic and electrophysiological examination of RP patients and relatives according to pre-existing protocols were carried out. Results Three novel disease-causing mutations in RP1 were detected: Q686X, K705fsX712 and K722fsX737, predicting truncated proteins. One novel missense mutation, Thr752Met, was detected in one family but the mutation does not co-segregate in the family, thereby excluding this amino acid variation in the protein as a cause of the disease. We found the Arg677Ter mutation, previously reported in other populations, in two independent families, confirming that this mutation is also present in a Spanish population. Conclusion Most of the mutations reported in the RP1 gene associated with adRP are expected to encode mutant truncated proteins that are approximately one third or half of the size of wild type protein. Patients with mutations in RP1 showed mild RP with variability in phenotype severity. We also observed several cases of non-penetrant mutations.

  17. Monitoring and evaluation of the water quality of Budeasa Reservoir-Arges River, Romania.

    Science.gov (United States)

    Ion, Antoanela; Vladescu, Luminita; Badea, Irinel Adriana; Comanescu, Laura

    2016-09-01

    The purpose of this study was to monitor and record the specific characteristics and properties of the Arges River water in the Budeasa Reservoir (the principal water resources of municipal tap water of the big Romanian city Pitesti and surrounding area) for a period of 5 years (2005-2009). The monitored physical and chemical parameters were turbidity, pH, electrical conductivity, chemical oxygen demand, 5 days biochemical oxygen demand, free dissolved oxygen, nitrite, nitrate, ammonia nitrogen, chloride, total dissolved iron ions, sulfate, manganese, phosphate, total alkalinity, and total hardness. The results were discussed in correlation with the precipitation values during the study. Monthly and annual values of each parameter determined in the period January 2005-December 2009 were used as a basis for the classification of Budeasa Reservoir water, according to the European legislation, as well as for assessing its quality as a drinking water supply. Principal component analysis and Pearson correlation coefficients were used as statistical procedures in order to evaluate the data obtained during this study.

  18. The effect of climate on populations of grape moths in the vineyard Stefanesti-Arges

    Directory of Open Access Journals (Sweden)

    Daniela BĂRBUCEANU

    2005-05-01

    Full Text Available In the period 1998-2003, in the vineyard Stefanesti-Arges, observing was carried out, with the help of pheromone traps, of the population of grape moths, Eupoecilia ambiguella Hb. and Lobesia botrana Den et Schiff. The different climatic preferences of both species, establish that, in accordance with the yearly conditions, one of the two species is dominated by the other. The year 1998 is the only year when the percentages of captures of the two species are nearly: 57% - L. botrana and 42,89% - E. ambiguella, while in 1999 is the only year when E. ambiguella dominated in number the species L. botrana - 66,28%. For meeting this situation it needed two years of consecutives optimum climate for the activity of species E. ambiguella (like the year 1998 and 1999. In the period 2000-2003, the species Lobesia botrana was dominated by E. ambiguella, but as a result of very raised temperatures of summers of years 2000, 2002 and 2003, the population of L. botrana diminished very much and there was no need for fighting back treatments.

  19. Analgesic and Anti-Inflammatory Activities of Leaf Extract of Mallotus repandus (Willd. Muell. Arg.

    Directory of Open Access Journals (Sweden)

    Md. Mahadi Hasan

    2014-01-01

    Full Text Available In folk medicine Mallotus repandus (Willd. Muell. Arg. is used to treat muscle pain, itching, fever, rheumatic arthritis, snake bite, hepatitis, and liver cirrhosis. This study aimed to evaluate the antinociceptive as well as the anti-inflammatory activities of the methanol extract of leaf. The leaves were extracted with methanol following hot extraction and tested for the presence of phytochemical constituents. Analgesic and anti-inflammatory activities were evaluated using acetic acid induced writhing test, xylene induced ear edema, cotton pellet induced granuloma, and tail immersion methods at doses of 500, 1000, and 2000 mg/kg body weight. The presence of flavonoids, saponins, and tannins was identified in the extract. The extract exhibited considerable antinociceptive and anti-inflammatory activities against four classical models of pain. In acetic acid induced writhing, xylene induced ear edema, and cotton pellet granuloma models, the extract revealed dose dependent activity. Additionally, it increased latency time in tail immersion model. It can be concluded that M. repandus possesses significant antinociceptive potential. These findings suggest that this plant can be used as a potential source of new antinociceptive and anti-inflammatory candidates. The activity of methanol extract is most likely mediated through central and peripheral inhibitory mechanisms. This study justified the traditional use of leaf part of this plant.

  20. A novel fibrinogen mutation (γ Thr277Arg) causes hereditary hypofibrinogenemia in a Chinese family.

    Science.gov (United States)

    Zhu, Liqing; Wang, Mingshan; Xie, Haixiao; Jin, Yanhui; Yang, Lihong; Xu, Pengfei

    2013-09-01

    Congenital hypofibrinogenemia is a rare disorder caused by heterozygous mutations in one of the three fibrinogen genes--fibrinogen α-chain (FGA), fibrinogen β-chain (FGB) and fibrinogen γ-chain (FGG)--which code for the Aα, Bβ and γ chains, respectively. In this study, we identified a genetic defect in the FGG underlying the hypofibrinogenemia. The proposita had a prolonged blood clotting time (thrombin time 24.5 s, prothrombin time 16.8 s) and a low level of plasma fibrinogen (0.71 g/l by Clauss method and 0.79 g/l by immunoturbidimetry). DNA screening of the whole fibrinogen gene revealed a heterozygous GC mutation at nucleotide 7482 in her FGG gene. Her father and her half-brother are also heterozygous for this mutation. This mutation contributes to Thr277 → Arg in the γ chain of fibrinogen. To the best of our knowledge, this is the first report of such a mutation that is associated with hypofibrinogenemia.

  1. Anatomia da madeira de Phyllanthus sellowianus Müll. Arg. (Phyllanthaceae

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    Luciano Denardi

    2013-01-01

    Full Text Available The present study aimed to describe the wood anatomy of Phyllanthus sellowianus Müll. Arg.(Phyllanthaceae, as well as the research of anatomical characters associated with the flexibility of their stems, desirable characteristic for bioengineering works. Belonging to the group of reophyllous and with up to three meters in height, the species occurs naturally in the south of Brazil, Uruguay and Argentina. The exsicat and wood sample from a P. sellowianus, located in the municipality of Santa Maria, Rio Grande do Sul state, Brazi, were deposited in the University Federal de Santa Maria under the numbers HDCF 5587 e 5588. Standard technique was used to set the histological lamina of wood. For macerations used the method of Jeffrey. The Wood anatomy description followed the IAWA Committee (1989. The microscopic analysis revealed the presence vassels of small diameter (< 60µm, radial multiples of 2 to 4 units, and simple perforation plates, obliquely, axial parenchyma absent, heterogeneous rays, with 1-3 cells large, and gelatinous fibers. The absence of axial parenchyma and laticifer tubes is compatible to the Phyllanthaceae group. The presence of perforated ray cells is, probably, the main anatomic component for differentiation of P. sellowianus from the other species of the genera. The abundant presence of gelatinous fibers certainly is one of the main anatomical characteristics related to the high flexibility of its stem.

  2. The tetrapeptide Arg-Leu-Tyr-Glu inhibits VEGF-induced angiogenesis

    Energy Technology Data Exchange (ETDEWEB)

    Baek, Yi-Yong; Lee, Dong-Keon [Department of Molecular and Cellular Biochemistry, School of Medicine, Kangwon National University, Chuncheon, Gangwon-do, 200-702 (Korea, Republic of); So, Ju-Hoon; Kim, Cheol-Hee [Department of Biology, Chungnam National University, Daejeon, 305-764 (Korea, Republic of); Jeoung, Dooil [Department of Biochemistry, College of Natural Sciences, Kangwon National University, Chuncheon, Gangwon-do, 200-702 (Korea, Republic of); Lee, Hansoo [Department of Life Sciences, College of Natural Sciences, Kangwon National University, Chuncheon, Gangwon-do, 200-702 (Korea, Republic of); Choe, Jongseon [Department of Immunology, School of Medicine, Kangwon National University, Chuncheon, Gangwon-do, 200-702 (Korea, Republic of); Won, Moo-Ho [Department of Neurobiology, School of Medicine, Kangwon National University, Chuncheon, Gangwon-do, 200-702 (Korea, Republic of); Ha, Kwon-Soo [Department of Molecular and Cellular Biochemistry, School of Medicine, Kangwon National University, Chuncheon, Gangwon-do, 200-702 (Korea, Republic of); Kwon, Young-Guen [Department of Biochemistry, College of Life Science and Biotechnology, Yonsei University, Seoul, 120-752 (Korea, Republic of); Kim, Young-Myeong, E-mail: ymkim@kangwon.ac.kr [Department of Molecular and Cellular Biochemistry, School of Medicine, Kangwon National University, Chuncheon, Gangwon-do, 200-702 (Korea, Republic of)

    2015-08-07

    Kringle 5, derived from plasminogen, is highly capable of inhibiting angiogenesis. Here, we have designed and synthesized 10 tetrapeptides, based on the amino acid properties of the core tetrapeptide Lys-Leu-Tyr-Asp (KLYD) originating from anti-angiogenic kringle 5 of human plasminogen. Of these, Arg-Leu-Tyr-Glu (RLYE) effectively inhibited vascular endothelial growth factor (VEGF)-induced endothelial cell proliferation, migration and tube formation, with an IC{sub 50} of 0.06–0.08 nM, which was about ten-fold lower than that of the control peptide KLYD (0.79 nM), as well as suppressed developmental angiogenesis in a zebrafish model. Furthermore, this peptide effectively inhibited the cellular events that precede angiogenesis, such as ERK and eNOS phosphorylation and nitric oxide production, in endothelial cells stimulated with VEGF. Collectively, these data demonstrate that RLYE is a potent anti-angiogenic peptide that targets the VEGF signaling pathway. - Highlights: • The tetrapeptide RLYE inhibited VEGF-induced angiogenesis in vitro. • RLYE also suppressed neovascularization in a zebrafish model. • Its effect was correlated with inhibition of VEGF-induced ERK and eNOS activation. • RLYE may be used as a therapeutic drug for angiogenesis-related diseases.

  3. Arg333 and Arg334 in the COOH terminus of the human P2Y1 receptor are crucial for Gq coupling.

    Science.gov (United States)

    Ding, Zhongren; Tuluc, Florin; Bandivadekar, Kavita R; Zhang, Lili; Jin, Jianguo; Kunapuli, Satya P

    2005-03-01

    The P2Y(1) ADP receptor activates G(q) and causes increases in intracellular Ca(2+) concentration through stimulation of PLC. In this study, we investigated the role of the amino acid residues in the COOH terminus of the human P2Y(1) receptor in G(q) activation. Stimulation of Chinese hamster ovary (CHO-K1) cells stably expressing the wild-type human P2Y(1) receptor (P2Y(1)-WT cells), P2Y(1)-DeltaR340-L373, or P2Y(1)-DeltaD356-L373 with 2-methylthio-ADP (2-MeSADP) caused inositol phosphate production. In contrast, cells expressing P2Y(1)-DeltaT330-L373, a mutant lacking the entire COOH terminus, completely lost their response to 2-MeSADP. Similar data were obtained by using these cell lines and measuring Ca(2+) mobilization upon stimulation with 2-MeSADP, indicating that the 10 amino acids (330TFRRRLSRAT339) in the COOH terminus of the human P2Y(1) receptor are essential for G(q) coupling. Radioligand binding demonstrated that both the P2Y(1)-WT and P2Y(1)-DeltaT330-L373-expressing cells have almost equal binding of [(3)H]MRS2279, a P2Y(1) receptor antagonist, indicating that COOH-terminal truncation did not drastically affect the conformation of the receptor. CHO-K1 cells expressing a chimeric P2Y(12) receptor with the P2Y(1) COOH terminus failed to elicit G(q) functional responses, indicating that the P2Y(1) COOH terminus is essential but not sufficient for G(q) activation. Finally, cells expressing a double-mutant P2Y(1) receptor (R333A/R334A) in the conserved BBXXB region of the COOH terminus of the G(q)-activating P2Y receptors completely lost their functional ability to activate G(q). We conclude that the two arginine residues (R333R334) in the COOH terminus of the human P2Y(1) receptor are essential for G(q) coupling.

  4. Development and adaptation of the ELBIS Elbe river information system into a water quality information system of the ARGE-ELBE working group. Final report; Entwicklung und Anpassung des Elbeinformationssystems ELBIS fuer ein Gewaessergueteinformationssystem der ARGE-ELBE. Abschlussbericht

    Energy Technology Data Exchange (ETDEWEB)

    Frank, B.; Prange, A.

    2001-07-01

    The geo-information system ELBIS was installed in mid-November 2000 to provide information on the web on Elbe water quality and other information. It presents the current results of the national Elbe monitoring programme of the ARGE Elbe working group, as well as the results of the 'Elbe 2000' programme of the nineties. [German] Das Geo-Informationssystem ELBIS informiert seit November 2000 im Internet ueber die Wasserqualitaet und verwandte Elbe-Themen. Mittels einer geografischen Oberflaeche stellt es die fortlaufenden, aktuellen Ergebnisse der nationalen Elbe-Messprogramme der ARGE Elbe (Arbeitsgemeinschaft fuer die Reinhaltung der Elbe) dar sowie Forschungsdaten aus der 'Elbe 2000'-Schadstoffforschung der neunziger Jahre. Auf einfache und zeitgemaesse Weise macht ELBIS diese Ergebnisse der interessierten Oeffentlichkeit zugaenglich und bietet Fachleuten einen umfassenden Ueberblick ueber die Datenlage. (orig.)

  5. Development and adaptation of the ELBIS Elbe river information system into a water quality information system of the ARGE-ELBE working group. Final report; Entwicklung und Anpassung des Elbeinformationssystems ELBIS fuer ein Gewaessergueteinformationssystem der ARGE-ELBE. Abschlussbericht

    Energy Technology Data Exchange (ETDEWEB)

    Frank, B.; Prange, A.

    2001-07-01

    The geo-information system ELBIS was installed in mid-November 2000 to provide information on the web on Elbe water quality and other information. It presents the current results of the national Elbe monitoring programme of the ARGE Elbe working group, as well as the results of the 'Elbe 2000' programme of the nineties. [German] Das Geo-Informationssystem ELBIS informiert seit November 2000 im Internet ueber die Wasserqualitaet und verwandte Elbe-Themen. Mittels einer geografischen Oberflaeche stellt es die fortlaufenden, aktuellen Ergebnisse der nationalen Elbe-Messprogramme der ARGE Elbe (Arbeitsgemeinschaft fuer die Reinhaltung der Elbe) dar sowie Forschungsdaten aus der 'Elbe 2000'-Schadstoffforschung der neunziger Jahre. Auf einfache und zeitgemaesse Weise macht ELBIS diese Ergebnisse der interessierten Oeffentlichkeit zugaenglich und bietet Fachleuten einen umfassenden Ueberblick ueber die Datenlage. (orig.)

  6. Synthesis and electrochemical properties of ferrocene-peptides Boc-HN-Fca-Gly-Gly-Tyr-Arg-OMe%二茂铁-肽Boc-HN-Fca-Gly-Gly-Tyr-Arg-OMe的合成和电化学性质

    Institute of Scientific and Technical Information of China (English)

    王芳斌; 谭月敏; 刘又年; 曾冬铭; 向娟

    2007-01-01

    以二茂铁二甲酸、甘氨酸、酪氨酸和精氨酸为原料,通过多步反应,合成新的二茂铁-肽衍生物Boc-HN-Fca-Gly-Gly-Tyr-Arg-OMe(7)(Fca代表二茂铁氨基酸)和Boc-HN-Fca-Gly-Gly-Tyr-Arg-OH(8),对寡聚肽的合成进行研究;对合成产物进行红外光谱和氢核磁共振谱表征,并运用循环伏安(CV)方法对产物的电化学性能进行研究.研究结果表明:产物(7)和(8)的产率分别为83.5%和80.2%;Boc-HN-Fca-Gly-Gly-Tyr-Arg-OMe的循环伏安扫描结果是氧化峰和还原峰电位分别为0.385 V和0.346 V,峰电位之差△Ep为41 mV,峰电流密度之比Jpa/Jpc为1.055;对于Boc-HN-Fca-Gly-Gly-Tyr-Arg-OH,其氧化峰和还原峰电位分别为0.532 V和0.453 V,峰电位之差△Ep为79 mV,峰电流密度之比Jpa/Jpc为0.928.

  7. FGFR4 Gly388Arg polymorphism contributes to prostate cancer development and progression: A meta-analysis of 2618 cases and 2305 controls

    Directory of Open Access Journals (Sweden)

    Zhang Zheng D

    2011-02-01

    Full Text Available Abstract Background Fibroblast growth factor receptor 4 (FGFR4 displays multiple biological activities, including mitogenic and angiogenic activity, and plays important roles in the etiology and progression of prostate cancer. Gly388Arg polymorphism in FGFR4 gene has been reported to be involved in prostate cancer incidence and aggressiveness in several studies. To derive a more precise estimation of the relationship, a meta-analysis was performed. Methods Odds ratios (ORs with 95% confidence intervals (CIs were estimated to assess the association. Results The Arg388 allele increased prostate cancer risk compared with Gly388 allele (OR = 1.17, 95% CI = 1.07-1.29. When stratified by race, there was a significantly increased prostate cancer risk in Asian and Caucasian populations. Moreover, prostate cancer patients with Arg/Arg genotype had a 1.34-fold increased risk of advanced prostate cancer (95% CI: 1.03-1.74 compared with those with Gly/Gly+Gly/Arg genotype. Conclusion This meta-analysis showed the evidence that FGFR4 Gly388Arg polymorphism was associated with an increased risk of prostate cancer development and progression, suggesting that FGFR4 Gly388Arg polymorphism could be a marker for prostate cancer development and progression.

  8. KIF6 719Arg Carrier Status Association with Homocysteine and C-Reactive Protein in Amnestic Mild Cognitive Impairment and Alzheimer’s Disease Patients

    Directory of Open Access Journals (Sweden)

    Michael Malek-Ahmadi

    2013-01-01

    Full Text Available Recent research has demonstrated associations between statin use, KIF6 719Arg carrier status, and cholesterol levels and amnestic mild cognitive impairment (aMCI and Alzheimer’s disease (AD patients. The association between 719Arg carrier status with homocysteine (tHcy and c-reactive protein (CRP levels in aMCI and AD has not been previously investigated. Data from 175 aMCI and AD patients were used for the analysis. 719Arg carriers had significantly lower levels of tHcy than noncarriers (P=0.02. No significant difference in CRP levels between 719Arg carriers and noncarriers was present (P=0.37. Logistic regression yielded no significant effect for 719Arg status on CRP [OR = 1.79 (0.85, 3.83, P=0.13] but did demonstrate a significant effect for tHcy [OR = 0.44 (0.23, 0.83, P=0.01] after adjusting for ApoE ε4 carrier status, age, gender, and statin use. This study is the first to explore the relationship between KIF6 719Arg carrier status with tHcy and CRP levels. 719Arg carriers were more likely to have normal tHcy levels after adjusting for ApoE ε4 status, age, gender, and statin use. These results suggest that the KIF6 gene might influence cardiovascular pathways associated with AD.

  9. Effect of the Arg389Gly β₁-adrenoceptor polymorphism on plasma renin activity and heart rate, and the genotype-dependent response to metoprolol treatment.

    Science.gov (United States)

    Petersen, Morten; Andersen, Jon T; Jimenez-Solem, Espen; Broedbaek, Kasper; Hjelvang, Brian R; Henriksen, Trine; Frandsen, Erik; Forman, Julie L; Torp-Pedersen, Christian; Køber, Lars; Poulsen, Henrik E

    2012-09-01

    1. A gene-drug interaction has been indicated between β₁-adrenoceptor-selective beta-blockers and the Arg389Gly polymorphism (rs1801253) in the adrenergic beta-1 receptor gene (ADRB1). In the present study, we investigated the effect of the ADRB1 Arg389Gly polymorphism on plasma renin activity (PRA) and heart rate (HR), as well as genotype-dependent responses to metoprolol and exercise. 2. Twenty-nine healthy male subjects participated in two treatment periods (placebo and 200 mg/day metoprolol). A 15 min submaximal exercise test was performed after each treatment period and PRA and HR were measured before and after exercise. 3. Before exercise, median PRA was lower in Gly/Gly subjects than in Arg/Arg subjects after both placebo (P = 0.030) and metoprolol (P = 0.020) treatment. After placebo, the exercise-induced increase in PRA was greater in Gly/Gly than Arg/Gly and Arg/Arg subjects (P = 0.033). The linear association between log(PRA) and log(metoprolol concentration) varied significantly between genotypes (P = 0.024). In Gly/Gly subjects, PRA decreased significantly with metoprolol concentration before (P = 0.025) and after exercise (P concentration had no effect on PRA. The effect of metoprolol concentration on PRA in Gly/Gly subjects was enhanced by exercise (P = 0.044). No significant differences in HR were seen between genotype groups. 4. Resting PRA was lower in Gly/Gly than Arg/Arg subjects and the effect of exercise and metoprolol concentration on PRA was stronger in Gly/Gly subjects than with the other two genotypes. Thus, Gly/Gly heart failure patients may require lower doses of metoprolol than other patients to block neurohumoral hyperactivity.

  10. The role of Arg(78) in the metabotropic glutamate receptor mGlu(1) for agonist binding and selectivity

    DEFF Research Database (Denmark)

    Jensen, Anders A.; Sheppard, P O; O'Hara, P J

    2000-01-01

    likely, through the formation of an ionic bond between its positively charged side chain and the distal acid group of the agonists. Furthermore, the different impact of the two mutations on (S)-glutamic acid and (S)-quisqualic acid potencies strongly indicates that while Arg(78) appears to be a common......The metabotropic glutamate receptors belong to family C of the G-protein coupled receptor superfamily. These receptors all possess large extracellular amino terminal domains, where agonist binding takes place. We have previously constructed a molecular model of the amino terminal domain of the m......Glu(1) receptor based on a weak amino acid sequence similarity with a family of bacterial periplasmic binding proteins (PBPs). The residues Ser(165) and Thr(188) were demonstrated to be involved in agonist binding to the receptor. Here, we report that mutation of Arg(78) in the mGlu(1b) receptor...

  11. Hepatic fibrinogen storage disease due to the fibrinogen γ375 Arg → Trp mutation "fibrinogen aguadilla" is present in Arabs

    Directory of Open Access Journals (Sweden)

    Abdulrahman Al-Hussaini

    2014-01-01

    Full Text Available The mutation γ375Arg → Trp (fibrinogen Aguadilla is one of four mutations (Brescia, Aguadilla, Angers, and AI duPont capable of causing hepatic storage of fibrinogen. It has been observed in four children from the Caribbean, Europe, and Japan, suffering from cryptogenic liver disease. We report the first case of hepatic fibrinogen storage disease in Arabs due to a mutation in the fibrinogen γ-chain gene in a 3-year-old Syrian girl presenting with elevated liver enzymes. The finding of an impressive accumulation of fibrinogen in liver cells raised the suspicion of endoplasmic reticulum storage disease. Sequencing of the fibrinogen genes revealed a γ375Arg → Trp mutation (fibrinogen Aguadilla in the child and in her father. In conclusion, when confronted with chronic hepatitis of unknown origin, one should check the plasma fibrinogen level and look carefully for the presence of hepatocellular intracytoplasmic globular inclusions to exclude hepatic fibrinogen storage disease.

  12. Ecología, etnobotánica y etnofarmacología del argán (Argania spinosa)

    OpenAIRE

    López Sáez, José Antonio; Alba Sánchez, Francisca

    2009-01-01

    Argania spinosa es una especie leñosa endémica del suroeste de Marruecos que ha jugado un papel fundamental, desde un punto de vista etnobotánico, en la economía local de los pueblos bereberes. El aceite de argán se extrae de manera artesanal de los frutos de esta planta siguiendo un modelo etnográfico de gran interés. En el presente trabajo se sintetizan, mediante una profunda revisión bibliográfica, los conocimientos referidos a la ecología, etnobotánica y etnofarmacología del argán y se de...

  13. Isolation of L-3-phenyllactyl-Leu-Arg-Asn-NH2 (Antho-RNamide), a sea anemone neuropeptide containing an unusual amino-terminal blocking group

    DEFF Research Database (Denmark)

    Grimmelikhuijzen, C J; Rinehart, K L; Jacob, E

    1990-01-01

    -phenyllactyl-Leu-Arg-Asn-NH2. By using reversed-phase HPLC and a chiral mobile phase, it was shown that the 3-phenyllactyl group had the L configuration. Immunocytochemical staining with antiserum against Arg-Asn-NH2 showed that L-3-phenyllactyl-Leu-Arg-Asn-NH2 (Antho-RNamide) was localized in neurons of sea...... anemones. The L-3-phenyllactyl group has not been found earlier in neuropeptides of vertebrates or higher invertebrates. We propose that this residue renders Antho-RNamide resistant to nonspecific aminopeptidases, thereby increasing the stability of the peptide after neuronal release....

  14. The control in cis of the position and the amount of the ARG4 meiotic double-strand break of Saccharomyces cerevisiae.

    OpenAIRE

    de Massy, B.; Nicolas, A.

    1993-01-01

    During meiosis, a transient DNA double-strand break (DSB) occurs in the promoter region (positions -200/-185) of the Saccharomyces cerevisiae ARG4 gene and is a likely intermediate in the initiation of meiotic gene conversion events in this region. We report here a functional analysis of the ARG4 DSB based on the study of various deletions in this chromosomal region. We have identified several cis-acting elements located within the -465/+3 region of the ARG4 promoter that control the formatio...

  15. Differential coupling of Arg- and Gly389 polymorphic forms of the β1-adrenergic receptor leads to pathogenic cardiac gene regulatory programs

    OpenAIRE

    2008-01-01

    The β1-adrenergic receptor (β1AR; ADRB1) polymorphism Arg389Gly is located in an intracellular loop and is associated with distinct human and mouse cardiovascular phenotypes. To test the hypothesis that β1-Arg389 and β1-Gly389 alleles could differentially couple to pathways beyond that of classic Gs-adenylyl cyclase (AC)/cAMP signaling, we performed comparative gene expression profile analyses on hearts from wild-type and transgenic mice that expressed either human β1-Arg389 or β1-Gly389 rece...

  16. Leptin receptor Gln223Arg polymorphism and breast cancer risk in Nigerian women: A case control study

    Directory of Open Access Journals (Sweden)

    Anyanwu Stanley N

    2008-11-01

    Full Text Available Abstract Background Leptin, a 16 kDa polypeptide hormone, implicated in various physiological processes, exerts its action through the leptin receptor, a member of the class I cytokine receptor family. Both leptin and leptin receptor have recently been implicated in processes leading to breast cancer initiation and progression in animal models and humans. An A to G transition mutation in codon 223 in exon 6 of the leptin receptor gene, resulting in glutamine to arginine substitution (Gln223Arg, lies within the first of two putative leptin-binding regions and may be associated with impaired signaling capacity of the leptin receptor. This study was designed to assess the role of this polymorphism in breast cancer susceptibility in Nigerian women. Methods We utilized a polymerase chain reaction (PCR-based restriction fragment length polymorphism (RFLP assay to evaluate the association between the Gln223Arg polymorphism of the leptin receptor gene and breast risk in Nigeria in a case control study involving 209 women with breast cancer and 209 controls without the disease. Study participants were recruited from surgical outpatient clinics and surgical wards of four University Teaching Hospitals located in Midwestern and southeastern Nigeria between September 2002 and April 2004. Results Premenopausal women carrying at least one LEPR 223Arg allele were at a modestly increased risk of breast cancer after adjusting for confounders (OR = 1.8, 95% confidence interval [CI] 1.0–3.2, p = 0.07. There was no association with postmenopausal breast cancer risk (OR = 0.9, 95% CI 0.4–1.8, p = 0.68. Conclusion Our results suggest that the LEPR Gln223Arg polymorphism in the extracellular domain of the LEPR receptor gene is associated with a modestly increased risk of premenopausal breast cancer in Nigerian women.

  17. Association between TP53 gene Arg72Pro polymorphism and Wilms' tumor risk in a Chinese population.

    Science.gov (United States)

    Fu, Wen; Zhuo, Zhen-Jian; Jia, Wei; Zhu, Jinhong; Zhu, Shi-Bo; Lin, Ze-Feng; Wang, Feng-Hua; Xia, Huimin; He, Jing; Liu, Guo-Chang

    2017-01-01

    Wilms' tumor is one of the most prevalent pediatric malignancies, ranking fourth in childhood cancer worldwide. TP53 is a critical tumor suppressor gene, which encodes a 53 kDa protein, p53. The p53 functions to protect against cancer by regulating cell cycle and apoptosis and maintaining DNA integrity. TP53 gene is highly polymorphic. Several TP53 gene polymorphisms have been considered to be associated with cancer risk. Of them, a nonsynonymous polymorphism, Arg72Pro (rs1042522 C>G), has been most extensively studied for the association with cancer risk; however, few studies have investigated its effect on Wilms' tumor. Because of the central role of p53 in cell cycle control, the TP53 gene Arg72Pro polymorphism is also a good potential candidate predisposition locus for this pediatric cancer. We genotyped this polymorphism in 145 patients and 531 cancer-free controls recruited from Chinese children by Taqman methodology. Overall, our result suggested a lack of association between the TP53 gene Arg72Pro polymorphism and Wilms' tumor. In the stratified analysis, we found that carriers of CG/GG genotypes had a significantly increased Wilms' tumor risk in children not older than 18 months (adjusted odds ratio =2.04, 95% confidence interval =1.003-4.13, P=0.049) compared with CC genotype carriers. Our study indicated that the TP53 gene Arg72Pro polymorphism may have a weak, age-related effect on Wilms' tumor risk in Chinese children. These findings need further validations in other populations with larger sample size.

  18. Association between TP53 gene Arg72Pro polymorphism and Wilms’ tumor risk in a Chinese population

    Science.gov (United States)

    Fu, Wen; Zhuo, Zhen-Jian; Jia, Wei; Zhu, Jinhong; Zhu, Shi-Bo; Lin, Ze-Feng; Wang, Feng-Hua; Xia, Huimin; He, Jing; Liu, Guo-Chang

    2017-01-01

    Wilms’ tumor is one of the most prevalent pediatric malignancies, ranking fourth in childhood cancer worldwide. TP53 is a critical tumor suppressor gene, which encodes a 53 kDa protein, p53. The p53 functions to protect against cancer by regulating cell cycle and apoptosis and maintaining DNA integrity. TP53 gene is highly polymorphic. Several TP53 gene polymorphisms have been considered to be associated with cancer risk. Of them, a nonsynonymous polymorphism, Arg72Pro (rs1042522 C>G), has been most extensively studied for the association with cancer risk; however, few studies have investigated its effect on Wilms’ tumor. Because of the central role of p53 in cell cycle control, the TP53 gene Arg72Pro polymorphism is also a good potential candidate predisposition locus for this pediatric cancer. We genotyped this polymorphism in 145 patients and 531 cancer-free controls recruited from Chinese children by Taqman methodology. Overall, our result suggested a lack of association between the TP53 gene Arg72Pro polymorphism and Wilms’ tumor. In the stratified analysis, we found that carriers of CG/GG genotypes had a significantly increased Wilms’ tumor risk in children not older than 18 months (adjusted odds ratio =2.04, 95% confidence interval =1.003–4.13, P=0.049) compared with CC genotype carriers. Our study indicated that the TP53 gene Arg72Pro polymorphism may have a weak, age-related effect on Wilms’ tumor risk in Chinese children. These findings need further validations in other populations with larger sample size. PMID:28260929

  19. β1-adrenergic receptor(Arg389Gly) polymorphism and response to bisoprolol in patients with chronic heart failure

    Institute of Scientific and Technical Information of China (English)

    俞文萍

    2006-01-01

    Objective The purpose of this study was to investigate the relation between the Arg389Gly polymorphism of theβ1-AR gene and chronic heart failure (CHF) and to evaluate the effect of this polymorphism on clinical response toβ-adrenoceptor blockade (bisoprolol) in patients with CHF. Methods One hundred and ten patients with stable CHF receiving basic therapy for heart failure were included. Before initiation and 3 months af-

  20. Examination of soluble integrin resistant mutants of foot-and-mouth disease virus

    Directory of Open Access Journals (Sweden)

    Lawrence Paul

    2013-01-01

    Full Text Available Abstract Background Foot-and-mouth disease virus (FMDV initiates infection via recognition of one of at least four cell-surface integrin molecules αvβ1, αvβ3, αvβ6, or αvβ8 by a highly conserved Arg-Gly-Asp (RGD amino acid sequence motif located in the G-H loop of VP1. Within the animal host, the αvβ6 interaction is believed to be the most relevant. Sub-neutralizing levels of soluble secreted αvβ6 (ssαvβ6 was used as a selective pressure during passages in vitro to explore the plasticity of that interaction. Results Genetically stable soluble integrin resistant (SIR FMDV mutants derived from A24 Cruzeiro were selected after just 3 passages in cell culture in the presence of sub-neutralizing levels of ssαvβ6. SIR mutants were characterized by: replication on selective cell lines, plaque morphology, relative sensitivity to ssαvβ6 neutralization, relative ability to utilize αvβ6 for infection, as well as sequence and structural changes. All SIR mutants maintained an affinity for αvβ6. Some developed the ability to attach to cells expressing heparan sulfate (HS proteoglycan, while others appear to have developed affinity for a still unknown third receptor. Two classes of SIR mutants were selected that were highly or moderately resistant to neutralization by ssαvβ6. Highly resistant mutants displayed a G145D substitution (RGD to RDD, while moderately resistant viruses exhibited a L150P/R substitution at the conserved RGD + 4 position. VP1 G-H loop homology models for the A-type SIR mutants illustrated potential structural changes within the integrin-binding motif by these 2 groups of mutations. Treatment of O1 Campos with ssαvβ6 resulted in 3 SIR mutants with a positively charged VP3 mutation allowing for HS binding. Conclusions These findings illustrate how FMDV particles rapidly gain resistance to soluble receptor prophylactic measures in vitro. Two different serotypes developed distinct capsid mutations to circumvent the

  1. Cyclo(L-Pro-D-Arg): a new antibacterial and antitumour diketopiperazine from Bacillus cereus associated with a rhabditid entomopathogenic.

    Science.gov (United States)

    Kumar, S Nishanth; Mohandas, C; Nambisan, Bala; Sreerag, R S; Jayaprakas, C A

    2014-05-01

    In continuation of our search for new antimicrobial secondary metabolites from Bacillus cereus associated with rhabditid entomopathogenic nematode, a new microbial diketopiperazine, cyclo(L-Pro-D-Arg), was isolated from the ethyl acetate extract of fermented modified nutrient broth. The chemical structures of the isolated compounds were identified based on their 1D, 2D NMR and high-resolution electrospray ionisation-mass spectroscopy data. Antibacterial activity of the compound was determined by minimum inhibitory concentration and disc diffusion method against medically important bacteria, and the compound was recorded to have significant antibacterial activity against test bacteria. The highest activity was recorded against Klebsiella pneumoniae (1 μg/mL). Cyclo(L-Pro-D-Arg) was recorded to have significant antitumor activity against HeLa cells (IC50 value 50 μg/mL), and this compound was recorded to have no cytotoxicity against normal monkey kidney cells (VERO) up to 100 μg/mL). To the best of our knowledge, this is the first time that cyclo(L-Pro-D-Arg) has been isolated from a microbial natural source.

  2. Rapid Progression of Sporadic ALS in a Patient Carrying SOD1 p.Gly13Arg Mutation

    Science.gov (United States)

    Kim, Myung-Jin; Bae, Jae-Han; Kim, Jeong-Min; Kim, Hye Ryoun; Yoon, Byung-Nam; Sung, Jung-Joon

    2016-01-01

    Amyotrophic lateral sclerosis (ALS), the most common adult onset motor neuron disease, is pathologically characterized by progressive loss of the upper and lower motor neurons. Mutations in the Cu/Zn superoxide dismutase gene (SOD1) account for about 20% of familial ALS cases and a small percentage of sporadic ALS (SALS) cases, and have revealed a validated genotype-phenotype correlation. Herein, we report a p.Gly13Arg mutation in SOD1 exon 1 in a patient with SALS who presented with a rapidly progressive course, predominantly affecting the lower motor neurons. A 48-year-old man presented with progressive weakness and muscle atrophy of the left upper and lower limbs, followed by muscle fasciculation and cramping. The clinical features of the patient were clearly suggestive of ALS, and implied a sporadic form with rapid progression, predominantly affecting the lower motor neurons. Sequencing of the SOD1 gene by PCR revealed a missense mutation of G to C (c.37G>C) in exon 1, and amino acid substitution of glycine by arginine (p.Gly13Arg). This is the first case identifying the p.Gly13Arg mutation of SOD1 in the Korean population, and clinical assessments of this patient revealed a different phenotype compared with other cases. PMID:28035186

  3. Pharmacobotanic characterization of young stems and stem barks of Rauvolfia sellowii Müll. Arg., Apocynaceae

    Directory of Open Access Journals (Sweden)

    Leopoldo Clemente Baratto

    2010-09-01

    Full Text Available Rauvolfia sellowii Müll. Arg. (Apocynaceae, a Brazilian native tree rich in indole alkaloids, is known as "pau-pra-tudo" and popularly used as hypocholesterolemic, hypoglycemic and antihypertensive. The aim of this work was to study the anatomy of the young stems and stem barks of this medicinal plant, in order to contribute to the identification of the species as a drug. The plant material was fixed and prepared according to standard microtechniques. The young stems have remaining epidermis, but a suberified peridermis is evident. The phellogen is located in the cortical region, forming suber externally. Underneath the phellogen, lies the phelloderm and collenchymatic region. In the cortex, there are numerous laticifers and some fibers. There is an incomplete sclerenchymatic sheath, consisting of several groups of fibers and stone cells. The stem has internal phloem ordered as isolated groups side by side. Numerous laticifers, calcium oxalate crystals, idioblasts and amyloplasts are found in the cortex, phloem, xylem and pith. The stem bark has many layers of suber and cortical parenchyma, a sheath composed of fibers and stone cells totally lignified, and external phloem. These anatomical characteristic, taken together, can be used as quality control parameters for this species.Rauvolfia sellowii Müll. Arg. (Apocynaceae, uma árvore nativa brasileira rica em alcaloides indólicos, é conhecida como "pau-pra-tudo" e utilizada popularmente como hipocolesterolêmica, hipoglicêmica e anti-hipertensiva. O presente trabalho teve como objetivo caracterizar anatomicamente o caule jovem e a casca caulinar dessa planta medicinal, a fim de contribuir para a identificação e autenticidade da droga. O material vegetal foi fixado e submetido às microtécnicas usuais. O caule jovem possui epiderme remanescente, porém uma periderme suberificada é observada. O felogênio instala-se na região cortical, formando súber externamente. Subjacentes ao felog

  4. Laxative activities of Mareya micrantha (Benth. Müll. Arg. (Euphorbiaceae leaf aqueous extract in rats

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    Djaman Joseph A

    2010-02-01

    Full Text Available Abstract Background Mareya micrantha (Benth. Müll. Arg. (Euphorbiaceae is a shrub that is commonly used in Côte d'Ivoire (West Africa for the treatment of constipation and as an ocytocic drug. The present study was carried out to investigate the laxative activity of Mareya micrantha in albino's Wistar rats. Methods Rats were divided in 5 groups of 5 animals each, first group as control, second group served as standard (sodium picosulfate while group 3, 4 and 5 were treated with leaf aqueous extract of Mareya micrantha at doses of 100, 200 and 400 mg/kg body weight (b.w., per os respectively. The laxative activity was determined based on the weight of the faeces matter. The effects of the leaves aqueous extract of Mareya micrantha and castor oil were also evaluated on intestinal transit, intestinal fluid accumulation and ions secretion. Results Phytochemicals screening of the extract revealed the presence of flavonoids, alkaloids, tannins, polyphenols, sterols and polyterpenes. The aqueous extract of Mareya micrantha applied orally (100, 200 and 400 mg/kg; p.o., produced significant laxative activity and reduced loperamide induced constipation in dose dependant manner. The effect of the extract at 200 and 400 mg/kg (p.o. was similar to that of reference drug sodium picosulfate (5 mg/kg, p.o. The same doses of the extract (200 and 400 mg/kg, p.o. produced a significant increase (p -, Na+, K+ and Ca2+ in the intestinal fluid (p Conclusions The results showed that the aqueous extract of Mareya micrantha has a significant laxative activity and supports its traditional use in herbal medicine.

  5. In vivo andin vitro phytochemical and antibacterial efficacy ofBaliospermum montanum(Willd.)Muell.Arg

    Institute of Scientific and Technical Information of China (English)

    JohnsonM; WeselyEG; ZahirHussain MI; SelvanN

    2010-01-01

    Objective:To evaluate the phytochemical and anti-bacterial potential of mother plantsin vivo andin vitro derived callus ofBaliospermum montanum (B. montanum) (Willd.) Muell.-Arg. leaves and root.Methods:Thein vitro derived rootlets and leaves segments ofB. montanum were cut into0.5-0.7 cm in length and cultured on Murashige and Skoog solid medium supplemented with 3% sucrose, gelled with0.7% agar and different concentration of2, 4-Deither alone or in combinations. The preliminary phytochemical screening was performed by Harborne method. Antibacterial efficacy was performed by well diffusion method and incubated for24 h at37 ℃.Results:The highest percentage of callus formation (leaves segments86.9±0.56; root segments78.7±0.51) was obtained on Murashige and Skoog’s basal medium supplemented with 3% sucrose and2.0 mg/L of 2, 4-Dichlorophenoxy acetic acid. The phytochemical study revealed the high quantity presence of steroids, triterpenoids, glycosides, saponins, alkaloids, flavanoids, phenolic compounds, tannins, sugars etc of root and leaves derived calli. The ethanol extract of leaves segment derived calli ofB. montanum showed the maximum solubility and antimicrobial activity with the MIC ranged from 100to 200 μL.Conclusions: The preliminary phytochemical study confirmed that the calli mediated tissues showed the higher percentage of metabolite constituents and extraction value compared to thein vivo leaves and roots. The present study observation suggested that a possibility to establish high yielding genotypes byin vitro culture for production of medicinally important bioactive compounds.

  6. IL23R(Arg381Gln) functional polymorphism is associated with active pulmonary tuberculosis severity.

    Science.gov (United States)

    Ben-Selma, Walid; Boukadida, Jalel

    2012-08-01

    The purpose of our study was to investigate the association between a functional single nucleotide polymorphism (SNP) in the interleukin-23 receptor gene (IL23R; rs11209026, 1142 G(wild type) → A(reduced function), Arg381Gln) and disease severity outcome in pulmonary tuberculosis (TB) in the Tunisian population. SNP was investigated in a population of 168 patients with active pulmonary TB (cases were stratified into patients with minimal/moderate lung involvement, i.e., patients with minimal/moderate disease [Pmd], and patients with extensive lung involvement, i.e., patients with active disease [Pad]) and 150 healthy subjects. Genotype analyses were carried out using the PCR-restriction fragment length polymorphism method. We have found that the IL23R reduced-function allele 1142A and genotypes AA and AG were overrepresented, especially in the Pad subgroup compared with the control group (51% versus 18% [P = 10(-8)], 33% versus 5% [P = 10(-8)], and 36% versus 26% [P = 5 × 10(-3)], respectively). Additionally, comparison of the Pad and the Pmd groups showed that the A allele and AA genotype seemed to be associated with 2.79-fold (P = 4 × 10(-5)) and 7.74-fold (P = 10(-5)) increased risks of TB with minimal/moderate lung involvement, respectively. Our results demonstrate that the reduced-function polymorphism 1142G → A encoded by IL23R influences the outcome of disease severity of active pulmonary TB in Tunisian patients.

  7. The Storage Stability of Anthocyanins in Mao (Antidesma thwaitesianum Müll. Arg. Juice and Concentrate

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    Prommakool Arunya

    2016-01-01

    Full Text Available Mao or Makmao (Antidesma thwaitesianum Müll. Arg. is a wild plant found in the northeast of Thailand. Mao is one of fruits which are source of anthocyanins. Mao fruits are used for juice and concentrate which are consumed for healthy drinks. Determination of the kinetic parameters is essential to predict the quality changes and stability of anthocyanins in Mao juice and concentrate that occur during storage. The purpose of this research was to study the degradation of anthocyanins in Mao juice and concentrate during storage at 5, 30 and 37°C for 15 days. The storage stability of Mao anthocyanins was studied in 15°Brix juice and 45°Brix concentrate. The degradation kinetic (k, half-life (t1/2, activation energy (Ea and Q10 values for Mao anthocyanins degradation were determined. The results indicated that analysis of kinetic data for the degradation of anthocyanins followed a first-order reaction. An increase storage temperature from 5 to 30 and 37°C increased k value of anthocyanins in Mao juice and concentrate. Increasing storage temperature decreased t1/2 value of anthocyanins in both concentrations. At 5, 30 and 37°C, the t1/2 of anthocyanins decreased from 35 to 13 and 5 days for Mao juice and 32 to 25 and 21 days for Mao concentrate. The Ea value of the anthocyanins degradation in Mao juice and concentrate were 38.03 and 8.42 kJ/mol, respectively. Q10 values of both Mao juice and concentrate at 30-37°C were higher than those were storaged at 5-30°C. Thus higher stability of anthocyanins was achieved by using concentration and storage at lower temperature.

  8. Recent trends in phytochemistry, ethnobotany and pharmacological significance of Alchornea cordifolia (Schumach. & Thonn.) Muell. Arg.

    Science.gov (United States)

    Boniface, Pone Kamdem; Ferreira, Sabrina Baptista; Kaiser, Carlos Roland

    2016-09-15

    Alchornea cordifolia (Schumach. & Thonn.) Muell. Arg. (Euphorbiaceae) (A. cordifolia) is widely distributed throughout tropical Africa, where it is used extensively in traditional medicine. Conditions for which the plant has enjoyed wide use are: coughs, gonorrhoea, infertility, prostatitis, bacterial infections, diarrhoea, ulcers, pain, inflammation, fever and bronchial troubles. This review summarizes the achievements of the investigations in traditional uses, ethnobotany, phytochemistry, biological activities and toxicological profile of A. cordifolia; this review also describes the shortcomings of studies on this herbal drug and thus serves as the basis of further scientific research and development of this traditional herbal drug. A. cordifolia-related information was collected from various resources including published articles in peer-reviewed journals, unpublished materials, textbooks, government survey reports and scientific databases such as Scifinder®, Pubmed, Science Direct, Wiley, Springer, ACS, Scielo, Web of Science and other web search instruments (Google, Yahoo), published on the subject from 1950 to 2016. 'The Plant List' (www.theplantlist.org) and 'Kew Royal Botanic Gardens' (mpns.kew.org) were used to validate the scientific name of the plant. The literature revealed several reports on traditional uses, biological activities, chemical constituents and toxicological evaluation of A. cordifolia. The phytochemical information indicates identification of 95 compounds including fatty acids, terpenoids, flavonoids, phenolic acids, alkaloids, which exhibited various pharmacological activities such as wound healing, anti-inflammation, anticancer, antioxidant, immunomodulation, antidiarrhoeal, antimicrobial, antidepressant, hepatoprotective, antiplasmodial and anxiolytic. However, there are still significant gaps in the completeness of our understanding of A. cordifolia bioactivity, therapeutic value, and roles played by each of the numerous

  9. N-Protonated Isomers and Coulombic Barriers to Dissociation of Doubly Protonated Ala8Arg

    Science.gov (United States)

    Haeffner, Fredrik; Irikura, Karl K.

    2017-10-01

    Collision-induced dissociation (or tandem mass spectrometry, MS/MS) of a protonated peptide results in a spectrum of fragment ions that is useful for inferring amino acid sequence. This is now commonplace and a foundation of proteomics. The underlying chemical and physical processes are believed to be those familiar from physical organic chemistry and chemical kinetics. However, first-principles predictions remain intractable because of the conflicting necessities for high accuracy (to achieve qualitatively correct kinetics) and computational speed (to compensate for the high cost of reliable calculations on such large molecules). To make progress, shortcuts are needed. Inspired by the popular mobile proton model, we have previously proposed a simplified theoretical model in which the gas-phase fragmentation pattern of protonated peptides reflects the relative stabilities of N-protonated isomers, thus avoiding the need for transition-state information. For singly protonated Ala n ( n = 3-11), the resulting predictions were in qualitative agreement with the results from low-energy MS/MS experiments. Here, the comparison is extended to a model tryptic peptide, doubly protonated Ala8Arg. This is of interest because doubly protonated tryptic peptides are the most important in proteomics. In comparison with experimental results, our model seriously overpredicts the degree of backbone fragmentation at N9. We offer an improved model that corrects this deficiency. The principal change is to include Coulombic barriers, which hinder the separation of the product cations from each other. Coulombic barriers may be equally important in MS/MS of all multiply charged peptide ions. [Figure not available: see fulltext.

  10. Biochemical and histological characterization of tomato mutants

    Directory of Open Access Journals (Sweden)

    Carolina C. Monteiro

    2012-06-01

    Full Text Available Biochemical responses inherent to antioxidant systems as well morphological and anatomical properties of photomorphogenic, hormonal and developmental tomato mutants were investigated. Compared to the non-mutant Micro-Tom (MT, we observed that the malondialdehyde (MDA content was enhanced in the diageotropica (dgt and lutescent (l mutants, whilst the highest levels of hydrogen peroxide (H2O2 were observed in high pigment 1 (hp1 and aurea (au mutants. The analyses of antioxidant enzymes revealed that all mutants exhibited reduced catalase (CAT activity when compared to MT. Guaiacol peroxidase (GPOX was enhanced in both sitiens (sit and notabilis (not mutants, whereas in not mutant there was an increase in ascorbate peroxidase (APX. Based on PAGE analysis, the activities of glutathione reductase (GR isoforms III, IV, V and VI were increased in l leaves, while the activity of superoxide dismutase (SOD isoform III was reduced in leaves of sit, epi, Never ripe (Nr and green flesh (gf mutants. Microscopic analyses revealed that hp1 and au showed an increase in leaf intercellular spaces, whereas sit exhibited a decrease. The au and hp1 mutants also exhibited a decreased in the number of leaf trichomes. The characterization of these mutants is essential for their future use in plant development and ecophysiology studies, such as abiotic and biotic stresses on the oxidative metabolism.Neste trabalho, analisamos as respostas bioquímicas inerentes ao sistema antioxidante, assim como propriedades morfológicas e anatômicas de mutantes fotomorfogenéticos e hormonais de tomateiro. Comparados ao não mutante Micro-Tom (MT, observamos que o conteúdo de malondialdeído (MDA aumentou nos mutantes diageotropica (dgt e lutescent (l, enquanto os maiores níveis de H2O2 foram encontrados nos mutantes high pigment 1 (hp1 e aurea (au. Análises de enzimas antioxidantes mostraram que todos os mutantes reduziram a atividade de catalase (CAT quando comparado a MT. A

  11. 含有困难氨基酸序列Arg-Arg的醋酸阿基瑞林的固相合成及其动力学研究%Solid Phase Synthesis of Argireline Acetate with Difficult Amino Acid Series Arg-Arg and its Reaction Kinetics

    Institute of Scientific and Technical Information of China (English)

    朱亮亮; 绳则翠; 陈英文; 沈树宝

    2013-01-01

    着重探讨大位阻氨基酸Fmoc-Arg(Pbf)-OH与NH2-Arg(Pbf)-Rink Amide-AM之间的缩合反应,同时对缩合反应进行动力学实验,以期能够获得一条最佳的醋酸阿基瑞林合成路线并对工业化生产进行指导。采用自制新型多肽固相合成反应器,集成过滤、搅拌、控温、鼓泡四重功能。通过对缩合过程中各反应条件的优化,确立了一条最佳的醋酸阿基瑞林合成路线:NMP/DCM(V:V=2:1)为反应溶剂,HBTU/HOBt/DIEA为缩合试剂,氨基酸与树脂摩尔比3:1,40℃下反应2.5 h,粗肽产率高达90.2%。经RP-HPLC分析,醋酸阿基瑞林粗品纯度为83.5%。同时采用微分法对大位阻氨基酸Fmoc-Arg(Pbf)-OH 与NH2-Arg(Pbf)-Rink Amide-AM之间的缩合反应进行动力学研究,测定了温度变化对反应速率的影响,并获得了活化酯方法下缩合反应的宏观动力学参数:反应级数为1.67,表观活化能为37.4 kJ×mol-1。%In order to obtain a best synthetic route of Argireline Acetate and the guidance of its industrial production, this paper focuses on the condensation reaction between sterically hindered amino acid Fmoc-Arg (Pbf)-OH and NH2-Arg(Pbf)-Rink Amide-AM with self-designed reactor, and its kinetic experiments were carried out at the same time. In self-designed reactor, the four functions of filtration, stir, temperature control and bubbling were combined. By optimizing the reaction conditions in the condensation process, a best synthetic route of Argireline Acetate was established, and its optimum reaction conditions were found as follows:the molar ratio of amino acid to resin 3:1, reaction time 2.5 h, temperature 40℃, NMP/DCM (2:1, V/V) as the solvent and HBTU/HOBt/DIEA as coupling reagents. Under above conditions, the highest yield of Argireline Acetate is 90.2%, and its purity is 83.5% by RP-HPLC analysis. The macroscopic kinetics of condensation reaction of Fmoc-Arg-(Pbf)-OH and NH2-Arg(Pbf)-Rink Amide-AM resin

  12. Normal CBF values by the ARG method using IMP SPECT. Comparison with a conventional microsphere model method

    Energy Technology Data Exchange (ETDEWEB)

    Ito, Hiroshi; Koyama, Masamichi; Kawashima, Ryuta; Ono, Shuichi; Fukuda, Hiroshi [Tohoku Univ., Sendai (Japan). Inst. of Development, Aging and Cancer; Ishii, Kiyoshi; Kinoshita, Toshifumi

    1996-02-01

    N-isopropyl-p[{sup 123}I]iodoamphetamine (IMP) has been used as a flow tracer for SPECT, and measurement of cerebral blood flow (CBF) using IMP has been performed by conventional microsphere model method (MS method). Recently, the ARG method for measuring CBF by using IMP with one SPECT scan and one point blood sampling has been developed. This method was based on two-compartment model. In the present study, normal CBF values were measured in 10 healthy male subjects (mean age{+-}S.D.: 29.8{+-}6.01, age range: 23-41) by ARG and MS methods. The mean CBF values ({+-}S.D.) for ARG method in which V{sub d} value was assumed to be 50 ml/ml were 41.7{+-}9.4, 31.1{+-}5.0, 40.7{+-}9.7, 41.5{+-}10.0, 38.2{+-}9.2, 39.0{+-}9.4, 41.9{+-}10.6, 38.7{+-}8.0 and 30.0{+-}7.7 ml/100 ml/min in the cerebellum, pons, thalamus, basal ganglia, frontal, temporal, parietal, occipital lobe cortex and centrum semiovale, respectively. The mean CBF values for the MS method were 46.8{+-}8.4, 37.5{+-}5.6, 45.8{+-}8.6, 46.5{+-}8.9, 43.7{+-}8.3, 44.4{+-}8.7, 46.8{+-}9.3, 44.3{+-}7.3 and 36.3{+-}8.1 ml/100 ml/min, respectively. The mean CBF values in the cerebral cortex region for ARG method were lower than those previously reported by PET. This would be caused by low first-pass extraction fraction of IMP compared with oxygen-15 labeled water. The mean CBF values for the MS method were higher than those for ARG method against previous studies. As reasons for this, errors in estimation of SPECT brain counts at 8 min in the MS method were considered. (author).

  13. The Arg233Lys AQP0 mutation disturbs aquaporin0-calmodulin interaction causing polymorphic congenital cataract.

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    Shanshan Hu

    Full Text Available Calmodulin (CaM directly interacts with the aquaporin 0 (AQP0 C-terminus in a calcium dependent manner to regulate the water permeability of AQP0. We previously identified a missense mutation (p.R233K in the putative CaM binding domain of AQP0 C-terminus in a congenital cataract family. This study was aimed at exploring the potential pathogenesis of this mutation causative of cataract and mainly identifying how it influenced the binding of AQP0 to CaM. Wild type and R233K mutant AQP0 with EGFP-tag were transfected separately into Hela cells to determine the expression and subcellular localizations. The co-immunoprecipitation (CoIP assay was used to detect the interaction between AQP0 and CaM. AQP0 C-terminus peptides were synthesized with and without R233K, and the binding abilities of these peptides to CaM were assessed using a fluorescence binding assay. Localizations of wild type and R233K mutant AQP0 were determined from EGFP fluorescence, and the chimeric proteins were both localized abundantly in the plasma membrane. Protein expression levels of the culture cells showed no significant difference between them. The results from CoIP assay implied that R233K mutant presented more weakly in association with CaM than wild type AQP0. The AQP0 C-terminal mutant peptide was found to have 2.5-fold lower binding affinity to CaM than wild type peptide. These results suggested that R233K mutation did not affect the expression, location and trafficking of the protein but did influence the interaction between AQP0 and CaM. The binding affinity of AQP0 C-terminus to CaM was significantly reduced. Due to lack of the modulation of the Ca2+-calmodulin complex, the water permeability of AQP0 was subsequently augmented, which might lead to the development of this cataract.

  14. Energy expenditure, body composition and insulin response to glucose in male twins discordant for the Trp64Arg polymorphism of the beta3-adrenergic receptor gene

    DEFF Research Database (Denmark)

    Højlund, K; Christiansen, C; Bjørnsbo, K S;

    2006-01-01

    and environmental background, the Trp64Arg polymorphism of the beta3AR gene is associated with lower fat mass, fasting insulin levels and an appropriate insulin response to glucose. Thus, heterozygosity for the Trp64Arg variant is unlikely to increase the risk of obesity, insulin resistance or type 2 diabetes.......AIM: The tryptophan to arginine change in position 64 (Trp64Arg) polymorphism of the beta3-adrenergic receptor (beta3AR) gene has been associated with an increased prevalence of obesity, insulin resistance and type 2 diabetes. In this, decreased rates of energy expenditure and impaired insulin......-ray absorptiometry scanning and energy expenditure by indirect and direct calorimetry. RESULTS: Twins heterozygous for the Trp64Arg polymorphism showed significantly lower fat mass independent of the method used, and significantly lower fasting insulin and glucose concentrations compared with their homozygous wild...

  15. The Effect of the Arg389Gly Beta-1 Adrenoceptor Polymorphism on Plasma Renin Activity and Heart Rate and the Genotype-Dependent Response to Metoprolol Treatment

    DEFF Research Database (Denmark)

    Petersen, Morten; Andersen, Jon T; Jimenez-Solem, Espen

    2012-01-01

    A gene-drug interaction has been indicated between beta-1 selective beta-blockers and the Arg389Gly polymorphism (rs1801253) in the adrenergic beta-1 receptor gene (ADRB1). We studied the effect of the ADRB1 Arg389Gly polymorphism on plasma renin activity (PRA) and heart rate (HR) and the genotype......(metoprolol concentration) varied significantly between genotypes (P = 0.024). In Gly/Gly subjects, PRA decreased significantly with metoprolol concentration before (P = 0.025) and after exercise (P effect on PRA. The effect of metoprolol concentration...... on PRA in Gly/Gly subjects was enhanced by exercise (P = 0.044). No significant differences in HR were seen between genotype groups. Resting PRA was lower in Gly/Gly than in Arg/Arg subjects, and the effect of exercise and metoprolol concentration on PRA was stronger in Gly/Gly subjects than...

  16. ADH1B Arg47His polymorphism is associated with esophageal cancer risk in high-incidence Asian population: evidence from a meta-analysis.

    Science.gov (United States)

    Zhang, Guohong; Mai, Ruiqin; Huang, Bo

    2010-10-27

    Incidence of Esophageal squamous cell carcinoma (ESCC) is prevalent in Asian populations, especially in the ones from the "Asian esophageal cancer belt" along the Silk Road and the ones from East Asia (including Japan). Silk Road and Eastern Asia population genetics are relevant to the ancient population migration from central China. The Arg47His (rs1229984) polymorphism of ADH1B is the highest in East Asians, and ancient migrations along the Silk Road were thought to be contributive to a frequent ADH1B*47His allele in Central Asians. This polymorphism was identified as responsible for susceptibility in the first large-scale genome-wide association study of ESCC and that's explained by its modulation of alcohol oxidization capability. To investigate the association of ADH1B Arg47His with ESCC in Asian populations under a common ancestry scenario of the susceptibility loci, we combined all available studies into a meta-analysis. A dataset composed of 4,220 cases and 8,946 controls from twelve studies of Asian populations was analyzed for ADH1B Arg47His association with ESCC and its interactions with alcohol drinking and ALDH2 Glu504Lys. Heterogeneity among studies and their publication bias were also tested. The ADH1B*47Arg allele was found to be associated to increased risk of ESCC, with the odds ratios (OR) being 1.62 (95% CI: 1.49-1.76) and 3.86 (2.96-5.03) for the His/Arg and the Arg/Arg genotypes, respectively. When compared with the His/His genotype of non-drinkers, the Arg/Arg genotype can interact with alcohol drinking and greatly increase the risk of ESCC (OR = 20.69, 95%CI: 5.09-84.13). Statistical tests also showed gene-gene interaction of ADH1B Arg+ with ALDH2 Lys+ can bring more risk to ESCC (OR  = 13.46, 95% CI: 2.32-78.07). Revealed by this meta-analysis, ADH1B*47Arg as a common ancestral allele can significantly increase the risk of ESCC in Asians, especially when coupled with alcohol drinking or the ALDH2*504Lys allele.

  17. ADH1B Arg47His polymorphism is associated with esophageal cancer risk in high-incidence Asian population: evidence from a meta-analysis.

    Directory of Open Access Journals (Sweden)

    Guohong Zhang

    Full Text Available BACKGROUND AND OBJECTIVES: Incidence of Esophageal squamous cell carcinoma (ESCC is prevalent in Asian populations, especially in the ones from the "Asian esophageal cancer belt" along the Silk Road and the ones from East Asia (including Japan. Silk Road and Eastern Asia population genetics are relevant to the ancient population migration from central China. The Arg47His (rs1229984 polymorphism of ADH1B is the highest in East Asians, and ancient migrations along the Silk Road were thought to be contributive to a frequent ADH1B*47His allele in Central Asians. This polymorphism was identified as responsible for susceptibility in the first large-scale genome-wide association study of ESCC and that's explained by its modulation of alcohol oxidization capability. To investigate the association of ADH1B Arg47His with ESCC in Asian populations under a common ancestry scenario of the susceptibility loci, we combined all available studies into a meta-analysis. METHODS: A dataset composed of 4,220 cases and 8,946 controls from twelve studies of Asian populations was analyzed for ADH1B Arg47His association with ESCC and its interactions with alcohol drinking and ALDH2 Glu504Lys. Heterogeneity among studies and their publication bias were also tested. RESULTS: The ADH1B*47Arg allele was found to be associated to increased risk of ESCC, with the odds ratios (OR being 1.62 (95% CI: 1.49-1.76 and 3.86 (2.96-5.03 for the His/Arg and the Arg/Arg genotypes, respectively. When compared with the His/His genotype of non-drinkers, the Arg/Arg genotype can interact with alcohol drinking and greatly increase the risk of ESCC (OR = 20.69, 95%CI: 5.09-84.13. Statistical tests also showed gene-gene interaction of ADH1B Arg+ with ALDH2 Lys+ can bring more risk to ESCC (OR  = 13.46, 95% CI: 2.32-78.07. CONCLUSION: Revealed by this meta-analysis, ADH1B*47Arg as a common ancestral allele can significantly increase the risk of ESCC in Asians, especially when coupled

  18. Study on culturing Trichodema mutants

    Institute of Scientific and Technical Information of China (English)

    CHEN Jian-ai; WANG Wei-ming

    2004-01-01

    @@ Trichodema mutants strains T5, T0803, T1010, T1003were cultured in different conditions and media, also in the presence of fungicides at 40 mg/kg (CK or procymidone + chlorothalonil, or maneb or phosethyl-Al) . The pH values of media were 5, 6, 7 and 8 and hyphae were grown at temperatures of 15, 20, 25 and 30 ℃. After being cultured for 3, 4, 5, or 6 days, the strains were transferred at a lower temperature to sporulate (20℃) Obtained data were analyzed statistically, with the orthogonal array and ranges (R) differing dependes on the treatments (R = 40.0,42.4, 48.0, 62.8,107.0). The results indicated that the most important factor was the nature of the strain (R =107.0), while the change in temperature and time of cultivation produced the lowest effect (R =40.0). Each factor variance was significant and A3B4C2D1E3 was the optimum combined condition, in which strain T1010 grew more quickly and sporulated most.

  19. CMPD: cancer mutant proteome database.

    Science.gov (United States)

    Huang, Po-Jung; Lee, Chi-Ching; Tan, Bertrand Chin-Ming; Yeh, Yuan-Ming; Julie Chu, Lichieh; Chen, Ting-Wen; Chang, Kai-Ping; Lee, Cheng-Yang; Gan, Ruei-Chi; Liu, Hsuan; Tang, Petrus

    2015-01-01

    Whole-exome sequencing, which centres on the protein coding regions of disease/cancer associated genes, represents the most cost-effective method to-date for deciphering the association between genetic alterations and diseases. Large-scale whole exome/genome sequencing projects have been launched by various institutions, such as NCI, Broad Institute and TCGA, to provide a comprehensive catalogue of coding variants in diverse tissue samples and cell lines. Further functional and clinical interrogation of these sequence variations must rely on extensive cross-platforms integration of sequencing information and a proteome database that explicitly and comprehensively archives the corresponding mutated peptide sequences. While such data resource is a critical for the mass spectrometry-based proteomic analysis of exomic variants, no database is currently available for the collection of mutant protein sequences that correspond to recent large-scale genomic data. To address this issue and serve as bridge to integrate genomic and proteomics datasets, CMPD (http://cgbc.cgu.edu.tw/cmpd) collected over 2 millions genetic alterations, which not only facilitates the confirmation and examination of potential cancer biomarkers but also provides an invaluable resource for translational medicine research and opportunities to identify mutated proteins encoded by mutated genes.

  20. Proteomic analysis of the effects of aged garlic extract and its FruArg component on lipopolysaccharide-induced neuroinflammatory response in microglial cells.

    Directory of Open Access Journals (Sweden)

    Hui Zhou

    Full Text Available Aged garlic extract (AGE is widely used as a dietary supplement, and is claimed to promote human health through anti-oxidant/anti-inflammatory activities with hypolipidemic, antiplatelet and neuroprotective effects. Prior studies of AGE have mainly focused on its organosulfur compounds, with little attention paid to its carbohydrate derivatives, such as N-α-(1-deoxy-D-fructos-1-yl-L-arginine (FruArg. The goal of this study is to investigate actions of AGE and FruArg on antioxidative and neuroinflammatory responses in lipopolysaccharide (LPS-activated murine BV-2 microglial cells using a proteomic approach. Our data show that both AGE and FruArg can significantly inhibit LPS-induced nitric oxide (NO production in BV-2 cells. Quantitative proteomic analysis by combining two dimensional differential in-gel electrophoresis (2D-DIGE with mass spectrometry revealed that expressions of 26 proteins were significantly altered upon LPS exposure, while levels of 20 and 21 proteins exhibited significant changes in response to AGE and FruArg treatments, respectively, in LPS-stimulated BV-2 cells. Notably, approximate 78% of the proteins responding to AGE and FruArg treatments are in common, suggesting that FruArg is a major active component of AGE. MULTICOM-PDCN and Ingenuity Pathway Analyses indicate that the proteins differentially affected by treatment with AGE and FruArg are involved in inflammatory responses and the Nrf2-mediated oxidative stress response. Collectively, these results suggest that AGE and FruArg attenuate neuroinflammatory responses and promote resilience in LPS-activated BV-2 cells by suppressing NO production and by regulating expression of multiple protein targets associated with oxidative stress.

  1. Photoactivation of Mutant Isocitrate Dehydrogenase 2 Reveals Rapid Cancer-Associated Metabolic and Epigenetic Changes.

    Science.gov (United States)

    Walker, Olivia S; Elsässer, Simon J; Mahesh, Mohan; Bachman, Martin; Balasubramanian, Shankar; Chin, Jason W

    2016-01-27

    Isocitrate dehydrogenase is mutated at a key active site arginine residue (Arg172 in IDH2) in many cancers, leading to the synthesis of the oncometabolite (R)-2-hydroxyglutarate (2HG). To investigate the early events following acquisition of this mutation in mammalian cells we created a photoactivatable version of IDH2(R172K), in which K172 is replaced with a photocaged lysine (PCK), via genetic code expansion. Illumination of cells expressing this mutant protein led to a rapid increase in the levels of 2HG, with 2HG levels reaching those measured in patient tumor samples, within 8 h. 2HG accumulation is closely followed by a global decrease in 5-hydroxymethylcytosine (5-hmC) in DNA, demonstrating that perturbations in epigenetic DNA base modifications are an early consequence of mutant IDH2 in cells. Our results provide a paradigm for rapidly and synchronously uncloaking diverse oncogenic mutations in live cells to reveal the sequence of events through which they may ultimately cause transformation.

  2. Expression and purification of the Bordella Pertussis toxin S1 subunit mutant in Escherichia coli

    Institute of Scientific and Technical Information of China (English)

    Xiao L. Zhang; Quan M. Zou

    2007-01-01

    Bordella pertussis is the causative agent of whooping cough.Traditional vaccines against this disease are inherently reactogenic, thus research is currentlly focussed on the production of less reactive,acellular vaccines.Expression of candidate antigens for these vaccines in Escherichia coli would be preferable. Pertussis toxin S1 subunit plays a critical role in the bacterium-host interplay.The mutant(rS1) containing two key amino acids substitution(Arg9-Lys/Glu129-Gly)is nontoxin and immunogenic and while retaining the protective epitopes. In this study, the immunoprotective S1 fragment of pertussis toxin fusion was verified by restriction endonuclease analysis and Western immunoblotting. Escherichia coli carrying the recombinant plasmid(pQE-rS1)produced a 26 kDa protein that was recognized by antibodies specific to the S1. Expressed rS1 in E. coli was purified from the inclusion bodies. The N-terminal 6 histidines could easily be captured by Ni-NTA affinity chromatography. Then, the rS1 of interest was purified to 92% homogeneity. Antisera generated against the purified S1 mutant protein recognized the native toxin indicating that some, if not all, of the native epitope were conserved. Thus, this vaccine preparation is potentially applicable for the production of novel vaccines against B. pertussis infection.

  3. Structural and population-based evaluations of TBC1D1 p.Arg125Trp.

    Directory of Open Access Journals (Sweden)

    Tom G Richardson

    Full Text Available Obesity is now a leading cause of preventable death in the industrialised world. Understanding its genetic influences can enhance insight into molecular pathogenesis and potential therapeutic targets. A non-synonymous polymorphism (rs35859249, p.Arg125Trp in the N-terminal TBC1D1 phosphotyrosine-binding (PTB domain has shown a replicated association with familial obesity in women. We investigated these findings in the Avon Longitudinal Study of Parents and Children (ALSPAC, a large European birth cohort of mothers and offspring, and by generating a predicted model of the structure of this domain. Structural prediction involved the use of three separate algorithms; Robetta, HHpred/MODELLER and I-TASSER. We used the transmission disequilibrium test (TDT to investigate familial association in the ALSPAC study cohort (N = 2,292 mother-offspring pairs. Linear regression models were used to examine the association of genotype with mean measurements of adiposity (Body Mass Index (BMI, waist circumference and Dual-energy X-ray absorptiometry (DXA assessed fat mass, and logistic regression was used to examine the association with odds of obesity. Modelling showed that the R125W mutation occurs in a location of the TBC1D1 PTB domain that is predicted to have a function in a putative protein:protein interaction. We did not detect an association between R125W and BMI (mean per allele difference 0.27 kg/m(2 (95% Confidence Interval: 0.00, 0.53 P = 0.05 or obesity (odds ratio 1.01 (95% Confidence Interval: 0.77, 1.31, P = 0.96 in offspring after adjusting for multiple comparisons. Furthermore, there was no evidence to suggest that there was familial association between R125W and obesity (χ(2 = 0.06, P = 0.80. Our analysis suggests that R125W in TBC1D1 plays a role in the binding of an effector protein, but we find no evidence that the R125W variant is related to mean BMI or odds of obesity in a general population sample.

  4. Reducing ppGpp level rescues an extreme growth defect caused by mutant EF-Tu.

    Science.gov (United States)

    Bergman, Jessica M; Hammarlöf, Disa L; Hughes, Diarmaid

    2014-01-01

    Transcription and translation of mRNA's are coordinated processes in bacteria. We have previously shown that a mutant form of EF-Tu (Gln125Arg) in Salmonella Typhimurium with a reduced affinity for aa-tRNA, causes ribosome pausing, resulting in an increased rate of RNase E-mediated mRNA cleavage, causing extremely slow growth, even on rich medium. The slow growth phenotype is reversed by mutations that reduce RNase E activity. Here we asked whether the slow growth phenotype could be reversed by overexpression of a wild-type gene. We identified spoT (encoding ppGpp synthetase/hydrolase) as a gene that partially reversed the slow growth rate when overexpressed. We found that the slow-growing mutant had an abnormally high basal level of ppGpp that was reduced when spoT was overexpressed. Inactivating relA (encoding the ribosome-associated ppGpp synthetase) also reduced ppGpp levels and significantly increased growth rate. Because RelA responds specifically to deacylated tRNA in the ribosomal A-site this suggested that the tuf mutant had an increased level of deacylated tRNA relative to the wild-type. To test this hypothesis we measured the relative acylation levels of 4 families of tRNAs and found that proline isoacceptors were acylated at a lower level in the mutant strain relative to the wild-type. In addition, the level of the proS tRNA synthetase mRNA was significantly lower in the mutant strain. We suggest that an increased level of deacylated tRNA in the mutant strain stimulates RelA-mediated ppGpp production, causing changes in transcription pattern that are inappropriate for rich media conditions, and contributing to slow growth rate. Reducing ppGpp levels, by altering the activity of either SpoT or RelA, removes one cause of the slow growth and reveals the interconnectedness of intracellular regulatory mechanisms.

  5. Reducing ppGpp level rescues an extreme growth defect caused by mutant EF-Tu.

    Directory of Open Access Journals (Sweden)

    Jessica M Bergman

    Full Text Available Transcription and translation of mRNA's are coordinated processes in bacteria. We have previously shown that a mutant form of EF-Tu (Gln125Arg in Salmonella Typhimurium with a reduced affinity for aa-tRNA, causes ribosome pausing, resulting in an increased rate of RNase E-mediated mRNA cleavage, causing extremely slow growth, even on rich medium. The slow growth phenotype is reversed by mutations that reduce RNase E activity. Here we asked whether the slow growth phenotype could be reversed by overexpression of a wild-type gene. We identified spoT (encoding ppGpp synthetase/hydrolase as a gene that partially reversed the slow growth rate when overexpressed. We found that the slow-growing mutant had an abnormally high basal level of ppGpp that was reduced when spoT was overexpressed. Inactivating relA (encoding the ribosome-associated ppGpp synthetase also reduced ppGpp levels and significantly increased growth rate. Because RelA responds specifically to deacylated tRNA in the ribosomal A-site this suggested that the tuf mutant had an increased level of deacylated tRNA relative to the wild-type. To test this hypothesis we measured the relative acylation levels of 4 families of tRNAs and found that proline isoacceptors were acylated at a lower level in the mutant strain relative to the wild-type. In addition, the level of the proS tRNA synthetase mRNA was significantly lower in the mutant strain. We suggest that an increased level of deacylated tRNA in the mutant strain stimulates RelA-mediated ppGpp production, causing changes in transcription pattern that are inappropriate for rich media conditions, and contributing to slow growth rate. Reducing ppGpp levels, by altering the activity of either SpoT or RelA, removes one cause of the slow growth and reveals the interconnectedness of intracellular regulatory mechanisms.

  6. The [Lys(-2)-Arg(-1)-des(17-21)]-endothelin-1 peptide retains the specific Arg(-1)-Asp8 salt bridge but reveals discrepancies between NMR data and molecular dynamics simulations.

    Science.gov (United States)

    Kaas, Quentin; Aumelas, André; Kubo, Shigeru; Chino, Naoyoshi; Kobayashi, Yuji; Chiche, Laurent

    2002-09-17

    The [des(17-21)]-endothelin-1 (CSH-ET) and [Lys(-)(2)-Arg(-)(1)-des(17-21)]-endothelin-1 (KR-CSH-ET) peptides, designed by removing the five-residue hydrophobic tail from the endothelin-1 (ET-1) and [Lys(-)(2)-Arg(-)(1)]-endothelin-1 (KR-ET-1) peptides, respectively, were synthesized. Previous studies on KR-ET-1 showed that, in contrast to ET-1, this engineered compound displays a pH-dependent conformational change related to the formation of a stabilizing salt bridge between the Arg(-)(1) and Asp(8) side chains. CD and NMR spectra indicate that CSH-ET and KR-CSH-ET display conformational behavior similar to those of ET-1 and KR-ET-1, respectively. The short salt bridge-stabilized KR-CSH-ET peptide therefore appears to be an attractive elementary scaffold for drug design. The solution structure of the salt-bridged form of KR-CSH-ET was determined by NMR at pH 4.5 and is very similar to the corresponding form of the parent KR-ET-1 peptide. Molecular dynamics simulations of the salt-bridged form of KR-CSH-ET were performed using both the GB/SA implicit solvation scheme or an explicit solvation and the particle-mesh Ewald method for long-range electrostatic calculation. Unexpectedly, the Arg(-)(1)-Asp(8) salt bridge does not display in the simulation the stability that could be expected from the experimental data. The cooperative involvement of a cation-pi interaction in formation of the salt bridge has been hypothesized. Difficulties in accurately simulating cation-pi interactions might be responsible for the lack of stability in the simulation. At this time, however, no definitive explanation for the observed discrepancy between experiments and simulations is available, and further experimental studies appear to be necessary to fully understand in atomic detail the pH-dependent conformational change observed in the KR-ET-1 series.

  7. Physico-chemical properties of R140G and K141Q mutants of human small heat shock protein HspB1 associated with hereditary peripheral neuropathies.

    Science.gov (United States)

    Nefedova, Victoria V; Datskevich, Petr N; Sudnitsyna, Maria V; Strelkov, Sergei V; Gusev, Nikolai B

    2013-08-01

    Some physico-chemical properties of R140G and K141Q mutants of human small heat shock protein HspB1 associated with hereditary peripheral neuropathy were analyzed. Mutation K141Q did not affect intrinsic Trp fluorescence and interaction with hydrophobic probe bis-ANS, whereas mutation R140G decreased both intrinsic fluorescence and fluorescence of bis-ANS bound to HspB1. Both mutations decreased thermal stability of HspB1. Mutation R140G increased, whereas mutation K141Q decreased the rate of trypsinolysis of the central part (residues 5-188) of HspB1. Both the wild type HspB1 and its K141Q mutant formed large oligomers with apparent molecular weight ∼560 kDa. The R140G mutant formed two types of oligomers, i.e. large oligomers tending to aggregate and small oligomers with apparent molecular weight ∼70 kDa. The wild type HspB1 formed mixed homooligomers with R140G mutant with apparent molecular weight ∼610 kDa. The R140G mutant was unable to form high molecular weight heterooligomers with HspB6, whereas the K141Q mutant formed two types of heterooligomers with HspB6. In vitro measured chaperone-like activity of the wild type HspB1 was comparable with that of K141Q mutant and was much higher than that of R140G mutant. Mutations of homologous hot-spot Arg (R140G of HspB1 and R120G of αB-crystallin) induced similar changes in the properties of two small heat shock proteins, whereas mutations of two neighboring residues (R140 and K141) induced different changes in the properties of HspB1.

  8. Identification of Specific Effect of Chloride on the Spectral Properties and Structural Stability of Multiple Extracellular Glutamic Acid Mutants of Bacteriorhodopsin

    Science.gov (United States)

    Lazarova, Tzvetana; Mlynarczyk, Krzysztof; Querol, Enric; Tenchov, Boris; Filipek, Slawomir; Padrós, Esteve

    2016-01-01

    In the present work we combine spectroscopic, DSC and computational approaches to examine the multiple extracellular Glu mutants E204Q/E194Q, E204Q/E194Q/E9Q and E204Q/E194Q/E9Q/E74Q of bacteriorhodopsin by varying solvent ionic strength and composition. Absorption spectroscopy data reveal that the absorption maxima of multiple EC Glu mutants can be tuned by the chloride concentration in the solution. Visible Circular dichroism spectra imply that the specific binding of Cl- can modulate weakened exciton chromophore coupling and reestablish wild type-like bilobe spectral features of the mutants. The DSC data display reappearance of the reversible thermal transition, higher Tm of denaturation and an increase in the enthalpy of unfolding of the mutants in 1 M KCl solutions. Molecular dynamics simulations indicate high affinity binding of Cl- to Arg82 and to Gln204 and Gln194 residues in the mutants. Analysis of the experimental data suggests that simultaneous elimination of the negatively charged side chain of Glu194 and Glu204 is the major cause for mutants’ alterations. Specific Cl- binding efficiently coordinates distorted hydrogen bonding interactions of the EC region and reconstitutes the conformation and structure stability of mutated bR in WT-like fashion. PMID:27657718

  9. Selective pressure of antibiotics on ARGs and bacterial communities in manure-polluted freshwater-sediment microcosms

    Directory of Open Access Journals (Sweden)

    Wenguang eXiong

    2015-03-01

    Full Text Available The aim of this study was to investigate selective pressure of antibiotics on antibiotic resistance genes (ARGs and bacterial communities in manure-polluted aquatic environment. Three treatment groups were set up in freshwater-sediment microcosms: tetracyclines group, sulfonamides group and fluoroquinolones group. Sediment and water samples were collected on day 14 after treatment. Antibiotic concentrations, ARGs abundances and bacterial community composition were analyzed. Antibiotic concentrations were determined by ultra-performance liquid chromatography-electrospray tandem mass spectrometry. ARGs abundances were quantified by real time quantitative PCR. Bacterial community composition was analyzed based on amplicon sequencing. Of the three classes of antibiotics analyzed in the treatment groups, accumulation amounts were tetracyclines> fluoroquinolone> sulfonamides in the sediment samples, while they were sulfonamides> fluoroquinolone> tetracyclines in the water samples. In the treatment groups, the relative abundances of some tet resistance genes (tet(W and tet(X and plasmid-mediated quinolone resistance (PMQR genes (oqx(B and aac(6’-Ib in sediment samples were significantly higher than those in the paired water samples. Tetracyclines significantly selected the bacterial classes including Gammaproteobacteria, Clostridia, and the genera including Salmonella, Escherichia/Shigella, Clostridium, Stenotrophomonas in sediment samples. The significant selection on bacterial communities posed by sulfonamides and fluoroquinolones was also observed. The results indicated that sediment may supply an ideal setting for maintenance and persistence of tet resistance genes (tet(W and tet(X and PMQR genes (oqx(B and aac(6’-Ib under antibiotic pollution. The results also highlighted that antibiotics significantly selected specific bacterial communities including the taxa associated with opportunistic pathogens.

  10. Synthesis of Fluorine-18 Labeled Glucose-Lys-Arg-Gly-Asp-D-Phe as a Potential Tumor Imaging Agent

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Kyo Chul; Kim, Ji Sun; Sung, Hyun Ju; Jung, Jae Ho; An, Gwang Il; Chi, Dae Yoon [Korea Institute of Radiological and Medical Sciences, Seoul (Korea, Republic of); Lee, Byung Chul; Moon, Byung Seok; Choi, Tae Hyun; Chuna, Kwon Soo [Inha Univ., Inchon (Korea, Republic of)

    2005-07-01

    The {alpha}{sub v}{beta}{sub 3} integrin is an important receptor affecting tumor growth, metastatic potential on proliferating endothelial cells as well as on tumor cells of various origin, tumor-induced angiogenesis could be blocked by antagonizing the {alpha}{sub v}{beta}{sub 3} integrin with RGD. Therefore, {alpha}{sub v}{beta}{sub 3} integrin is a target for angiogenesis imaging that might be useful in assessing tumor-induced angiogenesis and identifying tumor metastasis. To design potent radiotracer for imaging angiogenesis containing a cRGD moiety should include low hepatic uptake in vivo. Tripeptide Arg-Gly-Asp (RGD), naturally existed in extracellular matrix proteins, is known to be the primary binding site of the {alpha}{sub v}{beta}{sub 3} integrin. The imaging of {alpha}{sub v}{beta}{sub 3} receptor expression will give the information of the metastatic ability of the tumor which is not available by [{sup 18}F]FDG. Our interest in developing new radiopharmaceuticals for in vivo visualization of angiogenesis has led us to synthesize derivatives of cRGD (cyclic arginineglycine-aspartic acid) that contains glucose moiety. Because sugar-protein interaction is a key step in metastasis and angiogenesis, it has also been proposed to play an intriguing role in imaging of tumor. We designed and synthesized two fluorine-18 labeled RGD glycopeptides . N-fluorobenzyl-diaminobutane-N'-glucose-Lys-Arg-Gly-Asp-D-Phe ([{sup 18}F]fluorobenzyl-glucose-KRGDf, and Nfluorobenzoyl- diaminobutane-N'-glucose-Lys-Arg-Gly-Asp-D-Phe ([{sup 18}F]fluorobenzoyl-glucose-KRGDf, from same precursor as a diagnostic tumor imaging agent for positron emission tomography (PET). Fluorine-18 labeled cRGD glycopeptides were prepared using two different simple labeling methods: one is reductive alkylation of an amine with [{sup 18}F]fluorobenzaldehyde and the other is amide condensation with [{sup 18}F]fluorobenzoic acid.

  11. Identification of Minor Secondary Metabolites from the Latex of Croton lechleri (Muell-Arg) and Evaluation of Their Antioxidant Activity

    OpenAIRE

    Maria Iorizzi; Francesco Visioli; Gelsomina Fico; Sara Vitalini; Franco Zollo; Fulvio Gala; Simona De Marino

    2008-01-01

    Dragon’s blood (Sangre de drago), a viscous red sap derived from Croton lechleri Muell-Arg (Euphorbiaceae), is extensively used by indigenous cultures of the Amazonian basin for its wound healing properties. The aim of this study was to identify the minor secondary metabolites and test the antioxidant activity of this sustance. A bioguided fractionation of the n-hexane, chloroform, n-butanol, and aqueous extracts led to the isolation of 15 compounds: three megastigmanes, four flavan-3-ol...

  12. Association between TP53 gene Arg72Pro polymorphism and Wilms’ tumor risk in a Chinese population

    Directory of Open Access Journals (Sweden)

    Fu W

    2017-02-01

    Full Text Available Wen Fu,1,2,* Zhen-Jian Zhuo,3,* Wei Jia,1,2 Jinhong Zhu,4 Shi-Bo Zhu,1,2 Ze-Feng Lin,1,2 Feng-Hua Wang,1,2 Huimin Xia,1,2 Jing He,1,2 Guo-Chang Liu1,2 1Department of Pediatric Urology, Guangzhou Women and Children’s Medical Center, Guangzhou Medical University, Guangzhou, Guangdong, 2Department of Pediatric Surgery, Guangzhou Institute of Pediatrics, Guangzhou Women and Children’s Medical Center, Guangzhou Medical University, Guangzhou, Guangdong, 3Faculty of Medicine, School of Chinese Medicine, The Chinese University of Hong Kong, Hong Kong, 4Molecular Epidemiology Laboratory, Department of Laboratory Medicine, Harbin Medical University Cancer Hospital, Harbin, Heilongjiang, People’s Republic of China *These authors contributed equally to this work Abstract: Wilms’ tumor is one of the most prevalent pediatric malignancies, ranking fourth in childhood cancer worldwide. TP53 is a critical tumor suppressor gene, which encodes a 53 kDa protein, p53. The p53 functions to protect against cancer by regulating cell cycle and apoptosis and maintaining DNA integrity. TP53 gene is highly polymorphic. Several TP53 gene polymorphisms have been considered to be associated with cancer risk. Of them, a nonsynonymous polymorphism, Arg72Pro (rs1042522 C>G, has been most extensively studied for the association with cancer risk; however, few studies have investigated its effect on Wilms’ tumor. Because of the central role of p53 in cell cycle control, the TP53 gene Arg72Pro polymorphism is also a good potential candidate predisposition locus for this pediatric cancer. We genotyped this polymorphism in 145 patients and 531 cancer-free controls recruited from Chinese children by Taqman methodology. Overall, our result suggested a lack of association between the TP53 gene Arg72Pro polymorphism and Wilms’ tumor. In the stratified analysis, we found that carriers of CG/GG genotypes had a significantly increased Wilms’ tumor risk in children not older

  13. Dynamic features of carboxy cytoglobin distal mutants investigated by molecular dynamics simulations.

    Science.gov (United States)

    Zhao, Cong; Du, Weihong

    2016-04-01

    Cytoglobin (Cgb) is a member of hemoprotein family with roles in NO metabolism, fibrosis, and tumourigenesis. Similar to other hemoproteins, Cgb structure and functions are markedly influenced by distal key residues. The sixth ligand His(81) (E7) is crucial to exogenous ligand binding, heme pocket conformation, and physiological roles of this protein. However, the effects of other key residues on heme pocket and protein biological functions are not well known. In this work, a molecular dynamics (MD) simulation study of two single mutants in CO-ligated Cgb (L46FCgbCO and L46VCgbCO) and two double mutants (L46FH81QCgbCO and L46VH81QCgbCO) was conducted to explore the effects of the key distal residues Leu(46)(B10) and His(81)(E7) on Cgb structure and functions. Results indicated that the distal mutation of B10 and E7 affected CgbCO dynamic properties on loop region fluctuation, internal cavity rearrangement, and heme motion. The distal conformation change was reflected by the distal key residues Gln(62) (CD3) and Arg(84)(E10). The hydrogen bond between heme propionates with CD3 or E10 residues were evidently influenced by B10/E7 mutation. Furthermore, heme pocket rearrangement was also observed based on the distal pocket volume and occurrence rate of inner cavities. The mutual effects of B10 and E7 residues on protein conformational rearrangement and other dynamic features were expressed in current MD studies of CgbCO and its distal mutants, suggesting their crucial role in heme pocket stabilization, ligand binding, and Cgb biological functions. The mutation of distal B10 and E7 residues affects the dynamic features of carboxy cytoglobin.

  14. Lack of association between the Trp719Arg polymorphism in kinesin-like protein 6 and coronary artery disease in 19 case-control studies

    Science.gov (United States)

    Assimes, Themistocles L; Hólm, Hilma; Kathiresan, Sekar; Reilly, Muredach P; Thorleifsson, Gudmar; Voight, Benjamin F; Erdmann, Jeanette; Willenborg, Christina; Vaidya, Dhananjay; Xie, Changchun; Patterson, Chris C; Morgan, Thomas M; Burnett, Mary Susan; Li, Mingyao; Hlatky, Mark A; Knowles, Joshua W; Thompson, John R; Absher, Devin; Iribarren, Carlos; Go, Alan; Fortmann, Stephen P; Sidney, Stephen; Risch, Neil; Tang, Hua; Myers, Richard M; Berger, Klaus; Stoll, Monika; Shah, Svati H.; Thorgeirsson, Gudmundur; Andersen, Karl; Havulinna, Aki S; Herrera, J. Enrique; Faraday, Nauder; Kim, Yoonhee; Kral, Brian G.; Mathias, Rasika; Ruczinski, Ingo; Suktitipat, Bhoom; Wilson, Alexander F; Yanek, Lisa R.; Becker, Lewis C; Linsel-Nitschke, Patrick; Lieb, Wolfgang; König, Inke R; Hengstenberg, Christian; Fischer, Marcus; Stark, Klaus; Reinhard, Wibke; Winogradow, Janina; Grassl, Martina; Grosshennig, Anika; Preuss, Michael; Eifert, Sandra; Schreiber, Stefan; Wichmann, H-Erich; Meisinger, Christa; Yee, Jean; Friedlander, Yechiel; Do, Ron; Meigs, James B; Williams, Gordon; Nathan, David M; MacRae, Calum A; Qu, Liming; Wilensky, Robert L; Matthai, William H.; Qasim, Atif N; Hakonarson, Hakon; Pichard, Augusto D; Kent, Kenneth M; Satler, Lowell; Lindsay, Joseph M; Waksman, Ron; Knouff, Christopher W; Waterworth, Dawn M; Walker, Max C; Mooser, Vincent; Marrugat, Jaume; Lucas, Gavin; Subirana, Isaac; Sala, Joan; Ramos, Rafael; Martinelli, Nicola; Olivieri, Oliviero; Trabetti, Elisabetta; Malerba, Giovanni; Pignatti, Pier Franco; Guiducci, Candace; Mirel, Daniel; Parkin, Melissa; Hirschhorn, Joel N; Asselta, Rosanna; Duga, Stefano; Musunuru, Kiran; Daly, Mark J; Purcell, Shaun; Braund, Peter S; Wright, Benjamin J; Balmforth, Anthony J; Ball, Stephen G; Ouwehand, Willem H; Deloukas, Panos; Scholz, Michael; Cambien, Francois; Huge, Andreas; Scheffold, Thomas; Salomaa, Veikko; Girelli, Domenico; Granger, Christopher B.; Peltonen, Leena; McKeown, Pascal P; Altshuler, David; Melander, Olle; Devaney, Joseph M; Epstein, Stephen E; Rader, Daniel J; Elosua, Roberto; Engert, James C; Anand, Sonia S; Hall, Alistair S; Ziegler, Andreas; O’Donnell, Christopher J; Spertus, John A; Siscovick, David; Schwartz, Stephen M; Becker, Diane; Thorsteinsdottir, Unnur; Stefansson, Kari; Schunkert, Heribert; Samani, Nilesh J; Quertermous, Thomas

    2011-01-01

    Objectives We sought to replicate the association between the kinesin-like protein 6 (KIF6) Trp719Arg polymorphism (rs20455) and clinical coronary artery disease (CAD). Background Recent prospective studies suggest that carriers of the 719Arg allele in KIF6 are at increased risk of clinical CAD compared with non-carriers. Methods The KIF6 Trp719Arg polymorphism (rs20455) was genotyped in nineteen case-control studies of non-fatal CAD either as part of a genome-wide association study or in a formal attempt to replicate the initial positive reports. Results Over 17 000 cases and 39 000 controls of European descent as well as a modest number of South Asians, African Americans, Hispanics, East Asians, and admixed cases and controls were successfully genotyped. None of the nineteen studies demonstrated an increased risk of CAD in carriers of the 719Arg allele compared with non-carriers. Regression analyses and fixed effect meta-analyses ruled out with high degree of confidence an increase of ≥2% in the risk of CAD among European 719Arg carriers. We also observed no increase in the risk of CAD among 719Arg carriers in the subset of Europeans with early onset disease (<50 years of age for males and <60 years for females) compared with similarly aged controls as well as all non-European subgroups. Conclusions The KIF6 Trp719Arg polymorphism was not associated with the risk of clinical CAD in this large replication study. PMID:20933357

  15. Comparative functional characterization of novel non-syndromic GJB2 gene variant p.Gly45Arg and lethal syndromic variant p.Gly45Glu

    Science.gov (United States)

    Gordhandas, Jeenal A.; Pique, Lynn

    2016-01-01

    We characterized a novel GJB2 missense variant, c.133G>A, p.Gly45Arg, and compared it with the only other variant at the same amino acid position of the connexin 26 protein (Cx26) reported to date: c.134G>A, p.Gly45Glu. Whereas both variants are associated with hearing loss and are dominantly inherited, p.Gly45Glu has been implicated in the rare fatal keratitis-ichthyosis-deafness (KID) syndrome, which results in cutaneous infections and septicemia with premature demise in the first year of life. In contrast, p.Gly45Arg appears to be non-syndromic. Subcellular localization experiments in transiently co-transfected HeLa cells demonstrated that Cx26-WT (wild-type) and p.Gly45Arg form gap junctions, whereas Cx26-WT with p.Gly45Glu protein does not. The substitution of a nonpolar amino acid glycine in wildtype Cx26 at position 45 with a negatively charged glutamic acid (acidic) has previously been shown to interfere with Ca2+ regulation of hemichannel gating and to inhibit the formation of gap junctions, resulting in cell death. The novel variant p.Gly45Arg, however, changes this glycine to a positively charged arginine (basic), resulting in the formation of dysfunctional gap junctions that selectively affect the permeation of negatively charged inositol 1,4,5-trisphosphate (IP3) and contribute to hearing loss. Cx26 p.Gly45Arg transfected cells, unlike cells transfected with p.Gly45Glu, thrived at physiologic Ca2+ concentrations, suggesting that Ca2+ regulation of hemichannel gating is unaffected in Cx26 p.Gly45Arg transfected cells. Thus, the two oppositely charged amino acids that replace the highly conserved uncharged glycine in p.Gly45Glu and p.Gly45Arg, respectively, produce strikingly different effects on the structure and function of the Cx26 protein. PMID:27761313

  16. Muscle development in mdx mutant mice.

    Science.gov (United States)

    Dangain, J; Vrbova, G

    1984-01-01

    Mechanical and contractile properties of tibialis anterior (TA) muscles from X-linked muscular dystrophic (mdx) mutant mice at different stages of development are compared to those of muscles from normal control animals. There is no difference between the tension output, speeds of contraction and relaxation, and weight of TA muscles from mutant adults and normal control animals. However, it is found that in 3-4-week-old mutant animals, tension output and muscle weight are very much reduced, and half relaxation time is prolonged. Thus, during this stage of development, muscles from mdx mice do not function properly. Histological examination of these muscles provides further evidence that, in these animals, rapid muscle destruction occurs at a particular time of development and that it is followed by complete recovery. This new mutant therefore presents an interesting case of muscle destruction and rapid regeneration. However, it is not an adequate model for Duchenne muscular dystrophy.

  17. Targeting ESR1-Mutant Breast Cancer

    Science.gov (United States)

    2015-09-01

    AWARD NUMBER: W81XWH-14-1-0359 TITLE: Targeting ESR1-Mutant Breast Cancer PRINCIPAL INVESTIGATOR: Dr. Sarat Chandarlapaty CONTRACTING...31 Aug 2015 4. TITLE AND SUBTITLE Targeting ESR1-Mutant Breast Cancer 5a. CONTRACT NUMBER 5b. GRANT NUMBER W81XWH-14-1-0359 5c. PROGRAM ELEMENT...mutations found in breast cancer using both structural and cell based assays. We have now have evidence for the effects of the most recurrent

  18. Targeting ESR1-Mutant Breast Cancer

    Science.gov (United States)

    2015-09-01

    Introduction Approximately 70% of ER+ breast cancers harbor expression of the estrogen receptor and are dependent upon its activity for various aspects of the...resistance to current FDA approved ER antagonists, but that more potent and selective estrogen receptor antagonists will be sufficiently active to...antagonists and their potency against ER mutants both in vitro and in vivo . Targeting ESR1-Mutant Breast Cancer W81XWH-14-1-0359 9 4. Impact A) Impact

  19. A Review on the Salt Bridge Between ASP177 and ARG163 of Wild-Type Rabbit Prion Protein

    CERN Document Server

    Zhang, Jiapu

    2014-01-01

    Prion diseases are invariably fatal and highly infectious neurodegenerative diseases that affect a wide variety of mammalian species such as sheep and goats, cattle, deer, elks, humans and mice etc., but rabbits have a low susceptibility to be infected by prion diseases with respect to other species. The stability of rabbit prion protein is due to its highly ordered beta2-alpha2 loop [PLoS One 5 (10) e13273 (2010); Journal of Biological Chemistry 285 (41) 31682-31693 (2010)] and a helix-capping motif within this loop [PLoS One 8 (5) e63047 (2013)]. The beta2-alpha2 loop has been a focus in prion studies. For this loop we found a salt bridge linkage ASP177-ARG163 (O-N) [Journal of Theoretical Biology 342 (7 February 2014) 70-82 (2014)]. Some scientists said on the 2FJ3.pdb NMR file of the rabbit prion protein, the distance of ASP177-ARG163 (O-N) gives the salt bridge of about 10 angstroms which is nearly null in terms of energy thus think our result is wrong. This opinion is clearly wrong simply due to the 3O7...

  20. The Lys234Arg Substitution in the Enzyme SHV-72 Is a Determinant for Resistance to Clavulanic Acid Inhibition▿

    Science.gov (United States)

    Mendonça, Nuno; Manageiro, Vera; Robin, Frédéric; Salgado, M. José; Ferreira, Eugénia; Caniça, Manuela; Bonnet, Richard

    2008-01-01

    The new β-lactamase SHV-72 was isolated from clinical Klebsiella pneumoniae INSRA1229, which exhibited the unusual association of resistance to the amoxicillin-clavulanic acid combination (MIC, 64 μg/ml) and susceptibility to cephalosporins, aztreonam, and imipenem. SHV-72 (pI 7.6) harbored the three amino acid substitutions Ile8Phe, Ala146Val, and Lys234Arg. SHV-72 had high catalytic efficiency against penicillins (kcat/Km, 35 to 287 μM−1·s−1) and no activity against oxyimino β-lactams. The concentration of clavulanic acid necessary to inhibit the enzyme activity by 50% was 10-fold higher for SHV-72 than for SHV-1. Molecular-dynamics simulation suggested that the Lys234Arg substitution in SHV-72 stabilized an atypical conformation of the Ser130 side chain, which moved the Oγ atom of Ser130 around 3.5 Å away from the key Oγ atom of the reactive serine (Ser70). This movement may therefore decrease the susceptibility to clavulanic acid by preventing cross-linking between Ser130 and Ser70. PMID:18316518

  1. Cationic amino acid transporters and Salmonella Typhimurium ArgT collectively regulate arginine availability towards intracellular Salmonella growth.

    Directory of Open Access Journals (Sweden)

    Priyanka Das

    Full Text Available Cationic amino acid transporters (mCAT1 and mCAT2B regulate the arginine availability in macrophages. How in the infected cell a pathogen can alter the arginine metabolism of the host remains to be understood. We reveal here a novel mechanism by which Salmonella exploit mCAT1 and mCAT2B to acquire host arginine towards its own intracellular growth within antigen presenting cells. We demonstrate that Salmonella infected bone marrow derived macrophages and dendritic cells show enhanced arginine uptake and increased expression of mCAT1 and mCAT2B. We show that the mCAT1 transporter is in close proximity to Salmonella containing vacuole (SCV specifically by live intracellular Salmonella in order to access the macrophage cytosolic arginine pool. Further, Lysosome associated membrane protein 1, a marker of SCV, also was found to colocalize with mCAT1 in the Salmonella infected cell. The intra vacuolar Salmonella then acquire the host arginine via its own arginine transporter, ArgT for growth. The argT knockout strain was unable to acquire host arginine and was attenuated in growth in both macrophages and in mice model of infection. Together, these data reveal survival strategies by which virulent Salmonella adapt to the harsh conditions prevailing in the infected host cells.

  2. A novel role for the fibrinogen Asn-Gly-Arg (NGR) motif in platelet function.

    Science.gov (United States)

    Moriarty, Róisín; McManus, Ciara A; Lambert, Matthew; Tilley, Thea; Devocelle, Marc; Brennan, Marian; Kerrigan, Steven W; Cox, Dermot

    2015-02-01

    The integrin αIIbβ3 on resting platelets can bind to immobilised fibrinogen resulting in platelet spreading and activation but requires activation to bind to soluble fibrinogen. αIIbβ3 is known to interact with the general integrin-recognition motif RGD (arginine-glycine-aspartate) as well as the fibrinogen-specific γ-chain dodecapeptide; however, it is not known how fibrinogen binding triggers platelet activation. NGR (asparagine-glycine-arginine) is another integrin-recognition sequence present in fibrinogen and this study aims to determine if it plays a role in the interaction between fibrinogen and αIIbβ3. NGR-containing peptides inhibited resting platelet adhesion to fibrinogen with an IC50 of 175 µM but failed to inhibit the adhesion of activated platelets to fibrinogen (IC50> 500 µM). Resting platelet adhesion to mutant fibrinogens lacking the NGR sequences was reduced compared to normal fibrinogen under both static and shear conditions (200 s⁻¹). However, pre-activated platelets were able to fully spread on all types of fibrinogen. Thus, the NGR motif in fibrinogen is the site that is primarily responsible for the interaction with resting αIIbβ3 and is responsible for triggering platelet activation.

  3. The Effect of Selective D- or Nα-Methyl Arginine Substitution on the Activity of the Proline-Rich Antimicrobial Peptide, Chex1-Arg20

    Science.gov (United States)

    Li, Wenyi; Sun, Zhe; O'Brien-Simpson, Neil M.; Otvos, Laszlo; Reynolds, Eric C.; Hossain, Mohammed A.; Separovic, Frances; Wade, John D.

    2017-01-01

    In vivo pharmacokinetics studies have shown that the proline-rich antimicrobial peptide, A3-APO, which is a discontinuous dimer of the peptide, Chex1-Arg20, undergoes degradation to small fragments at positions Pro6-Arg7 and Val19-Arg20. With the aim of minimizing or abolishing this degradation, a series of Chex1-Arg20 analogs were prepared via Fmoc/tBu solid phase peptide synthesis with D-arginine or, in some cases, peptide backbone Nα-methylated arginine, substitution at these sites. All the peptides were tested for antibacterial activity against the Gram-negative bacterium Klebsiella pneumoniae. The resulting activity of position-7 substitution of Chex1-Arg20 analogs showed that arginine-7 is a crucial residue for maintaining activity against K. pneumoniae. However, arginine-20 substitution had a much less deleterious effect on the antibacterial activity of the peptide. Moreover, none of these peptides displayed any cytotoxicity to HEK and H-4-II-E mammalian cells. These results will aid the development of more effective and stable PrAMPs via judicious amino acid substitutions. PMID:28154813

  4. The effect of selective D- or N(alpha)-methyl arginine substitution on the activity of the proline-rich antimicrobial peptide, Chex1-Arg20.

    Science.gov (United States)

    Li, Wenyi; Sun, Zhe; O'Brien-Simpson, Neil M.; Otvos, Laszlo; Reynolds, Eric C.; Hossain, Mohammed A.; Separovic, Frances; Wade, John D.

    2017-01-01

    In vivo pharmacokinetics studies have shown that the proline-rich antimicrobial peptide, A3-APO, which is a discontinuous dimer of the peptide, Chex1-Arg20, undergoes degradation to small fragments at positions Pro6-Arg7 and Val19-Arg20. With the aim of minimizing or abolishing this degradation, a series of Chex1-Arg20 analogues were prepared via Fmoc/tBu solid phase peptide synthesis with D-arginine or, in some cases, peptide backbone N-methylated arginine, substitution at these sites. All the peptides were tested for antibacterial activity against the Gram-negative bacterium Klebsiella pneumoniae. The resulting activity of position-7 substitution of Chex1-Arg20 analogues showed that arginine-7 is a crucial residue for maintaining activity against K. pneumoniae. However, arginine-20 substitution had a much less deleterious effect on the antibacterial activity of the peptide. Moreover, none of these peptides displayed any cytotoxicity to HEK and H-4-II-E mammalian cells. These results will aid the development of more effective and stable PrAMPs via judicious amino acid substitutions.

  5. Correlation between SULT1A1 Arg213His Gene Polymorphisms and Uterine Leiomyomas%SULT1A1基因Arg213His 位点多态性与子宫肌瘤发生的关联性

    Institute of Scientific and Technical Information of China (English)

    周超; 林林; 张英姿; 徐天和; 张磊磊

    2011-01-01

    目的:探讨硫酸氨基转移酶(Sulfotransferase,SULT)1A1基因Arg213His位点多态性与鲁北地区汉族女性子宫肌瘤的关系.方法:以病例-对照的研究方法,采用聚合酶链式反应-限制性片段长度多态性(PCR-RFLP)方法检测了123例子宫肌瘤患者和123例匹配对照者的SULT1A1基因Arg213His位点的基因型,应用条件Logistic回归等方法分析基因多态性与子宫肌瘤的关系.结果:1)SULT1A1基因Arg/Arg、Arg/His、His/His 3种基因型在子宫肌瘤与对照组中的分布存在显著性差异(P=0.011);2)与Arg/Arg基因型相比,Arg/His、His/His危险度均增加,分别为2.321倍和1.985倍(P=0.003和P=0.468);3)His等位基因可显著增加患子宫肌瘤的危险性(P=0.003,OR调整=2.296,95%CI为1.325~3.978).结论:SULT1A1基因Arg213His位点基因多态性与鲁北地区汉族女性子宫肌瘤的发生有关,增加了子宫肌瘤的患病风险.%Objective: To study the correlation between polymorphisms of SULT1A1 with the risk of uterine leiomyoma interaction among Han Chinese in Northern QiLu.Methods: Arg213His genotypes of the SULT1A1 gene were detected using polymerase chain reaction-restriction fragment length polymorphism in a case-control study.A total of 123 cases of uterine leiomyomas and 123 controls were included.Multivariate logistic regression analysis was used to estimate the risk of developing uterine leiomyomas associated with environmental exposures of the SULT1A1 genotype.Results: ( 1 ) The SULT1A1 polymorphisms of the Arg/Arg, Arg/His, and His/His genotypes in the uterine leiomyoma group and the control group were significantly different ( P = 0.011 ); ( 2 ) Comparedwith that of the Arg/Arg genotype, the risk of the Arg/His was genotype and His/His was both increased, by 2.321 times and 1.985 times ( P = 0.003 and P = 0.468 ).( 3 ) The risk was significantly higher among uterine leiomyoma patients with the His allele ( OR adjusted = 2.296, 95 % CI 1.325-3.978, P = 0

  6. 大位阻氨基酸Fmoc-Arg(Pbf)-OH与Rink Amide-AM树脂的高效缩合%Highly efficient condensation of sterically hindered amino acid fmoc-arg(Pbf)-OH and rink amide-AM resin

    Institute of Scientific and Technical Information of China (English)

    朱亮亮; 绳则翠; 周成; 祝社民; 陈英文; 沈树宝

    2012-01-01

    The coupling reactions of Fmoc-Arg( Pbf) -OH and Rink Amide-AM Resin with symmetrical anhydride method, active ester method and 2,6-dichlorobenoyl chloride (DCB) method, respectively, were investigated in a self-designed reactor which congregated the stirring,filtration,and bubbling function together. The effects of reaction strategy, solvent system, reaction time, molar ratio of reactants and stirring method on the yield of condensation reaction were performed. The results indicate that the DIC/HOBt/DMAP strategy is the best method. The optimal reaction conditions are shown as follows: 3 : 1 of molar ratio of the reactants, 3 h of reaction time and DMA/DCM ( 1 : 1, V/ V) as a solvent using nitrogen-assisted magnetic stir system. The highest yield of the condensation reaction is 93% .%采用集自动搅拌、过滤、鼓泡等多重功能于一体的自制多肽固相合成反应器,以对称酸酐法、活化酯法、2,6-二氯苯甲酰氯(DCB)法研究了大位阻氨基酸Fmoc-Arg (Pbf)-OH与Rink Amide-AM树脂的连接反应工艺.探讨了催化体系、溶剂体系、反应时间、反应物配比以及搅拌方式对合成Fmoc-Arg(Pbf)-Rink Amide-AM树脂反应的影响.结果表明,采用活化酯法(DIC/HOBt/DMAP)时连接率最高,最佳反应条件为:在采用N2辅助磁力拌系统,以体积比为1∶1的DMA/DCM为反应溶剂,氨基酸与树脂物质的量的比为3∶1,反应时间为3h时,连接率高达93%.

  7. Characterisation of a C1qtnf5 Ser163Arg knock-in mouse model of late-onset retinal macular degeneration.

    Directory of Open Access Journals (Sweden)

    Xinhua Shu

    Full Text Available A single founder mutation resulting in a Ser163Arg substitution in the C1QTNF5 gene product causes autosomal dominant late-onset retinal macular degeneration (L-ORMD in humans, which has clinical and pathological features resembling age-related macular degeneration. We generated and characterised a mouse "knock-in" model carrying the Ser163Arg mutation in the orthologous murine C1qtnf5 gene by site-directed mutagenesis and homologous recombination into mouse embryonic stem cells. Biochemical, immunological, electron microscopic, fundus autofluorescence, electroretinography and laser photocoagulation analyses were used to characterise the mouse model. Heterozygous and homozygous knock-in mice showed no significant abnormality in any of the above measures at time points up to 2 years. This result contrasts with another C1qtnf5 Ser163Arg knock-in mouse which showed most of the features of L-ORMD but differed in genetic background and targeting construct.

  8. Studies of associations between the Arg389Gly polymorphism of the beta1-adrenergic receptor gene (ADRB1) and hypertension and obesity in 7677 Danish white subjects

    DEFF Research Database (Denmark)

    Gjesing, A P; Andersen, G; Albrechtsen, A

    2007-01-01

    Activation of the beta(1)-adrenergic receptor (ADRB1) causes increased lipolysis in adipose tissue and enhances cardiac output. Analysis of the association of the functional ADRB1 Arg389Gly variant with obesity and hypertension has given ambiguous results. To clarify the potential impact of this ......Activation of the beta(1)-adrenergic receptor (ADRB1) causes increased lipolysis in adipose tissue and enhances cardiac output. Analysis of the association of the functional ADRB1 Arg389Gly variant with obesity and hypertension has given ambiguous results. To clarify the potential impact...... of this variant on obesity and hypertension in the general population, we examined the Arg389Gly variant in a relatively large-scale population-based study....

  9. 纤维蛋白原α链Arg16His突变导致遗传性异常纤维蛋白原血症%Inherited dysfibrinogenemia caused by Arg16His mutation in α chain of fibrinogen

    Institute of Scientific and Technical Information of China (English)

    赵小娟; 阮长耿; 王兆钺; 江明华; 张威; 曹丽娟; 马珍妮; 董宁征; 白霞; 余自强

    2010-01-01

    Objective To analyze the phenotype and genotype of a family with inherited dysfibrinogenemia.Methods Assays of coagulation,including activated partial thromboplastin time(APIT),prothrombin time(PT)and thrombin time(TT),were carried out with Stago Compact in the proband and his family members.The activity and antigen of fibrinogen in plasma were determined by Clanss and immunotur bidimetry,respectively.Fibrinogen and its constituent were analyzed by Western blot with nonreducing 4%-20% SDS-polyacrylamide gel electrophoresis(PAGE).All exons and exon-intron boundaries of fibringen genes FGA,FGB and FGG were analyzed by PCR and then direct sequencing.Results The proband had normal APTT and PT.but prolonged TT.The activity of fibrinogen in plasma wag decreased while its antigen level was normal.These abnormalities were also found in his mother and a sister.Genetic analysis revealed heterozygous G1233A in the exon 2 of FGA originating from his mother.which resulted in Arg16His missense mutation.Conclusion Inherited dysfibrinogenemia was caused by Arg16His mutation in exon 2 of FGA,and this iS the first case reported in a Chinese family.%目的 对1例遗传性异常纤维蛋白原血症家系进行表型和基因型分析.方法 用血凝仪检测先证者家系3代6人外周血活化部分凝血酶时间(APTT)、凝血酶原时间(PT)和凝血酶时间(TT).纤维蛋白原活性和抗原分别用Clauss法和免疫比浊法检测,Western blot检测血浆纤维蛋白原及其片段分布.PCR扩增纤维蛋白原基因FGA、FGB和FGG所有外显子及其侧翼序列,PCR产物纯化后直接测序进行基因分析.结果 先证者APTT、PT正常,而TT明显延长;纤维蛋白原抗原正常,而纤维蛋白原活性降低,先证者母亲和胞姐表型与之相似.基因分析显示先证者纤维蛋白原FGA基因2号外显子g1233→a杂合碱基改变(密码子GGT→CAT),导致Arg16His错义突变,该突变来源于母系.结论 纤维蛋白原α链Arg16His杂合错义

  10. Phanerochaete mutants with enhanced ligninolytic activity

    Energy Technology Data Exchange (ETDEWEB)

    Kakar, S.N.; Perez, A.; Gonzales, J.

    1993-06-01

    In addition to lignin, the white rot fungus Phanerochaete chrysosporium has the ability to degrade a wide spectrum of recalcitrant organopollutants in soils and aqueous media. Although some of the organic compounds are degraded under nonligninolytic conditions, most are degraded under ligninolytic conditions with the involvement of the extracellular enzymes, lignin peroxidases, and manganese-dependent peroxidases, which are produced as secondary metabolites triggered by conditions of nutrient starvation (e.g., nitrogen limitation). The fungus and its enzymes can thus provide alternative technologies for bioremediation, biopulping, biobleaching, and other industrial applications. The efficiency and effectiveness of the fungus can be enhanced by increasing production and secretion of the important enzymes in large quantities and as primary metabolites under enriched conditions. One way this can be achieved is through isolation of mutants that are deregulated or are hyperproducers or supersecretors of key enzymes under enriched conditions. Through ultraviolet-light and gamma-rays mutagenesis we have isolated a variety of mutants, some of which produce key enzymes of the ligninolytic system under high-nitrogen growth conditions. One of the mutants produced 272 units (U) of lignin peroxidases enzyme activity per liter after nine days under high nitrogen. The mutant and the parent strains produced up to 54 U/L and 62 U/L, respectively, of the enzyme activity under low-nitrogen growth conditions during this period. In some experiments the mutant showed 281 U/L of enzyme activity under high nitrogen after 17 days.

  11. Gene encoding γ-carbonic anhydrase is cotranscribed with argC and induced in response to stationary phase and high CO2 in Azospirillum brasilense Sp7

    Directory of Open Access Journals (Sweden)

    Mishra Mukti N

    2010-07-01

    Full Text Available Abstract Background Carbonic anhydrase (CA is a ubiquitous enzyme catalyzing the reversible hydration of CO2 to bicarbonate, a reaction underlying diverse biochemical and physiological processes. Gamma class carbonic anhydrases (γ-CAs are widespread in prokaryotes but their physiological roles remain elusive. At present, only γ-CA of Methanosarcina thermophila (Cam has been shown to have CA activity. Genome analysis of a rhizobacterium Azospirillum brasilense, revealed occurrence of ORFs encoding one β-CA and two γ-CAs. Results One of the putative γ-CA encoding genes of A. brasilense was cloned and overexpressed in E. coli. Electrometric assays for CA activity of the whole cell extracts overexpressing recombinant GCA1 did not show CO2 hydration activity. Reverse transcription-PCR analysis indicated that gca1 in A. brasilense is co-transcribed with its upstream gene annotated as argC, which encodes a putative N-acetyl-γ-glutamate-phosphate reductase. 5'-RACE also demonstrated that there was no transcription start site between argC and gca1, and the transcription start site located upstream of argC transcribed both the genes (argC-gca1. Using transcriptional fusions of argC-gca1 upstream region with promoterless lacZ, we further demonstrated that gca1 upstream region did not have any promoter and its transcription occurred from a promoter located in the argC upstream region. The transcription of argC-gca1 operon was upregulated in stationary phase and at elevated CO2 atmosphere. Conclusions This study shows lack of CO2 hydration activity in a recombinant protein expressed from a gene predicted to encode a γ-carbonic anhydrase in A. brasilense although it cross reacts with anti-Cam antibody raised against a well characterized γ-CA. The organization and regulation of this gene along with the putative argC gene suggests its involvement in arginine biosynthetic pathway instead of the predicted CO2 hydration.

  12. Genotyping and meta-analysis of KIF6 Trp719Arg polymorphism in South Indian Coronary Artery Disease patients: A case–control study

    Directory of Open Access Journals (Sweden)

    Durairajpandian Vishnuprabu

    2015-09-01

    Full Text Available The KIF6 719Arg allele is an interesting genomic variant widely screened in various populations and is reported to be associated with the risk of Coronary Artery Disease (CAD and statin treatment outcome. Recent population based clinical studies and large-scale meta-analyses pondered over the role of 719Arg variant in CAD risk and treatment response. We screened the KIF6 Trp719Arg polymorphism (rs20455 in south Indian CAD patients in a case–control approach. A total of 1042 samples (510 CAD patients and 532 controls were screened for the KIF6 Trp719Arg SNP by TaqMan SNP genotyping assay, followed by meta-analysis of the genotype data of non-Europeans reports. The 719Arg risk genotype (GG was observed in 29.6% of CAD cases and in 30.1% of controls with an odds ratio (OR of 1.07 (95% CI: 0.76–1.50, p value = 0.709. No significant difference in the genotype frequency was observed between CAD and controls in both dominant model (AG + GG vs AA and allelic model (719Arg vs 719Trp with an OR of 1.11 (p = 0.491 and 1.03 (p = 0.767, respectively. The covariate analysis indicated that smoking & alcohol consumption increased the risk for MI among CAD patients. Meta-analysis showed that the KIF6 719Arg allele is not associated with CAD risk in both fixed effect (p = 0.515, OR = 1.023, 95% CI = 0.956–1.094 and random effect (p = 0.547, OR = 1.022, 95% CI = 0.953–1.096. The symmetrical shape of the Egger's funnel plots revealed that there is no publication bias. These results suggest that there is no association of KIF6 719Arg allele with CAD risk in South Indian population and the meta-analysis confirms the same among non-European population.

  13. Enhanced cellulase production in mutants of Thermomonospora

    Energy Technology Data Exchange (ETDEWEB)

    Fennington, G.; Lupo, D.; Stutzenberger, F.

    1982-01-01

    Thermomonospora curvata, a thermophilic actinomycete, secretes multiple forms of endo-beta, 1-4-glucanase (EG) when grown on cellulose-mineral salts liquid medium. The EG activity (measured as carboxymethyl cellulose hydrolysis) was separated by ion exchange chromatography into three distinct components which differed in their kinetic properties. Exposure of T. curvata to ultraviolet light, N-nitrosoguanidine, or ethane methyl sulfonate produced mutants with enhanced EG production. Selection of colonies which cleared cellulose agar plants containing 2-deoxyglucose or glycerol yielded mutants having 1.5 to 2.6 times the extracellular EG and saccharifying activity (measured by filter-paper and cotton-fiber hydrolysis). The secretion of extracellular protein was increased proportionally in mutant cultures. (Refs. 40).

  14. High Persister Mutants in Mycobacterium tuberculosis.

    Directory of Open Access Journals (Sweden)

    Heather L Torrey

    Full Text Available Mycobacterium tuberculosis forms drug-tolerant persister cells that are the probable cause of its recalcitrance to antibiotic therapy. While genetically identical to the rest of the population, persisters are dormant, which protects them from killing by bactericidal antibiotics. The mechanism of persister formation in M. tuberculosis is not well understood. In this study, we selected for high persister (hip mutants and characterized them by whole genome sequencing and transcriptome analysis. In parallel, we identified and characterized clinical isolates that naturally produce high levels of persisters. We compared the hip mutants obtained in vitro with clinical isolates to identify candidate persister genes. Genes involved in lipid biosynthesis, carbon metabolism, toxin-antitoxin systems, and transcriptional regulators were among those identified. We also found that clinical hip isolates exhibited greater ex vivo survival than the low persister isolates. Our data suggest that M. tuberculosis persister formation involves multiple pathways, and hip mutants may contribute to the recalcitrance of the infection.

  15. Escherichia coli mutants with a temperature-sensitive alcohol dehydrogenase.

    OpenAIRE

    Lorowitz, W; Clark, D.

    1982-01-01

    Mutants of Escherichia coli resistant to allyl alcohol were selected. Such mutants were found to lack alcohol dehydrogenase. In addition, mutants with temperature-sensitive alcohol dehydrogenase activity were obtained. These mutations, designated adhE, are all located at the previously described adh regulatory locus. Most adhE mutants were also defective in acetaldehyde dehydrogenase activity.

  16. Native Mutant Huntingtin in Human Brain

    Science.gov (United States)

    Sapp, Ellen; Valencia, Antonio; Li, Xueyi; Aronin, Neil; Kegel, Kimberly B.; Vonsattel, Jean-Paul; Young, Anne B.; Wexler, Nancy; DiFiglia, Marian

    2012-01-01

    Huntington disease (HD) is caused by polyglutamine expansion in the N terminus of huntingtin (htt). Analysis of human postmortem brain lysates by SDS-PAGE and Western blot reveals htt as full-length and fragmented. Here we used Blue Native PAGE (BNP) and Western blots to study native htt in human postmortem brain. Antisera against htt detected a single band broadly migrating at 575–850 kDa in control brain and at 650–885 kDa in heterozygous and Venezuelan homozygous HD brains. Anti-polyglutamine antisera detected full-length mutant htt in HD brain. There was little htt cleavage even if lysates were pretreated with trypsin, indicating a property of native htt to resist protease cleavage. A soluble mutant htt fragment of about 180 kDa was detected with anti-htt antibody Ab1 (htt-(1–17)) and increased when lysates were treated with denaturants (SDS, 8 m urea, DTT, or trypsin) before BNP. Wild-type htt was more resistant to denaturants. Based on migration of in vitro translated htt fragments, the 180-kDa segment terminated ≈htt 670–880 amino acids. If second dimension SDS-PAGE followed BNP, the 180-kDa mutant htt was absent, and 43–50 kDa htt fragments appeared. Brain lysates from two HD mouse models expressed native full-length htt; a mutant fragment formed if lysates were pretreated with 8 m urea + DTT. Native full-length mutant htt in embryonic HD140Q/140Q mouse primary neurons was intact during cell death and when cell lysates were exposed to denaturants before BNP. Thus, native mutant htt occurs in brain and primary neurons as a soluble full-length monomer. PMID:22375012

  17. Aging Kit mutant mice develop cardiomyopathy.

    Directory of Open Access Journals (Sweden)

    Lei Ye

    Full Text Available Both bone marrow (BM and myocardium contain progenitor cells expressing the c-Kit tyrosine kinase. The aims of this study were to determine the effects of c-Kit mutations on: i. myocardial c-Kit(+ cells counts and ii. the stability of left ventricular (LV contractile function and structure during aging. LV structure and contractile function were evaluated (echocardiography in two groups of Kit mutant (W/Wv and W41/W42 and in wild type (WT mice at 4 and 12 months of age and the effects of the mutations on LV mass, vascular density and the numbers of proliferating cells were also determined. In 4 month old Kit mutant and WT mice, LV ejection fractions (EF and LV fractional shortening rates (FS were comparable. At 12 months of age EF and FS were significantly decreased and LV mass was significantly increased only in W41/W42 mice. Myocardial vascular densities and c-Kit(+ cell numbers were significantly reduced in both mutant groups when compared to WT hearts. Replacement of mutant BM with WT BM at 4 months of age did not prevent these abnormalities in either mutant group although they were somewhat attenuated in the W/Wv group. Notably BM transplantation did not prevent the development of cardiomyopathy in 12 month W41/W42 mice. The data suggest that decreased numbers and functional capacities of c-Kit(+ cardiac resident progenitor cells may be the basis of the cardiomyopathy in W41/W42 mice and although defects in mutant BM progenitor cells may prove to be contributory, they are not causal.

  18. Behavioral characterization of system xc- mutant mice.

    Science.gov (United States)

    McCullagh, Elizabeth A; Featherstone, David E

    2014-05-15

    The slc7a11 gene encodes xCT, an essential component of 'system xc-', a plasma membrane exchanger that imports cystine and exports glutamate. Slc7a11 is expressed primarily in the brain, but its role there is not clear. We performed behavioral tests on two different strains of homozygous slc7a11 mutant mice ('sut' and 'xCT'), as well as heteroallelic offspring of these two strains ('xCT/sut') and their associated genetic backgrounds. Homozygous sut mutant males showed reduced spontaneous alternation in spontaneous alternation tasks as well as reduced movement in an open field maze, but xCT and xCT/sut strains did not show significant changes in these tasks compared to appropriate controls. Neither xCT nor sut mutants showed differences from controls in rotarod tests. Female behavioral phenotypes were independent of estrus cycle stage. To ensure that homozygous xCT, sut, and xCT/sut strains all represent protein null alleles, we measured whole brain xCT protein levels using immunoblots. xCT, sut and xCT/sut strains showed no detectable xCT protein expression, confirming them as null alleles. Previously published microdialysis experiments showed reduced striatal glutamate in xCT mutants. Using the same methods, we measured reduced interstitial glutamate levels in the striatum but not cerebellum of sut mutants. However, we detected no glutamate change in the striatum or cerebellum of sut/xCT mice. We detected no changes in whole brain EAAT-1, -2, or -3 expression. We conclude that the behavioral and chemical differences exist between slc7a11 mutant strains, but we were unable to definitively attribute any of these differences to loss of system xc-.

  19. COMPARATIVE STUDY OF TOURIST TRAFFIC AT THE CITY, COUNTY AND HOUSE IN CÂMPULUNG MUSCEL ARGES

    Directory of Open Access Journals (Sweden)

    Adelaida Cristina HONTUŞ

    2013-01-01

    Full Text Available This paper is based on a comparative analysis of tourism demand and supply and the development of mountain tourism tendinţelorde plannaţional and internationally, the degree of capitalization of the mountain tourism in Romania, focusing on assessment of tourism in the tourist resort Campulung Muscel County Arges. The objective of this study consists of a comparative analysis of tourist traffic at the county level tourist resort and tourist board level. To perform this study we used a series of statistical data provided by the representatives of the City of Delhi Muscel on tourism and tourist traffic statistics on the county level and at the boarding house.

  20. Ornithological researches on the Goleşti Dam Lake (Argeş county, Romania during 2003 - 2010

    Directory of Open Access Journals (Sweden)

    Denisa CONETE

    2012-05-01

    Full Text Available 199 bird species were observed in the Goleşti Dam Lake, from the middle hydrographical basin of the Argeş River, during 2003 – 2010 (minimum – 64 in January and maximum – 141 in April. Their distribution regarding the constancy, dominancy and Dzuba index of ecological signification were considered. There are two eudominant species (Anas platyrhynchos and Aythya ferina and two dominant species (Aythya fuligula and Larus ridibundus. The variation in the number of individuals during the year is strongly determined by the temperature variation that leads to the diminishing or increasing of the food resources. In winter, when the temperatures were low in the north (and the waters froze, the birds arrived in great number in our area but only a few of them remained in the area for breeding.

  1. Evaluation of analgesic and anti-inflammatory effects of aerial parts of Breynia rhamnoides Muell.-Arg.

    Directory of Open Access Journals (Sweden)

    Shadiya C. K.

    2015-09-01

    Full Text Available The plant Breynia rhamnoides Muell. Arg (family-Phyllanthaceae/Euphorbiaceae was reported to have numerous potent constituents and the same plant was used for many traditional medicines. Phytochemical analysis of ethanolic extracts of Breynia rhamnoides revealed the presence of many active compounds such as flavonoids, glycosides, phenolic compounds, terpenoids, steroids and saponins. Present work describes the extraction and evaluation of analgesic and anti-inflammatory activities of aerial part extract of Breynia rhamnoides. Acute toxicity studies were performed as per OECD-423 guidelines. Toxicity signs and symptoms were not observed. Anti-inflammatory activity was evaluated by carrageenan-induced paw edema model and analgesic activity by Eddy’s hot plate method and Acetic acid induced writhing method in Wistar albino rats. The extract exhibited significant anti- inflammatory and peripheral analgesic activity.

  2. Apert Syndrome: Molecularly Confirmed C.758C>G (P.Pro253Arg) in FGFR2

    Energy Technology Data Exchange (ETDEWEB)

    Cha Gon, Lee, E-mail: leechagon@eulji.ac.kr [Department of Pediatrics, Eulji General Hospital, College of Medicine, Eulji University, 68 Hangeulbiseok-ro, Nowon-gu, Seoul 139-711 (Korea, Republic of)

    2016-03-21

    A 5-day-old girl was referred to our clinic for evaluation of congenital malformations. She was identified with a pathogenic mutation c.758C>G (p.Pro253Arg) in FGFR2 gene using targeted exome sequencing. The de novo mutation was confirmed with Sanger sequencing in the patient and her parents. She showed occipital plagiocephaly with frontal bossing (Figure A and B). Skull frontal and lateral radiography revealed fusion of most of the sutures except coronal suture, with convolutional markings (Figure D and E). She had complete cleft palate (Figure C). Her fused bilateral hands showed type II syndactyly with complete syndactyly between the ring and the little fingers (Figure F1-F3). Both toes were simple syndactyly with side-to-side fusion of skin (Figure G1-)

  3. Congenital erythropoietic porphyria with two mutations of the uroporphyrinogen III synthase gene (Cys73Arg, Thr228Met).

    Science.gov (United States)

    Gucev, Zoran; Slavevska, Nevenka; Tasic, Velibor; Laban, Nevenka; Pop-Jordanova, Nada; Danilovski, Dragan; Woolf, Jacqueline; Cole, Duncan

    2011-05-01

    Congenital erythropoietic porphyria (CEP) is an autosomal recessive inborn error of metabolism that results from the markedly deficient activity of uroporphyrinogen III synthase (UROS). We describe a 14-year-old girl with red urine since infancy, progressive blistering and scarring of the skin, and moderate hemolytic anemia. After years of skin damage, her face is mutilated; she has a bald patch on the scalp, hypertrichosis of the neck, areas of skin darkening, and limited joint movements of the hands. Total urine excretion and fecal total porphyrin were both markedly raised above normal levels. Sequencing of the UROS gene identified two mutations causing CEP (Cys73Arg, Thr228Met). The patient lesions are progressing. Bone marrow transplantation and/or gene therapy are proposed as the next steps in her treatment. In brief, we describe a CEP with confirmed two pathogenic mutations, severe phenotype and discuss the various treatment options available.

  4. Congenital erythropoietic porphyria with two mutations of the uroporphyrinogen III synthase gene (Cys73Arg, Thr228Met

    Directory of Open Access Journals (Sweden)

    Zoran Gucev

    2011-01-01

    Full Text Available Congenital erythropoietic porphyria (CEP is an autosomal recessive inborn error of metabolism that results from the markedly deficient activity of uroporphyrinogen III synthase (UROS. We describe a 14-year-old girl with red urine since infancy, progressive blistering and scarring of the skin, and moderate hemolytic anemia. After years of skin damage, her face is mutilated; she has a bald patch on the scalp, hypertrichosis of the neck, areas of skin darkening, and limited joint movements of the hands. Total urine excretion and fecal total porphyrin were both markedly raised above normal levels. Sequencing of the UROS gene identified two mutations causing CEP (Cys73Arg, Thr228Met. The patient lesions are progressing. Bone marrow transplantation and/or gene therapy are proposed as the next steps in her treatment. In brief, we describe a CEP with confirmed two pathogenic mutations, severe phenotype and discuss the various treatment options available.

  5. Targeting ESR1-Mutant Breast Cancer

    Science.gov (United States)

    2015-09-01

    cell line, biochemical and structural biology techniques to uncover the best candidate drugs for the clinical targeting of these mutants. Targeting...ESR1-­‐Mutant  Breast  Cancer   W81XWH-­‐14-­‐1-­‐0360   4   2. Keywords Estrogen Receptor Acquired Drug Resistance Metastatic Breast...preparations for publication: 1) “ESR1 Somatic Mutations Y537S and D538G Confer Breast Cancer Endocrine Resistance by Stabilizing the Active AF-2 conformation

  6. Regiospecificity determinants of human heme oxygenase: differential NADPH- and ascorbate-dependent heme cleavage by the R183E mutant.

    Science.gov (United States)

    Wang, Jinling; Lad, Latesh; Poulos, Thomas L; Ortiz de Montellano, Paul R

    2005-01-28

    The ability of the human heme oxygenase-1 (hHO-1) R183E mutant to oxidize heme in reactions supported by either NADPH-cytochrome P450 reductase or ascorbic acid has been compared. The NADPH-dependent reaction, like that of wild-type hHO-1, yields exclusively biliverdin IXalpha. In contrast, the R183E mutant with ascorbic acid as the reductant produces biliverdin IXalpha (79 +/- 4%), IXdelta (19 +/- 3%), and a trace of IXbeta. In the presence of superoxide dismutase and catalase, the yield of biliverdin IXdelta is decreased to 8 +/- 1% with a corresponding increase in biliverdin IXalpha. Spectroscopic analysis of the NADPH-dependent reaction shows that the R183E ferric biliverdin complex accumulates, because reduction of the iron, which is required for sequential iron and biliverdin release, is impaired. Reversal of the charge at position 183 makes reduction of the iron more difficult. The crystal structure of the R183E mutant, determined in the ferric and ferrous-NO bound forms, shows that the heme primarily adopts the same orientation as in wild-type hHO-1. The structure of the Fe(II).NO complex suggests that an altered active site hydrogen bonding network supports catalysis in the R183E mutant. Furthermore, Arg-183 contributes to the regiospecificity of the wild-type enzyme, but its contribution is not critical. The results indicate that the ascorbate-dependent reaction is subject to a lower degree of regiochemical control than the NADPH-dependent reaction. Ascorbate may be able to reduce the R183E ferric and ferrous dioxygen complexes in active site conformations that cannot be reduced by NADPH-cytochrome P450 reductase.

  7. Ophthalmic manifestations in a Chinese family with familial amyloid polyneuropathy due to a TTR Gly83Arg mutation.

    Science.gov (United States)

    Liu, T; Zhang, B; Jin, X; Wang, W; Lee, J; Li, J; Yuan, H; Cheng, X

    2014-01-01

    To describe the characteristic ophthalmic phenotypes of a large Chinese family with familial amyloid polyneuropathy due to a missense mutation in transthyretin (TTR) (c.307 C>G). Twenty-seven individuals (12 affected, 15 unaffected) from a five-generation Chinese family underwent general medical examination and comprehensive ophthalmic examination, including best correct visual acuity, intraocular pressure measurements, Schirmer test, slitlamp examination, fundoscopy, and ocular ultrasonography. Histological examination of vitreous biopsies using Congo red staining and immunohistochemistry was performed. Cardiovascular magnetic resonance (CMR), electrocardiogram, and echocardiogram were used to evaluate cardiac amyloidosis. Electromyography was used to evaluate nerve function. All four exons of TTR were amplified by PCR, sequenced using a Bigdye terminator v3.1 cycle sequencing kit and analyzed on an ABI 3700XL Genetic Analyzer. All 12 affected individuals in the family had ocular manifestations, including severe vitreous opacities, secondary glaucoma, xerophthalmia, dyscoria, and attenuated retinal arteries. Congo red staining demonstrated amyloid deposits in the vitreous, and immunohistochemical staining confirmed the deposition of TTR proteins in the vitreous. Twelve individuals had polyneuropathy, and electromyography detected functional damage in peripheral nerves. One individual was diagnosed with cardiac amyloidosis by CMR. Direct sequencing revealed the heterozygous missense mutation in TTR (c.307 C>G p.Gly83Arg) in all 12 affected individuals. The mutation co-segregated with the disease phenotype and was absent in 100 normal controls. Vitreous opacity is very common in patients with the TTR Gly83Arg mutation; other clinical characteristics associated with the mutation include polyneuropathy and cardiac amyloidosis.

  8. Frequencies of the Arg16Gly, Gln27Glu and Thr164Ile Adrenoceptor β2 Polymorphisms among Omanis

    Directory of Open Access Journals (Sweden)

    Khalid Al-Balushi

    2015-11-01

    Full Text Available Objectives: This study aimed to assess the distribution of missense mutations in the adrenoceptor β2 (ADRB2 gene in an Omani cohort. Methods: This study was carried out between May 2014 and March 2015 at the Sultan Qaboos University, Muscat, Oman. Blood samples were taken from 316 unrelated Omani subjects. Genotyping for rs1042713 (c.46A>G, p.Arg16Gly, rs1042714 (c.79C>G, p.Gln27Glu and rs1800888 (c.491C>T, p.Thr164Ile polymorphisms was performed by real-time polymerase chain reaction using single nucleotide polymorphism (SNP genotyping assays. The allelic frequencies of these polymorphisms were estimated on the basis of the observed numbers of specific alleles from the genotype data for male and female subjects. The genotype frequencies for each polymorphism were tested for deviation from the Hardy-Weinberg equilibrium. Results: Gly16 and Glu27 were the most frequent variants found among the cohort (63% and 75%, respectively. The Ile164 variant was not detected in the study population. There was a significant linkage disequilibrium between the rs1042713 and rs1042714 SNPs (r2 = 0.209; P ≤0.001. The most observed haplotypes were Gly16-Gln27 and Arg16-Gln27 (0.37 and 0.38, respectively. The frequency of Gly16-Glu27 was 0.25, comprising all Glu27 carriers. Conclusion: The allelic distribution of variants in this Omani cohort was similar to distributions reported among Caucasian populations.

  9. Rapid Antibiotic Resistance Evolution of GASP Mutants

    Science.gov (United States)

    Zhang, Qiucen; Kim, Hyunsung; Pourmand, Nader; Austin, Robert

    2012-02-01

    The GASP phenotype in bacteria is due to a mutation which enables the bacteria to grow under high stress conditions where other bacteria stop growing. We probe using our Death Galaxy microenvironment how rapidly the GASP mutant can evolve resistance to mutagenic antibiotics compared to wild-type bacteria, and explore the genomic landscape changes due to the evolution of resistance.

  10. Generation and characterization of pigment mutants of ...

    African Journals Online (AJOL)

    acer

    deleterious effects on living organisms (Shigaeva et al.,. 1994); they are also ... It was cultured at 25±0.5°C with a fluorescent light intensity of approximately 6 ... mutants) resulted in three new colonies characterized by different green colors ...

  11. A dominant semi dwarf mutant in rice

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    @@ In the winter of 1997, a semi dwarf mutant was found in the F6 population of M9056/ R8018 xuan in Hainan Province. In the spring of 1998, the seeds were sown in Hefei, Anhui Province and the plant height of the population was measured at maturity.

  12. Nicotinamide ribosyl uptake mutants in Haemophilus influenzae.

    Science.gov (United States)

    Herbert, Mark; Sauer, Elizabeta; Smethurst, Graeme; Kraiss, Anita; Hilpert, Anna-Karina; Reidl, Joachim

    2003-09-01

    The gene for the nicotinamide riboside (NR) transporter (pnuC) was identified in Haemophilus influenzae. A pnuC mutant had only residual NR uptake and could survive in vitro with high concentrations of NR, but could not survive in vivo. PnuC may represent a target for the development of inhibitors for preventing H. influenzae disease.

  13. Mutant PTEN in Cancer : Worse Than Nothing

    NARCIS (Netherlands)

    Leslie, Nick R; den Hertog, Jeroen

    2014-01-01

    Tumor suppressors block the development of cancer and are often lost during tumor development. Papa et al. show that partial loss of normal PTEN tumor suppressor function can be compounded by additional disruption caused by the expression of inactive mutant PTEN protein. This has significant

  14. Colored HOMFLY polynomials can distinguish mutant knots

    CERN Document Server

    Nawata, Satoshi; Singh, Vivek Kumar

    2015-01-01

    We illustrate from the viewpoint of braiding operations on WZNW conformal blocks how colored HOMFLY polynomials with multiplicity structure can detect mutations. As an example, we explicitly evaluate the (2,1)-colored HOMFLY polynomials that distinguish a famous mutant pair, Kinoshita-Terasaka and Conway knot.

  15. Replication and pathogenicity of primer binding site mutants of SL3-3 murine leukemia viruses

    DEFF Research Database (Denmark)

    Lund, Anders Henrik; Schmidt, J; Luz, A

    1999-01-01

    delayed relative to that of the wild-type virus, molecular tumor analysis indicated that all the primer binding site-modified viruses induce T-cell lymphomas similar to those induced by the wild-type virus in terms of frequencies of genomic rearrangements within the T-cell receptor beta......) in undifferentiated embryonic cells. In this study we test whether SL3-3 MLV can replicate stably using tRNA primers other than the cognate tRNAPro and analyze the effect of altering the primer binding site sequence to match the 3' end of tRNA1Gln, tRNA3Lys, or tRNA1,2Arg in a mouse pathogenicity model. Contrary...... to findings from cell culture studies of primer binding site-modified human immunodeficiency virus type 1 and avian retroviruses, our findings were that SL3-3 MLV may stably and efficiently replicate with tRNA primers other than tRNAPro. Although lymphoma induction of the SL3-3 Lys3 mutant was significantly...

  16. Effects of temperature on the p53-DNA binding interactions and their dynamical behavior: comparing the wild type to the R248Q mutant.

    Directory of Open Access Journals (Sweden)

    Khaled Barakat

    Full Text Available BACKGROUND: The protein p53 plays an active role in the regulation of cell cycle. In about half of human cancers, the protein is inactivated by mutations located primarily in its DNA-binding domain. Interestingly, a number of these mutations possess temperature-induced DNA-binding characteristics. A striking example is the mutation of Arg248 into glutamine or tryptophan. These mutants are defective for binding to DNA at 310 K although they have been shown to bind specifically to several p53 response elements at sub-physiological temperatures (298-306 K. METHODOLOGY/PRINCIPAL FINDINGS: This important experimental finding motivated us to examine the effects of temperature on the structure and configuration of R248Q mutant and compare it to the wild type protein. Our aim is to determine how and where structural changes of mutant variants take place due to temperature changes. To answer these questions, we compared the mutant to the wild-type proteins from two different aspects. First, we investigated the systems at the atomistic level through their DNA-binding affinity, hydrogen bond networks and spatial distribution of water molecules. Next, we assessed changes in their long-lived conformational motions at the coarse-grained level through the collective dynamics of their side-chain and backbone atoms separately. CONCLUSIONS: The experimentally observed effect of temperature on the DNA-binding properties of p53 is reproduced. Analysis of atomistic and coarse-grained data reveal that changes in binding are determined by a few key residues and provide a rationale for the mutant-loss of binding at physiological temperatures. The findings can potentially enable a rescue strategy for the mutant structure.

  17. GAMPMS: Genetic algorithm managed peptide mutant screening.

    Science.gov (United States)

    Long, Thomas; McDougal, Owen M; Andersen, Tim

    2015-06-30

    The prominence of endogenous peptide ligands targeted to receptors makes peptides with the desired binding activity good molecular scaffolds for drug development. Minor modifications to a peptide's primary sequence can significantly alter its binding properties with a receptor, and screening collections of peptide mutants is a useful technique for probing the receptor-ligand binding domain. Unfortunately, the combinatorial growth of such collections can limit the number of mutations which can be explored using structure-based molecular docking techniques. Genetic algorithm managed peptide mutant screening (GAMPMS) uses a genetic algorithm to conduct a heuristic search of the peptide's mutation space for peptides with optimal binding activity, significantly reducing the computational requirements of the virtual screening. The GAMPMS procedure was implemented and used to explore the binding domain of the nicotinic acetylcholine receptor (nAChR) α3β2-isoform with a library of 64,000 α-conotoxin (α-CTx) MII peptide mutants. To assess GAMPMS's performance, it was compared with a virtual screening procedure that used AutoDock to predict the binding affinity of each of the α-CTx MII peptide mutants with the α3β2-nAChR. The GAMPMS implementation performed AutoDock simulations for as few as 1140 of the 64,000 α-CTx MII peptide mutants and could consistently identify a set of 10 peptides with an aggregated binding energy that was at least 98% of the aggregated binding energy of the 10 top peptides from the exhaustive AutoDock screening.

  18. Phanerochaete mutants with enhanced ligninolytic activity

    Energy Technology Data Exchange (ETDEWEB)

    Kakar, S.N.; Perez, A.; Gonzales, J. (Argonne National Lab., IL (United States))

    In addition to lignin, the white rot fungus Phanerochaete chrysosporium has the ability to degrade a wide spectrum of recalcitrant organopollutants in soils and aqueous media. Most of the organic compounds are degraded under ligninolytic conditions with the involvement of the extracellular enzymes, lignin peroxidases, and manganese-dependent peroxidases, which are produced as secondary metabolites triggered by conditions of nutrient starvation (e.g., nitrogen limitation). The fungus and its enzymes can thus provide alternative technologies for bioremediation, biopulping, biobleaching, and other industrial applications. The efficiency and effectiveness of the fungus can be enhanced by increasing production and secretion of the important enzymes in large quantities and as primary metabolites under enriched conditions. One way this can be achieved is through isolation of mutants that are deregulated, or are hyperproducers or supersectors of key enzymes under enriched conditions. Through UV-light and [gamma]-ray mutagenesis, the authors have isolated a variety of mutants, some of which produce key enzymes of the ligninolytic system under high-nitrogen growth conditions. One of the mutants, 76UV, produced 272 U of lignin peroxidases enzyme activity/L after 9 d under high nitrogen (although the parent strain does not produce this enzyme under these conditions). The mutant and the parent strains produced up to 54 and 62 U/L, respectively, of the enzyme activity under low-nitrogen growth conditions during this period. In some experiments, the mutant showed 281 U/L of enzyme activity under high nitrogen after 17 d. 17 refs., 1 fig., 3 tabs.

  19. MASS LOSS AND NITROGEN DYNAMICS DURING THE DECOMPOSITION OF A N-LABELED N2-FIXING EPOPHYTIC LICHEN, LOBARIA OREGANA (TUCK.) MULL. ARG.

    Science.gov (United States)

    We studied mass loss and nitrogen dynamics during fall and spring initiated decomposition of an N2-fixing epiphytic lichen, Lobaria oregana (Tuck.) Mull. Arg. using 15N. We developed a method of labeling lichens with 15N that involved spraying lichen material with a nutrient sol...

  20. Cell adhesion to fibrillin-1: identification of an Arg-Gly-Asp-dependent synergy region and a heparin-binding site that regulates focal adhesion formation

    DEFF Research Database (Denmark)

    Bax, Daniel V; Mahalingam, Yashithra; Cain, Stuart;

    2007-01-01

    We have defined the molecular basis of cell adhesion to fibrillin-1, the major structural component of extracellular microfibrils that are associated with elastic fibres. Using human dermal fibroblasts, and recombinant domain swap fragments containing the Arg-Gly-Asp motif, we have demonstrated a...

  1. Primary structure of the precursor for the sea anemone neuropeptide Antho-RFamide (less than Glu-Gly-Arg-Phe-NH2)

    DEFF Research Database (Denmark)

    Darmer, D; Schmutzler, C; Diekhoff, D;

    1991-01-01

    Neuropeptides containing the carboxylterminal sequence Arg-Phe-NH2 are found throughout the animal kingdom and are important substances mediating neuronal communication. Here, we have cloned the cDNA coding for the precursor protein of the sea anemone neuropeptide (Antho-RFamide) less than Glu...

  2. XRCC1 Arg399Gln polymorphism confers risk of breast cancer in American population: a meta-analysis of 10846 cases and 11723 controls.

    Directory of Open Access Journals (Sweden)

    Tao Bu

    Full Text Available BACKGROUND: In the X-ray repair cross-complementing group 1 (XRCC1 gene, a polymorphism, Arg399Gln (rs25487, has been shown to change neoconservative amino acid and thus result in alternation of DNA repair capacity. Numerous studies have investigated the association between Arg399Gln and breast cancer risk in the American population, but yielding inconsistent results. This study aimed to clarify the role of this polymorphism in susceptibility to breast cancer. METHODS: Literatures were searched in multiple databases including PubMed, Springer Link, Ovid, EBSCO and ScienceDirect databases up to April 2013. A comprehensive meta-analysis was conducted to estimate the overall odds ratio (OR, by integrating data from 18 case control studies of 10846 cases and 11723 controls in the American population. RESULTS: Overall, significant association was observed between the Arg399Gln polymorphism and breast cancer risk under the random-effects model (OR for dominant model = 1.12, 95% CI: 1.02-1.24, P heterogeneity = 0.003; OR for additive model = 1.07, 95% CI: 1.01-1.14, P heterogeneity = 0.017. Further sensitivity analysis supported the robust stability of this current result by showing similar ORs before and after removal of a single study. CONCLUSIONS: This meta-analysis suggests that the XRCC1 Arg399Gln polymorphism may significantly contribute to susceptibility of breast cancer in the American population.

  3. XRCC1 Arg399Gln Polymorphism Confers Risk of Breast Cancer in American Population: A Meta-Analysis of 10846 Cases and 11723 Controls

    Science.gov (United States)

    Sun, Yong; Zhao, Li; Peng, Yang; Zhou, Shudong; Li, Lixia; Chen, Sidong; Gao, Yanhui

    2014-01-01

    Background In the X-ray repair cross-complementing group 1 (XRCC1) gene, a polymorphism, Arg399Gln (rs25487), has been shown to change neoconservative amino acid and thus result in alternation of DNA repair capacity. Numerous studies have investigated the association between Arg399Gln and breast cancer risk in the American population, but yielding inconsistent results. This study aimed to clarify the role of this polymorphism in susceptibility to breast cancer. Methods Literatures were searched in multiple databases including PubMed, Springer Link, Ovid, EBSCO and ScienceDirect databases up to April 2013. A comprehensive meta-analysis was conducted to estimate the overall odds ratio (OR), by integrating data from 18 case control studies of 10846 cases and 11723 controls in the American population. Results Overall, significant association was observed between the Arg399Gln polymorphism and breast cancer risk under the random-effects model (OR for dominant model = 1.12, 95% CI: 1.02–1.24, Pheterogeneity = 0.003; OR for additive model = 1.07, 95% CI: 1.01–1.14, Pheterogeneity = 0.017). Further sensitivity analysis supported the robust stability of this current result by showing similar ORs before and after removal of a single study. Conclusions This meta-analysis suggests that the XRCC1 Arg399Gln polymorphism may significantly contribute to susceptibility of breast cancer in the American population. PMID:24489692

  4. High Incidence of Noonan Syndrome Features Including Short Stature and Pulmonic Stenosis in Patients carrying NF1 Missense Mutations Affecting p.Arg1809: Genotype-Phenotype Correlation.

    Science.gov (United States)

    Rojnueangnit, Kitiwan; Xie, Jing; Gomes, Alicia; Sharp, Angela; Callens, Tom; Chen, Yunjia; Liu, Ying; Cochran, Meagan; Abbott, Mary-Alice; Atkin, Joan; Babovic-Vuksanovic, Dusica; Barnett, Christopher P; Crenshaw, Melissa; Bartholomew, Dennis W; Basel, Lina; Bellus, Gary; Ben-Shachar, Shay; Bialer, Martin G; Bick, David; Blumberg, Bruce; Cortes, Fanny; David, Karen L; Destree, Anne; Duat-Rodriguez, Anna; Earl, Dawn; Escobar, Luis; Eswara, Marthanda; Ezquieta, Begona; Frayling, Ian M; Frydman, Moshe; Gardner, Kathy; Gripp, Karen W; Hernández-Chico, Concepcion; Heyrman, Kurt; Ibrahim, Jennifer; Janssens, Sandra; Keena, Beth A; Llano-Rivas, Isabel; Leppig, Kathy; McDonald, Marie; Misra, Vinod K; Mulbury, Jennifer; Narayanan, Vinodh; Orenstein, Naama; Galvin-Parton, Patricia; Pedro, Helio; Pivnick, Eniko K; Powell, Cynthia M; Randolph, Linda; Raskin, Salmo; Rosell, Jordi; Rubin, Karol; Seashore, Margretta; Schaaf, Christian P; Scheuerle, Angela; Schultz, Meredith; Schorry, Elizabeth; Schnur, Rhonda; Siqveland, Elizabeth; Tkachuk, Amanda; Tonsgard, James; Upadhyaya, Meena; Verma, Ishwar C; Wallace, Stephanie; Williams, Charles; Zackai, Elaine; Zonana, Jonathan; Lazaro, Conxi; Claes, Kathleen; Korf, Bruce; Martin, Yolanda; Legius, Eric; Messiaen, Ludwine

    2015-11-01

    Neurofibromatosis type 1 (NF1) is one of the most frequent genetic disorders, affecting 1:3,000 worldwide. Identification of genotype-phenotype correlations is challenging because of the wide range clinical variability, the progressive nature of the disorder, and extreme diversity of the mutational spectrum. We report 136 individuals with a distinct phenotype carrying one of five different NF1 missense mutations affecting p.Arg1809. Patients presented with multiple café-au-lait macules (CALM) with or without freckling and Lisch nodules, but no externally visible plexiform neurofibromas or clear cutaneous neurofibromas were found. About 25% of the individuals had Noonan-like features. Pulmonic stenosis and short stature were significantly more prevalent compared with classic cohorts (P NF1-patient showed two different somatic NF1 mutations, p.Arg1809Cys and a multi-exon deletion, providing genetic evidence that p.Arg1809Cys is a loss-of-function mutation in the melanocytes and causes a pigmentary phenotype. Constitutional missense mutations at p.Arg1809 affect 1.23% of unrelated NF1 probands in the UAB cohort, therefore this specific NF1 genotype-phenotype correlation will affect counseling and management of a significant number of patients.

  5. Site-directed mutagenesis of Arg58 and Asp86 of elongation factor Tu from Escherichia coli: effects on the GTPase reaction and aminoacyl-tRNA binding

    DEFF Research Database (Denmark)

    Knudsen, Charlotte Rohde; Clark, Brian F. C.

    1996-01-01

    Elongation factor Tu from Escherichia coli was mutated separately at positions Asp86 and Arg58, in order to shed light both on the GTPase mechanism of elongation factor Tu and on the binding of aminoacyl-tRNA. In addition, the binding of guanine nucleotides was investigated by determination...

  6. The effects of mutating Tyr9 and Arg15 on the structure, stability, conformational dynamics and mechanism of GSTA3-3

    CSIR Research Space (South Africa)

    Robertson, GJ

    2017-03-01

    Full Text Available GSTA1-1, its closest competitor in the Alpha class of GSTs. In order to examine the differences between Alpha class GSTs and to better elucidate the mechanism of GSTA3-3 the roles of Tyr9 and Arg15 were examined. Tyr9 is the major catalytic residue...

  7. Assessment of the Link between XRCC1 Arg399Gln Polymorphism and Breast Cancer: a Meta-Analysis in a Single Ethnic Group.

    Science.gov (United States)

    Chen, Xin-Ping; Wen, Hai-Feng; Zhang, Fan; Xu, Wei-Hua; Ma, Zhi-Chao; Fu, Sheng-Miao

    2017-04-01

    Although various individual studies have been conducted to determine the association between X-ray repair cross-complementing group 1 (XRCC1) Arg399Gln polymorphism and breast cancer, the results remain inconclusive. To assess the influence of XRCC1 Arg399Gln polymorphism on the risk of breast cancer, a metaanalysis was performed in a single ethnic group. Eligible studies were identified via databases such as PubMed, Springer Link, Ovid, Chinese Wanfang Data Knowledge Service Platform, Chinese National Knowledge Infrastructure, and Chinese Biology Medicine, throughout February 2016. Pooled odds ratios (ORs) and 95% confidence intervals (CIs) were used to assess the strengths of the associations. Ten studies documenting a total of 4732 breast cancer cases and 5677 controls were included in this metaanalysis. The results indicated no significant association between XRCC1 Arg399Gln polymorphism and breast cancer risk in both total analysis and subgroup analysis stratified by geographical areas and source of controls. This meta-analysis provided evidence that XRCC1 Arg399Gln variant might not be risk alleles for breast cancer susceptibility in the Chinese population. Further studies conducted in other ethnic groups are required for definite conclusions.

  8. IL-10 and ARG-1 Concentrations in Bone Marrow and Peripheral Blood of Metastatic Neuroblastoma Patients Do Not Associate with Clinical Outcome

    Directory of Open Access Journals (Sweden)

    Fabio Morandi

    2015-01-01

    Full Text Available The expression of the immunosuppressive molecules IL-10 and arginase 1 (ARG-1, and of FOXP3 and CD163, as markers of regulatory T cells (Treg and macrophages, respectively, was evaluated in bone marrow (BM and peripheral blood (PB samples collected at diagnosis from patients with metastatic neuroblastoma (NB. IL-10 and ARG-1 plasma concentrations were measured and the association of each parameter with patients’ outcome was tested. The percentages of immunosuppressive Treg and type-1 regulatory (Tr1 cells were also determined. In both BM and PB samples, IL-10 mRNA expression was higher in metastatic NB patients than in controls. IL-10 plasma concentration was higher in patients with NB regardless of stage. Neither IL-10 expression nor IL-10 plasma concentration significantly associated with patient survival. In PB samples from metastatic NB patients, ARG-1 and CD163 expression was higher than in controls but their expression did not associate with survival. Moreover, ARG-1 plasma concentration was lower than in controls, and no association with patient outcome was found. Finally, in metastatic NB patients, the percentage of circulating Treg was higher than in controls, whereas that of Tr1 cells was lower. In conclusion, although IL-10 concentration and Treg percentage were increased, their contribution to the natural history of metastatic NB appears uncertain.

  9. Nociceptive stimulation induces expression of Arc/Arg3.1 in the spinal cord with a preference for neurons containing enkephalin

    NARCIS (Netherlands)

    S.M. Hossaini (Mehdi); J.L.M. Jongen (Joost); K. Biesheuvel (Karla); D. Kuhl (Dietmar); J.C. Holstege (Jan)

    2010-01-01

    textabstractBackground: In pain processing, long term synaptic changes play an important role, especially during chronic pain. The immediate early gene Arc/Arg3.1 has been widely implicated in mediating long-term plasticity in telencephalic regions, such as the hippocampus and cortex. Accordingly, A

  10. Site-directed mutagenesis of Arg58 and Asp86 of elongation factor Tu from Escherichia coli: effects on the GTPase reaction and aminoacyl-tRNA binding

    DEFF Research Database (Denmark)

    Knudsen, Charlotte Rohde; Clark, Brian F. C.

    1996-01-01

    Elongation factor Tu from Escherichia coli was mutated separately at positions Asp86 and Arg58, in order to shed light both on the GTPase mechanism of elongation factor Tu and on the binding of aminoacyl-tRNA. In addition, the binding of guanine nucleotides was investigated by determination of th...

  11. Assessment of the Role of MAP Kinase in Mediating Activity-Dependent Transcriptional Activation of the Immediate Early Gene "Arc/Arg3.1" in the Dentate Gyrus in Vivo

    Science.gov (United States)

    Chotiner, Jennifer K.; Nielson, Jessica; Farris, Shannon; Lewandowski, Gail; Huang, Fen; Banos, Karla; de Leon, Ray; Steward, Oswald

    2010-01-01

    Different physiological and behavioral events activate transcription of "Arc/Arg3.1" in neurons in vivo, but the signal transduction pathways that mediate induction in particular situations remain to be defined. Here, we explore the relationships between induction of "Arc/Arg3.1" transcription in dentate granule cells in vivo and activation of…

  12. Assessment of the Role of MAP Kinase in Mediating Activity-Dependent Transcriptional Activation of the Immediate Early Gene "Arc/Arg3.1" in the Dentate Gyrus in Vivo

    Science.gov (United States)

    Chotiner, Jennifer K.; Nielson, Jessica; Farris, Shannon; Lewandowski, Gail; Huang, Fen; Banos, Karla; de Leon, Ray; Steward, Oswald

    2010-01-01

    Different physiological and behavioral events activate transcription of "Arc/Arg3.1" in neurons in vivo, but the signal transduction pathways that mediate induction in particular situations remain to be defined. Here, we explore the relationships between induction of "Arc/Arg3.1" transcription in dentate granule cells in vivo and activation of…

  13. Development and dissection of diagnostic SNP markers for the downy mildew resistance genes PlArg and Pl8 and maker-assisted gene pyramiding in sunflower (Helianthus annuus L.)

    Science.gov (United States)

    Downy mildew, which is caused by fungus Plasmopara halstedii (Farl.) Berlese & de Toni, is one of the most important diseases that affect sunflower production globally. Two downy mildew resistance genes, PlArg and Pl8, were discovered in the late 1980s. Over two decades, PlArg is still effective aga...

  14. Structural Characterization of Heme Environmental Mutants of CgHmuT that Shuttles Heme Molecules to Heme Transporters

    Directory of Open Access Journals (Sweden)

    Norifumi Muraki

    2016-05-01

    Full Text Available Corynebacteria contain a heme uptake system encoded in hmuTUV genes, in which HmuT protein acts as a heme binding protein to transport heme to the cognate transporter HmuUV. The crystal structure of HmuT from Corynebacterium glutamicum (CgHmuT reveals that heme is accommodated in the central cleft with His141 and Tyr240 as the axial ligands and that Tyr240 forms a hydrogen bond with Arg242. In this work, the crystal structures of H141A, Y240A, and R242A mutants were determined to understand the role of these residues for the heme binding of CgHmuT. Overall and heme environmental structures of these mutants were similar to those of the wild type, suggesting that there is little conformational change in the heme-binding cleft during heme transport reaction with binding and the dissociation of heme. A loss of one axial ligand or the hydrogen bonding interaction with Tyr240 resulted in an increase in the redox potential of the heme for CgHmuT to be reduced by dithionite, though the wild type was not reduced under physiological conditions. These results suggest that the heme environmental structure stabilizes the ferric heme binding in CgHmuT, which will be responsible for efficient heme uptake under aerobic conditions where Corynebacteria grow.

  15. The XRCC1 Arg194Trp polymorphism is significantly associated with lung adenocarcinoma: a case-control study in an Eastern European Caucasian group

    Directory of Open Access Journals (Sweden)

    Cătană A

    2015-11-01

    Full Text Available Andreea Cătană,1 Monica Pop,2 Bianca Domokos Hincu,2 Ioan V Pop,1 Felicia M Petrişor,1 Mihai D Porojan,3 Radu A Popp1 1Department of Molecular Sciences, 2Department of Pneumology, 3Department of Internal Medicine, Iuliu Haţieganu University of Medicine and Pharmacy, Cluj-Napoca, Romania Abstract: DNA repair plays an important role in maintaining the integrity of the genome by repairing DNA damage induced by carcinogens. Certain genetic polymorphisms that occur in DNA-repair genes may affect the ability to repair DNA defects, and may represent a risk factor in carcinogenesis. The gene XRCC1 is involved in DNA repair. The purpose of our study was to investigate the association between XRCC1 Arg194Trp and Arg399Gln polymorphisms and the risk of lung cancer in a Romanian population. We recruited 222 healthy controls and 102 patients with lung cancer. Genotypes were determined by multiplex polymerase chain-reaction restriction fragment-length polymorphism. Statistical analysis (odds ratio, recessive model revealed an increased risk for lung cancer for the homozygous 194Trp genotype (χ2=0.186, odds ratio 10.667, 95% confidence interval 1.309–86.933; P=0.007. Also, we found an association between the 194Trp allele and women with lung adenocarcinoma. In conclusion, the results of the study place the XRCC1 Arg194Trp polymorphism among independent risk factors for developing lung cancer. Keywords: lung cancer, XRCC1 Arg194Trp, XRCC1 Arg399Gln

  16. 妊娠湖羊补充 L-Arg 和 Se 对新生羔羊血清参数的影响%Effects of L-Arg and Selenium Supplementation to Pregnant Hu Sheep on Serum Parameters of Newborn Lambs

    Institute of Scientific and Technical Information of China (English)

    王连群; 侯芳; 郭同军; 李静溪; 马春晖

    2015-01-01

    为外源性精氨酸(Arg)和硒(Se)在养羊生产上的应用提供参考,选取体重相近、同期发情处理的已孕湖羊20只,随机平均分为4组(饲喂相同的基础日粮),即 Se 组:于妊娠期50 d 至分娩在基础日粮中额外添加 Se 0.2 mg/kg;Arg 组:于妊娠期130 d 至分娩每天静脉注射 L-Arg 70 mg/kg 体重;AS 组:Se 组和 Arg 组联用;对照组(CK):饲喂基础日粮。各组羊只分娩后立即采集新生羔羊的血液进行测定分析,以探明外源性 Arg 和 Se 通过母体对新生羔羊血清参数的影响。结果表明:1)与对照组比较,AS 组血清超氧化物歧化酶活力显著增加(P <0.05),Se 组和 AS 组血清谷胱甘肽过氧化物酶活力极显著增加(P <0.01)。2)AS组血清精氨酸含量显著高于 Se 组(P <0.05),极显著高于对照组(P <0.01);Arg 组血清精氨酸含量显著高于对照组(P <0.05)。Arg 组、AS 组的血清鸟氨酸含量均显著高于 Se 组和对照组(P <0.05)。3)AS组、Se 组的血清 Se 含量极显著高于 Arg 组和对照组(P <0.01)。妊娠湖羊日粮添加 Se 或与静脉注射L-Arg 联用能够提高新生羔羊机体的抗氧化性,单独进行静脉注射 L-Arg 或与日粮添加 Se 联用能显著增加新生羔羊血清中的 Arg 和鸟氨酸含量,有利于新生羔羊的生长发育。%To provide a reference for application of exogenous Arg and Se in sheep husbandry,20 pregnant Hu Sheep ewes with similar weight and estrus synchronization were selected and allotted randomly into four groups (five in each group,fed the same basal diet),namely:Se group (supplemented with 0.2 mg/kg of Se added to the basal diet every day during the 50 days of pregnancy to childbirth),Arg group (intravenous injection of 70 mg/kg BW d of L-Arg every day during the 130 days of pregnancy to childbirth),the AS group (supplemented with 0.2 mg/kg of Se

  17. Amuvatinib has cytotoxic effects against NRAS-mutant melanoma but not BRAF-mutant melanoma.

    Science.gov (United States)

    Fedorenko, Inna V; Fang, Bin; Koomen, John M; Gibney, Geoffrey T; Smalley, Keiran S M

    2014-10-01

    Effective targeted therapy strategies are still lacking for the 15-20% of melanoma patients whose melanomas are driven by oncogenic NRAS. Here, we report on the NRAS-specific behavior of amuvatinib, a kinase inhibitor with activity against c-KIT, Axl, PDGFRα, and Rad51. An analysis of BRAF-mutant and NRAS-mutant melanoma cell lines showed the NRAS-mutant cohort to be enriched for targets of amuvatinib, including Axl, c-KIT, and the Axl ligand Gas6. Increasing concentrations of amuvatinib selectively inhibited the growth of NRAS-mutant, but not BRAF-mutant melanoma cell lines, an effect associated with induction of S-phase and G2/M-phase cell cycle arrest and induction of apoptosis. Mechanistically, amuvatinib was noted to either inhibit Axl, AKT, and MAPK signaling or Axl and AKT signaling and to induce a DNA damage response. In three-dimensional cell culture experiments, amuvatinib was cytotoxic against NRAS-mutant melanoma cell lines. Thus, we show for the first time that amuvatinib has proapoptotic activity against melanoma cell lines, with selectivity observed for those harboring oncogenic NRAS.

  18. Substitution of the Lys linker with the β-Ala linker dramatically decreased the renal uptake of 99mTc-labeled Arg-X-Asp-conjugated and X-Ala-Asp-conjugated α-melanocyte stimulating hormone peptides.

    Science.gov (United States)

    Flook, Adam M; Yang, Jianquan; Miao, Yubin

    2014-11-13

    The purpose of this study was to examine whether the substitution of the Lys linker with the β-Ala could reduce the renal uptake of (99m)Tc-labeled Arg-X-Asp-conjugated and X-Ala-Asp-conjugated α-melanocyte stimulating hormone (α-MSH) peptides. RSD-β-Ala-(Arg(11))CCMSH (1) {c[Arg-Ser-Asp-dTyr-Asp]-β-Ala-Cys-Cys-Glu-His-dPhe-Arg-Trp-Cys-Arg-Pro-Val-NH2}, RTD-β-Ala-(Arg(11))CCMSH (2), RVD-β-Ala-(Arg(11))CCMSH (3), RAD-β-Ala-(Arg(11))CCMSH (4), NAD-β-Ala-(Arg(11))CCMSH (5), and EAD-β-Ala-(Arg(11))CCMSH (6) peptides were synthesized and evaluated for their melanocortin 1 (MC1) receptor binding affinities in B16/F1 melanoma cells. The biodistribution of their (99m)Tc-conjugates were determined in B16/F1 melanoma-bearing C57 mice. The substitution of the Lys linker with β-Ala linker dramatically reduced the renal uptake of all six (99m)Tc-peptides. (99m)Tc-4 exhibited the highest melanoma uptake (15.66 ± 6.19% ID/g) and the lowest kidney uptake (20.18 ± 3.86% ID/g) among these (99m)Tc-peptides at 2 h postinjection. The B16/F1 melanoma lesions could be clearly visualized by single photon emission computed tomography (SPECT)/CT using (99m)Tc-4 as an imaging probe.

  19. Report on a 2009 mini-demonstration of the ARG-US Radio Frequency Identification (RFID) system in transportation.

    Energy Technology Data Exchange (ETDEWEB)

    Tsai, H.; Chen, K.; Jusko, M.; Craig, B.; Liu, Y.; Decision and Information Sciences

    2009-11-23

    The Packaging Certification Program (PCP) of the U.S. Department of Energy (DOE) Environmental Management (EM), Office of Packaging and Transportation (EM-14), has developed a radio frequency identification (RFID) tracking and monitoring system for the management of nuclear materials during storage and transportation. The system, developed by the PCP team at Argonne National Laboratory, consists of hardware (Mk-series sensor tags, fixed and handheld readers, form factor for multiple drum types, seal integrity sensors, and enhanced battery management), software (application programming interface, ARG-US software for local and remote/web applications, secure server and database management), and cellular/satellite communication interfaces for vehicle tracking and item monitoring during transport. The ability of the above system to provide accurate, real-time tracking and monitoring of the status of multiple, certified containers of nuclear materials has been successfully demonstrated in a week-long, 1,700-mile DEMO performed in April 2008. While the feedback from the approximately fifty (50) stakeholders who participated in and/or observed the DEMO progression were very positive and encouraging, two major areas of further improvements - system integration and web application enhancement - were identified in the post-DEMO evaluation. The principal purpose of the MiniDemo described in this report was to verify these two specific improvements. The MiniDemo was conducted on August 28, 2009. In terms of system integration, a hybrid communication interface - combining the RFID item-monitoring features and a commercial vehicle tracking system by Qualcomm - was developed and implemented. In the MiniDemo, the new integrated system worked well in reporting tag status and vehicle location accurately and promptly. There was no incompatibility of components. The robust commercial communication gear, as expected, helped improve system reliability. The MiniDemo confirmed that system

  20. Report on a 2009 mini-demonstration of the ARG-US Radio Frequency Identification (RFID) system in transportation.

    Energy Technology Data Exchange (ETDEWEB)

    Tsai, H.; Chen, K.; Jusko, M.; Craig, B.; Liu, Y.; Decision and Information Sciences

    2009-11-23

    The Packaging Certification Program (PCP) of the U.S. Department of Energy (DOE) Environmental Management (EM), Office of Packaging and Transportation (EM-14), has developed a radio frequency identification (RFID) tracking and monitoring system for the management of nuclear materials during storage and transportation. The system, developed by the PCP team at Argonne National Laboratory, consists of hardware (Mk-series sensor tags, fixed and handheld readers, form factor for multiple drum types, seal integrity sensors, and enhanced battery management), software (application programming interface, ARG-US software for local and remote/web applications, secure server and database management), and cellular/satellite communication interfaces for vehicle tracking and item monitoring during transport. The ability of the above system to provide accurate, real-time tracking and monitoring of the status of multiple, certified containers of nuclear materials has been successfully demonstrated in a week-long, 1,700-mile DEMO performed in April 2008. While the feedback from the approximately fifty (50) stakeholders who participated in and/or observed the DEMO progression were very positive and encouraging, two major areas of further improvements - system integration and web application enhancement - were identified in the post-DEMO evaluation. The principal purpose of the MiniDemo described in this report was to verify these two specific improvements. The MiniDemo was conducted on August 28, 2009. In terms of system integration, a hybrid communication interface - combining the RFID item-monitoring features and a commercial vehicle tracking system by Qualcomm - was developed and implemented. In the MiniDemo, the new integrated system worked well in reporting tag status and vehicle location accurately and promptly. There was no incompatibility of components. The robust commercial communication gear, as expected, helped improve system reliability. The MiniDemo confirmed that system

  1. Characterization of a Legionella micdadei mip mutant

    DEFF Research Database (Denmark)

    O'Connell, W A; Bangsborg, Jette Marie; Cianciotto, N P

    1995-01-01

    The pathogenesis of Legionella micdadei is dependent upon its ability to infect alveolar phagocytes. To better understand the basis of intracellular infection by this organism, we examined the importance of its Mip surface protein. In Legionella pneumophila, Mip promotes infection of both human m...... into the phagocyte. Similarly, the mutant was less able to parasitize Hartmannella amoebae. Taken together, these data argue that Mip specifically potentiates intracellular growth by L. micdadei....

  2. Some Mutant Forms of Quantum Mechanics

    CERN Document Server

    Takeuchi, Tatsu; Lewis, Zachary; Minic, Djordje

    2013-01-01

    We construct a `mutant' form of quantum mechanics on a vector space over the finite Galois field GF(q). We find that the correlations in our model do not violate the Clauser-Horne-Shimony-Holt (CHSH) version of Bell's inequality, despite the fact that the predictions of this discretized quantum mechanics cannot be reproduced with any hidden variable theory. An alternative `mutation' is also suggested.

  3. Spontaneous Nif- mutants of Rhodopseudomonas capsulata.

    OpenAIRE

    Wall, J D; Love, J.; Quinn, S P

    1984-01-01

    Revertible, spontaneous Nif- mutants of Rhodopseudomonas capsulata have been shown to accumulate in cultures growing photosynthetically with an amino acid as the nitrogen source such that H2 is maximally produced. The majority of such strains carry mutations which are clustered in a short region of the chromosome, probably representing one or two genes. Because this cluster includes temperature-sensitive mutations, it is also likely that it identifies the structural gene of a polypeptide. The...

  4. Mutant chaperonin proteins: new tools for nanotechnology

    Energy Technology Data Exchange (ETDEWEB)

    Li, Y [SETI Institute, 515 N Whisman Road, Mountain View, CA 94043 (United States); Paavola, C D [NASA Ames Research Center, Bioengineering Branch, Mail Stop 239-15, Moffett Field, CA 94035 (United States); Kagawa, H [SETI Institute, 515 N Whisman Road, Mountain View, CA 94043 (United States); Chan, S L [SETI Institute, 515 N Whisman Road, Mountain View, CA 94043 (United States); Trent, J D [NASA Ames Research Center, Bioengineering Branch, Mail Stop 239-15, Moffett Field, CA 94035 (United States)

    2007-11-14

    Much effort has gone into finding peptides that bind potentially useful nanoparticles, but relatively little effort has focused on the scaffolds that organize these peptides into useful nanostructures. Chaperonins are protein complexes with 14-18 protein subunits that self-assemble into double-ring complexes and function as scaffolds for peptides or amino acids that bind metallic and semiconductor quantum dots. The utility of chaperonins as scaffolds depends on their structure and their ability to self-assemble into double-rings and higher-order structures, such as filaments and two-dimensional arrays. To better understand the structure of chaperonins, we constructed a model of a group II chaperonin and, based on this model, genetically constructed five mutant subunits with significant deletions. We expressed these mutants as recombinant proteins and observed by native polyacrylamide gel electrophoresis (PAGE) and transmission electron microscopy (TEM) that they all self-assembled into double rings. Our model predicted and TEM confirmed that these deletions did not significantly change the 17 nm diameter of the wild-type double rings, but decreased their height and opened their central cavities. Four of the five mutants formed higher-order structures: chains of rings, bundles of chains or filaments, and two-dimensional arrays, which we suggest can be useful nanostructures.

  5. Isolation of a novel mutant from Bacillus subtilis natto.

    Science.gov (United States)

    Yoshida, Kazuo

    2006-01-01

    For the construction of strains with full probiotics function in intestines, deoxycholate resistant mutants were isolated from Bacillus subtilis natto. The partial characterization of the mutants was carried out and described.

  6. Novel homozygous mutation, c.400C>T (p.Arg134*), in the PVRL1 gene underlies cleft lip/palate-ectodermal dysplasia syndrome in an Asian patient.

    Science.gov (United States)

    Yoshida, Kazue; Hayashi, Ryota; Fujita, Hideki; Kubota, Masaya; Kondo, Mai; Shimomura, Yutaka; Niizeki, Hironori

    2015-07-01

    Cleft lip/palate-ectodermal dysplasia syndrome is a rare, autosomal recessive disorder caused by homozygous loss-of-function mutations of the poliovirus receptor-like 1 (PVRL1) gene encoding nectin-1. Nectin-1 is a cell-cell adhesion molecule that is important for the initial step in the formation of adherens junctions and tight junctions; it is expressed in keratinocytes, neurons, and the developing face and palate. Clinical manifestations comprise a unique facial appearance with cleft lip/palate, ectodermal dysplasia, cutaneous syndactyly of the fingers and/or toes, and in some cases, mental retardation. We present the first report, to our knowledge, of an Asian individual with cleft lip/palate-ectodermal dysplasia syndrome with a novel PVRL1 mutation. A 7-year-old Japanese boy, the first child of a consanguineous marriage, showed hypohidrotic ectodermal dysplasia with sparse, brittle, fine, dry hair and hypodontia, the unique facial appearance with cleft lip/palate, cutaneous syndactyly of the fingers and mild mental retardation. Scanning electron microscopic examination of the hair demonstrated pili torti and pili trianguli et canaliculi. Mutation analysis of exon 2 of PVRL1 revealed a novel homozygous nonsense mutation, c.400C>T (p.Arg134*). His parents were heterozygous for the mutant alleles. All four PVRL1 mutations identified in cleft lip/palate-ectodermal dysplasia syndrome to date, including this study, resulted in truncated proteins that lack the transmembrane domain and intracellular domain of nectin-1, which is necessary to initiate the cell-cell adhesion process.

  7. Identification of minor secondary metabolites from the latex of Croton lechleri (Muell-Arg) and evaluation of their antioxidant activity.

    Science.gov (United States)

    De Marino, Simona; Gala, Fulvio; Zollo, Franco; Vitalini, Sara; Fico, Gelsomina; Visioli, Francesco; Iorizzi, Maria

    2008-01-01

    Dragon's blood (Sangre de drago), a viscous red sap derived from Croton lechleri Muell-Arg (Euphorbiaceae), is extensively used by indigenous cultures of the Amazonian basin for its wound healing properties. The aim of this study was to identify the minor secondary metabolites and test the antioxidant activity of this sustance. A bioguided fractionation of the n-hexane, chloroform, n-butanol, and aqueous extracts led to the isolation of 15 compounds: three megastigmanes, four flavan-3-ols, three phenylpropanoids, three lignans, a clerodane, and the alkaloid taspine. In addition to these known molecules, six compounds were isolated and identified for the first time in the latex: blumenol B, blumenol C, 4,5-dihydroblumenol A, erythro-guaiacyl-glyceryl-beta-O-4'- dihydroconiferyl ether, 2-[4-(3-hydroxypropyl)-2-methoxyphenoxy]-propane-1,3-diol and floribundic acid glucoside. Combinations of spectroscopic methods ((1)H-, (13)C- NMR and 2D-NMR experiments), ESI-MS, and literature comparisons were used for compound identification. In vitro antioxidant activities were assessed by DPPH, total antioxidant capacity and lipid peroxidation assays. Flavan-3-ols derivatives (as major phenolic compounds in the latex) exhibited the highest antioxidant activity.

  8. p.Pro4Arg mutation in LMNA gene: a new atypical progeria phenotype without metabolism abnormalities.

    Science.gov (United States)

    Guo, Hong; Luo, Na; Hao, Fei; Bai, Yun

    2014-08-01

    Hutchinson-Gilford progeria syndrome (HGPS) is a typical presenile disorder, with mutation in the LMNA gene. Besides HGPS, mutations in LMNA gene have also been reported in atypical progeroid syndrome (APS). The objective of the study was to investigate the phenotype and molecular basis of APS in a Chinese family. LMNA gene mutations were also reviewed to identify the phenotypic and pathogenic differences among APS. Two siblings in a non-consanguineous Chinese family with atypical progeria were reported. The clinical features were observed, including presenile manifestations such as bird-like facial appearance, generalized lipodystrophy involving the extremities and mottled hyperpigmentation on the trunk and extremities. A heterozygous mutation c.11C>G (p.Pro4Arg) of the LMNA gene was detected in the two patients. 28 different variants of the LMNA gene have been reported in APS families, spreading over almost all the 12 exons of the LMNA gene with some hot-spot regions. This is the first detailed description of an APS family without metabolism abnormalities. APS patients share most of the clinical features, but there may be some distinct features in different ethnic groups.

  9. Assessment of haemolytic, cytotoxic and free radical scavenging activities of an underutilized fruit, Baccaurea ramiflora Lour. (Roxb.) Muell. Arg.

    Science.gov (United States)

    Saha, Manas Ranjan; Dey, Priyankar; Chaudhuri, Tapas Kumar; Goyal, Arvind Kr; Sarker, Dilip De; Sen, Arnab

    2016-02-01

    Baccaurea ramiflora Lour. (Roxb.) Muell. Arg. is an underutilized juicy fruit bearing plant found in sub-Himalayan area, South China, Indo-Burma region, etc. The fruit is considered to be nutritive, and in this study, we evaluated its antioxidant, haemolytic and cytotoxic properties. The juice was examined for the quenching activity of hydroxyl radical, nitric oxide, singlet oxygen, peroxynitrite, total antioxidant activity (TAA), erythrocyte membrane stabilizing activity (EMSA) along with quantification of phenolic and flavonoid contents and also tested for its potential activity as iron chelator, inhibitor of lipid peroxidation and total reducing power. Principal component analysis (PCA) and hierarchical cluster analysis (HCA) were also performed to correlate antioxidant capacities with the phenolic and flavonoid content. Haemolytic activity on murine erythrocyte and MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) cytotoxic test was performed on murine splenocytes, thymocytes, hepatocytes and peritoneal exudates macrophage to examine the cytotoxic effect of its juice. The result exhibited its potent free radical scavenging activity. In case of TAA, DPPH (2, 2-diphenyl-1-picrylhydrazyl), EMSA and lipid peroxidation, the fruit juice was found to have significant (P cytotoxic test confirms that the juice does not contain any cytotoxic effect and the fruit is safe for consumption. Fourier transform infrared (FTIR) spectra analysis exhibited high possibility of presence of flavonoid compounds in the juice.

  10. Growth of endothelial cells on different concentrations of Gly-Arg-Gly-Asp photochemically grafted in polyethylene glycol modified polyurethane.

    Science.gov (United States)

    Lin, Y S; Wang, S S; Chung, T W; Wang, Y H; Chiou, S H; Hsu, J J; Chou, N K; Hsieh, K H; Chu, S H

    2001-08-01

    To improve endothelial cell adhesion and growth on the surface of polyethylene glycol modified polyurethane (PU-PEG), cell adhesive peptide Gly-Arg-Gly-Asp (GRGD) was photochemically grafted to the surface. The surface grafted GRGD-N-Succinimidyl-6-[4'-azido-2'-nitrophenylamino]hexanoate (SANPAH) on a PU-PEG surface was performed by adsorption and subsequent ultraviolet irradiation. Fourier transform infrared spectra (FTIR) and electron spectroscopy for chemical analysis (ESCA) confirmed the GRGD grafted to form a PU-PEG-GRGD surface. The composition fraction of nitrogen calculated from ESCA analysis for the PU-PEG-GRGD surface was well correlated with the concentration of GRGD to be immobilized. Human umbilical vein endothelial cells (ECs) were well adhered and growing on the PU-PEG-GRGD surface. Moreover, the viability of ECs growing on PU-PEG-GRGD surfaces, analyzed by MTT test, was also well correlated with the GRGD concentrations immobilized on the surface. With photochemical techniques, we could manipulate different contents of GRGD to form multiple regions of PU-PEG-GRGD surface that could enhance the growth of ECs on the surface, and the enhancement efficiency was well correlated with GRGD contents.

  11. Identification of Minor Secondary Metabolites from the Latex of Croton lechleri (Muell-Arg and Evaluation of Their Antioxidant Activity

    Directory of Open Access Journals (Sweden)

    Maria Iorizzi

    2008-06-01

    Full Text Available Dragon’s blood (Sangre de drago, a viscous red sap derived from Croton lechleri Muell-Arg (Euphorbiaceae, is extensively used by indigenous cultures of the Amazonian basin for its wound healing properties. The aim of this study was to identify the minor secondary metabolites and test the antioxidant activity of this sustance. A bioguided fractionation of the n-hexane, chloroform, n-butanol, and aqueous extracts led to the isolation of 15 compounds: three megastigmanes, four flavan-3-ols, three phenylpropanoids, three lignans, a clerodane, and the alkaloid taspine. In addition to these known molecules, six compounds were isolated and identified for the first time in the latex: blumenol B, blumenol C, 4,5-dihydroblumenol A, erythro-guaiacyl-glyceryl-β-O-4’- dihydroconiferyl ether, 2-[4-(3-hydroxypropyl-2-methoxyphenoxy]-propane-1,3-diol and floribundic acid glucoside. Combinations of spectroscopic methods (1H-, 13C- NMR and 2D-NMR experiments, ESI-MS, and literature comparisons were used for compound identification. In vitro antioxidant activities were assessed by DPPH, total antioxidant capacity and lipid peroxidation assays. Flavan-3-ols derivatives (as major phenolic compounds in the latex exhibited the highest antioxidant activity.

  12. Diverse Phenotypic Expression of Cardiomyopathies in a Family with TNNI3 p.Arg145Trp Mutation

    Science.gov (United States)

    Hwang, Ji-won; Jang, Mi-Ae; Jang, Shin Yi; Seo, Soo Hyun; Seong, Moon-Woo; Park, Sung Sup; Ki, Chang-Seok

    2017-01-01

    Genetic diagnosis of cardiomyopathies is challenging, due to the marked genetic and allelic heterogeneity and the lack of knowledge of the mutations that lead to clinical phenotypes. Here, we present the case of a large family, in which a single TNNI3 mutation caused variable phenotypic expression, ranging from restrictive cardiomyopathy (RCMP) to hypertrophic cardiomyopathy (HCMP) to near-normal phenotype. The proband was a 57-year-old female with HCMP. Examining the family history revealed that her elder sister had expired due to severe RCMP. Using a next-generation sequencing-based gene panel to analyze the proband, we identified a known TNNI3 gene mutation, c.433C>T, which is predicted to cause an amino acid substitution (p.Arg145Trp) in the highly conserved inhibitory region of the cardiac troponin I protein. Sanger sequencing confirmed that six relatives with RCMP or near-normal phenotypes also carried this mutation. To our knowledge, this is the first genetically confirmed family with diverse phenotypic expression of cardiomyopathies in Korea. Our findings demonstrate familial implications, where a single mutation in a sarcomere protein can cause diverse phenotypic expression of cardiomyopathies.

  13. Starter substrate specificities of wild-type and mutant polyketide synthases from Rutaceae.

    Science.gov (United States)

    Lukacin, Richard; Schreiner, Stephan; Silber, Katrin; Matern, Ulrich

    2005-02-01

    Chalcone synthases (CHSs) and acridone synthases (ACSs) belong to the superfamily of type III polyketide synthases (PKSs) and condense the starter substrate 4-coumaroyl-CoA or N-methylanthraniloyl-CoA with three malonyl-CoAs to produce flavonoids and acridone alkaloids, respectively. ACSs which have been cloned exclusively from Ruta graveolens share about 75-85% polypeptide sequence homology with CHSs from other plant families, while 90% similarity was observed with CHSs from Rutaceae, i.e., R. graveolens, Citrus sinensis and Dictamnus albus. CHSs cloned from many plants do not accept N-methylanthraniloyl-CoA as a starter substrate, whereas ACSs were shown to possess some side activity with 4-coumaroyl-CoA. The transformation of an ACS to a functional CHS with 10% residual ACS activity was accomplished previously by substitution of three amino acids through the corresponding residues from Ruta-CHS1 (Ser132Thr, Ala133Ser and Val265Phe). Therefore, the reverse triple mutation of Ruta-CHS1 (mutant R2) was generated, which affected only insignificantly the CHS activity and did not confer ACS activity. However, competitive inhibition of CHS activity by N-methylanthraniloyl-CoA was observed for the mutant in contrast to wild-type CHSs. Homology modeling of ACS2 with docking of 1,3-dihydroxy-N-methylacridone suggested that the starter substrates for CHS or ACS reaction are placed in different topographies in the active site pocket. Additional site specific substitutions (Asp205Pro/Thr206Asp/His207Ala or Arg60Thr and Val100Ala/Gly218Ala, respectively) diminished the CHS activity to 75-50% of the wild-type CHS1 without promoting ACS activity. The results suggest that conformational changes in the periphery beyond the active site cavity volumes determine the product formation by ACSs vs. CHSs in R. graveolens. It is likely that ACS has evolved from CHS, but the sole enlargement of the active site pocket as in CHS1 mutant R2 is insufficient to explain this process.

  14. Relationship between Trp64Arg mutation in the β3-adrenergic receptor gene and metabolic syndrome: a seven-year follow-up study

    Institute of Scientific and Technical Information of China (English)

    ZHU Lü-yun; HU Li-ye; LI Xiao-ling; WANG Guang-yu; SHAN Wei; MA Li-cheng; WANG Xiu-hui

    2010-01-01

    Background It has been shown that the β3-adrenergic receptor (β3-AR) gene Trp64Arg mutation was closely related to obesity and insulin resistance, and may be related to the prevalence of metabolic syndrome (MS). The aim of this study was to investigate the relationship between the 33-AR gene mutation and the prevalence of MS. Methods A seven-year follow-up study was initiated in 2000, with 496 samples of simplex obese subjects (body mass index ≥25 kg/m2) and 248 normal-weight subjects. According to the β3-AR genotypes, the subjects were classified as Trp64 homozygote group and Arg64 carrier group and after 7 years the prevalence of MS was determined. Results According to the baseline profile, there were no significant differences in the adiposity, blood pressure, lipid profile, fasting plasma glucose and fasting insulin between Trp64 homozygote group and Arg64 carrier group either in obesity or normal-weight subjects. The results of follow-up study indicated that in obese men the prevalence rate of MS was much higher in Arg64 carrier group than that in Trp64 homozygote group (54.76% vs. 40.85%, P <0.05), but there was no statistical difference in women of the above groups. The prevalence rate of MS in obese men of both Trp64 homozygote group and Arg64 carrier obese group were obviously higher than that in women of the above groups (40.85% vs. 18.27% and 54.76% vs 21.28%, all P <0.005). Differences were not statistically significant in the prevalence of MS for normal weight Trp64 homozygote group and normal weight Arg64 carrier group, either between men, between women, or between men and women. Comparison of populations indicated that no matter with the β3-AR gene mutation or not, the prevalence of MS in obese subjects was significantly higher than normal weight subjects (X2=28.240 and x2=15.586, all P <0.005). Logistic analysis showed that the mutation of β3-AR gene was associated with the prevalence of MS in men.

  15. pH-sensitive CDP-diglyceride synthetase mutants of Escherichia coli: phenotypic suppression by mutations at a second site.

    Science.gov (United States)

    Ganong, B R; Raetz, C R

    1983-01-01

    In Escherichia coli, mutations which lower the level of CDP-diglyceride synthetase are designated cds and map at min 4. The cds-8 mutation resulted in strikingly defective enzyme activity and also rendered cells pH sensitive for growth. Both the inhibition of growth and the massive accumulation of phosphatidic acid which occur in a cds-8 mutant at pH 8 were suppressed by mutations at a second locus, designated cdsS, which mapped between argG and gltB near min 68. The cdsS3 mutation by itself did not affect CDP-diglyceride synthetase activity in wild-type cells, but it caused a twofold stimulation of the residual activity present in strains harboring cds-8. Both the insensitivity to pH and the twofold stimulation of residual activity were lost by introduction of an F' strain carrying cdsS+ into a recA1 cds-8 cdsS3 host. When a culture of a cds-8 cdsS+ strain was shifted to pH 8, the residual specific activity of synthetase dropped by 75% within 100 min. In a cds-8 cdsS3 double mutant under the same conditions, the activity declined appreciably less, about to the level found in the cds-8 cdsS+ strain under permissive conditions (pH 6). Thus, it appears that mutations in the cdsS gene suppress the pH sensitivity of cds mutants by inhibiting the decay of residual CDP-diglyceride synthetase activity at the nonpermissive pH. The cdsS locus appears to be distinct from any known nonsense or missense suppressor. PMID:6296051

  16. Inducement and identification of an endosperm mutant in maize

    African Journals Online (AJOL)

    ajl yemi

    2011-11-30

    Nov 30, 2011 ... “super sweet” phenotype were derived from the mutated offspring. ... characteristics and distinguished molecular mechanism to the previous mutants of gene sh2, these three mutant lines are ...... Physical association of starch biosynthetic ... reduced seedling mutant in oilseed rape, Brassica napus, for.

  17. Acquisition of pro-oxidant activity of fALS-linked SOD1 mutants as revealed using circular dichroism and UV-resonance Raman spectroscopy

    Science.gov (United States)

    Fujimaki, Nobuhiro; Nishiya, Ken; Miura, Takashi; Nakabayashi, Takakazu

    2016-11-01

    The acquisition of pro-oxidant activity of the mutated form of human Cu, Zn-superoxide dismutase (SOD1) has been investigated to clarify the relationship between mutations in SOD1 and the pathogenesis of amyotrophic lateral sclerosis (ALS). Ala4 → Val (A4V) and Gly93 → Ala (G93A) mutants, which are representative ALS-linked SOD1 mutants, have been found to exhibit both the denaturation and the gain of pro-oxidant activity after incubation in the apo-form at a physiological condition of 37 °C and pH 7.4 and the rebinding of Cu2+. These characteristics are similar to those previously reported for the His43 → Arg (H43R) mutant. UV-resonance Raman spectra indicated that the coordination structure of the Cu-binding site catalyzing the oxidation reaction is the same among the denatured A4V, G93A, and H43R. Since wild-type SOD1 does not exhibit the denaturation in its apo-form at 37 °C and pH 7.4, the instability of the protein structure due to mutation can be considered as a significant factor that induces the denaturation and the subsequent pro-oxidant activity.

  18. Arg279 is the key regulator of coenzyme selectivity in the flavin-dependent ornithine monooxygenase SidA.

    Science.gov (United States)

    Robinson, Reeder; Franceschini, Stefano; Fedkenheuer, Michael; Rodriguez, Pedro J; Ellerbrock, Jacob; Romero, Elvira; Echandi, Maria Paulina; Martin Del Campo, Julia S; Sobrado, Pablo

    2014-04-01

    Siderophore A (SidA) is a flavin-dependent monooxygenase that catalyzes the NAD(P)H- and oxygen-dependent hydroxylation of ornithine in the biosynthesis of siderophores in Aspergillus fumigatus and is essential for virulence. SidA can utilize both NADPH or NADH for activity; however, the enzyme is selective for NADPH. Structural analysis shows that R279 interacts with the 2'-phosphate of NADPH. To probe the role of electrostatic interactions in coenzyme selectivity, R279 was mutated to both an alanine and a glutamate. The mutant proteins were active but highly uncoupled, oxidizing NADPH and producing hydrogen peroxide instead of hydroxylated ornithine. For wtSidA, the catalytic efficiency was 6-fold higher with NADPH as compared to NADH. For the R279A mutant the catalytic efficiency was the same with both coenyzmes, while for the R279E mutant the catalytic efficiency was 5-fold higher with NADH. The effects are mainly due to an increase in the KD values, as no major changes on the kcat or flavin reduction values were observed. Thus, the absence of a positive charge leads to no coenzyme selectivity while introduction of a negative charge leads to preference for NADH. Flavin fluorescence studies suggest altered interaction between the flavin and NADP⁺ in the mutant enzymes. The effects are caused by different binding modes of the coenzyme upon removal of the positive charge at position 279, as no major conformational changes were observed in the structure for R279A. The results indicate that the positive charge at position 279 is critical for tight binding of NADPH and efficient hydroxylation. Copyright © 2014 Elsevier B.V. All rights reserved.

  19. Esterase polymorphism in remanant populations of Aspidosperma polyneuron Müll.Arg. (Apocynaceae Polimorfismo de esterases em populações remanescentes de Aspidosperma polyneuron Müll.Arg. (Apocynaceae

    Directory of Open Access Journals (Sweden)

    Vanda Marilza de Carvalho

    2004-10-01

    Full Text Available The population genetic structure of the endangered tree species Aspidosperma polyneuron Mull.Arg. (Apocynaceae was reported based on analysis of esterase polymorphism in two remanant populations. Allelic variation was detected at three isoesterase loci (Est-3, Est-9, and Est-10. The proportion of polymorphic loci for both populations was 30% and deviation from Hardy-Weinberg equilibrium was observed for the Est-3 locus observed in the northern population. Segregation distortion and the lower level of observed and expected heterozygosity in this population were attributed to founder genotype. The high genetic identity values for northern and northwestern populations are in accordance with the low levels of interpopulation genetic divergence demonstrated by the F(ST (0.03 value. The F(IS value (0.23 indicated moderate levels of inbreeding. A. polyneuron can be indicated as an example of endangered species suggesting high genetic variation in contrast to the low genetic variation reported for endangered species. The esterase isozymes may be a good genetic marker for studies of natural A. polyneuron populations.A análise do polimorfismo de isozimas esterases foi usada para reportar a estrutura genética de duas populações remanecentes da espécie de árvore em extinção Aspidosperma polyneuron Müll.Arg. (Apocynaceae. Variação alélica foi detectada em três locos de isoesterases (Est-3, Est-9, e Est-10. A proporção de locos polimórficos de ambas as populações foi de 30%, sendo observado um desvio do equilíbrio de Hardy-Weinberg no loco Est-3 na população da região norte do Estado do Paraná. Uma distorção na segregação e um mais baixo nível de heterozigosidade observada e esperada nesta população foram atribuídos ao efeito do genótipo fundador. Os valores altos de identidade genética das populações do norte e noroeste do Estado estão de acordo com o baixo nível de divergência genética interpopulacional demonstrado

  20. Mutant prevention concentration and mutant selection window for 10 antimicrobial agents against Rhodococcus equi.

    Science.gov (United States)

    Berghaus, Londa J; Giguère, Steeve; Guldbech, Kristen

    2013-10-25

    The objectives of this study were to determine the mutant prevention concentration (MPC), time above the MPC and mutant selection window for 10 antimicrobial agents against Rhodococcus equi and to determine if the combination of a macrolide with rifampin would decrease emergence of resistant mutants. Antimicrobial agents investigated (erythromycin, clarithromycin, azithromycin, rifampin, amikacin, gentamicin, enrofloxacin, vancomycin, imipenem, and doxycycline) were selected based on in vitro activity and frequency of use in foals or people infected with R. equi. Each antimicrobial agent or combination of agents was evaluated against four virulent strains of R. equi. MPC were determined using an agar plate assay. Pharmacodynamic parameters were calculated using published plasma and pulmonary pharmacokinetic variables. There was a significant (Pequi. Copyright © 2013 Elsevier B.V. All rights reserved.

  1. The MECP2 variant c.925C>T (p.Arg309Trp) causes intellectual disability in both males and females without classic features of Rett syndrome.

    Science.gov (United States)

    Schönewolf-Greulich, B; Tejada, M-I; Stephens, K; Hadzsiev, K; Gauthier, J; Brøndum-Nielsen, K; Pfundt, R; Ravn, K; Maortua, H; Gener, B; Martínez-Bouzas, C; Piton, A; Rouleau, G; Clayton-Smith, J; Kleefstra, T; Bisgaard, A-M; Tümer, Z

    2016-06-01

    Missense MECP2 variants can have various phenotypic effects ranging from a normal phenotype to typical Rett syndrome (RTT). In females, the phenotype can also be influenced by the X-inactivation pattern. In this study, we present detailed clinical descriptions of six patients with a rare base-pair substitution affecting Arg309 at the C-terminal end of the transcriptional repression domain (TRD). All patients have intellectual disability and present with some RTT features, but they do not fulfill the clinical criteria for typical or atypical RTT. Most of the patients also have mild facial dysmorphism. Intriguingly, the mother of an affected male patient is an asymptomatic carrier of this variant. It is therefore likely that the p.(Arg309Trp) variation does not necessarily lead to male lethality, and it results in a wide range of clinical features in females, probably influenced by different X-inactivation patterns in target tissues.

  2. Utilización de otros láseres en odontología: Argón, Nd:YAP y Ho:YAG

    OpenAIRE

    Guinot Moya, Rosa; España Tost, A. J. (Antonio Jesús); Berini Aytés, Leonardo; Gay Escoda, Cosme

    2004-01-01

    Las ventajas que la incorporación del láser ha proporcionado a la Odontología abarcan todos los ámbitos. Así, los láseres de Argón, de Nd:YAP y de Ho:YAG, tienen aplicaciones muy concretas y de gran interés tanto en el campo de la terapéutica dental como en la especialida de cirugía bucal. Las aplicaciones principales del láser de Argón se centran en la polimerización de los materiales de restauración, en la endodoncia y dentro del ámbito de la cirugía bucal, en el corte de tejidos blandos, p...

  3. Change in microbial community in landfill refuse contaminated with antibiotics facilitates denitrification more than the increase in ARG over long-term

    Science.gov (United States)

    Wu, Dong; Chen, Guanzhou; Zhang, Xiaojun; Yang, Kai; Xie, Bing

    2017-01-01

    In this study, the addition of sulfamethazine (SMT) to landfill refuse decreased nitrogen intermediates (e.g. N2O and NO) and dinitrogen (N2) gas fluxes to 5%) for the dominant bacterial hosts (Rhodothermus, ~20%) harboring nosZ and norB genes that significantly correlated with nitrogen emission pattern, while sulfamethazine amendment completely reduced the relative abundance of the “original inhabitants” functioning to produce NOx gas reduction. The main ARG carriers (Pseudomonas) that were substantially enriched in the SMT group had lower levels of denitrifying functional genes, which could imply that denitrification is influenced more by bacterial dynamics than by abundance of ARGs under antibiotic pressures. PMID:28120869

  4. Characterisation of cuticular mutants in Arabidopsis thaliana

    OpenAIRE

    Faust, Andrea

    2006-01-01

    Plants are protected by the extracellular cuticle, which is made up of cutin, cutan and waxes. The cutin composition of a variety of plants has been known and models of the biosynthesis of cutin monomers exist but not many enzymes have been identified. It is generally accepted that a defect in the cuticle leads to an organ fusion phenotype. In the model plant A. thaliana many fusion mutants have been identified but the identification of genes involved have not lead to a complete picture of th...

  5. Effects of Arc/Arg3.1 gene deletion on rhythmic synchronization of hippocampal CA1 neurons during locomotor activity and sleep.

    Science.gov (United States)

    Malkki, Hemi A I; Mertens, Paul E C; Lankelma, Jan V; Vinck, Martin; van Schalkwijk, Frank J; van Mourik-Donga, Laura B; Battaglia, Francesco P; Mahlke, Claudia; Kuhl, Dietmar; Pennartz, Cyriel M A

    2016-05-01

    The activity-regulated cytoskeletal-associated protein/activity regulated gene (Arc/Arg3.1) is crucial for long-term synaptic plasticity and memory formation. However, the neurophysiological substrates of memory deficits occurring in the absence of Arc/Arg3.1 are unknown. We compared hippocampal CA1 single-unit and local field potential (LFP) activity in Arc/Arg3.1 knockout and wild-type mice during track running and flanking sleep periods. Locomotor activity, basic firing and spatial coding properties of CA1 cells in knockout mice were not different from wild-type mice. During active behavior, however, knockout animals showed a significantly shifted balance in LFP power, with a relative loss in high-frequency (beta-2 and gamma) bands compared to low-frequency bands. Moreover, during track-running, knockout mice showed a decrease in phase locking of spiking activity to LFP oscillations in theta, beta and gamma bands. Sleep architecture in knockout mice was not grossly abnormal. Sharp-wave ripples, which have been associated with memory consolidation and replay, showed only minor differences in dynamics and amplitude. Altogether, these findings suggest that Arc/Arg3.1 effects on memory formation are not only manifested at the level of molecular pathways regulating synaptic plasticity, but also at the systems level. The disrupted power balance in theta, beta and gamma rhythmicity and concomitant loss of spike-field phase locking may affect memory encoding during initial storage and memory consolidation stages.

  6. Potential impact of (rs 4645878) BAX promoter -248G>A and (rs 1042522) TP53 72Arg>pro polymorphisms on epithelial ovarian cancer patients.

    Science.gov (United States)

    Dholariya, S; Mir, R; Zuberi, M; Yadav, P; Gandhi, G; Khurana, N; Saxena, A; Ray, P C

    2016-01-01

    In India, Epithelial ovarian cancer has emerged as one of the most common malignancies affecting women. Tumor protein 53 (TP53) induces expression of the B cell lymphoma 2-associated X protein (BAX) gene by directly binding to the TP53-binding element in the BAX promoter. Therefore, we hypothesized that single-nucleotide polymorphism of BAX promoter -248G>A and TP53 72Arg>Pro gene may jointly contribute to ovarian cancer risk. This study aimed at exploring the association of BAX promoter -248G>A and TP53 72Arg>Pro gene polymorphism with risk of developing EOC and its clinicopathological features and to evaluate gene-gene interaction of these two polymorphisms with risk of developing EOC. The study was conducted on 70 Epithelial ovarian cancer patients and 70 healthy controls. Genotyping of p53 codon 72 and BAX promoter gene was examined by ASO-PCR and PICA-PCR, respectively. Odds ratios and 95 % confidence intervals were calculated. We found an increased cancer risk associated with the BAX AA (ORs = 4.1, 95 %, CI = 1.23-13.97) genotype. An increased risk was also associated with the TP53 Pro/Pro (OR = 4.4, 95 % CI = 1.40-13.99) and Arg/Pro genotype (OR = 2.3, 95 % CI = 1.13-4.86). The gene-gene interaction of these polymorphisms increased EOC risk in a more than additive manner (ORs for the presence of both BAX AA and TP53 Arg/Pro genotypes = 8.7, 95 % CI = 1.66-45.48). BAX GG genotype was associated with adverse staging of cancer (P = 0.01). The findings suggest that polymorphism of BAX and TP53 genes may be potential genetic modifiers for developing ovarian cancer.

  7. Lack of evidence of HPV etiology of prostate cancer following radical surgery and higher frequency of the Arg/Pro genotype in turkish men with prostate cancer

    Science.gov (United States)

    Aydin, Merve; Bozkurt, Aliseydi; Cikman, Aytekin; Gulhan, Baris; Karabakan, Mehmet; Gokce, Aysun; Alper, Murat; Kara, Murat

    2017-01-01

    ABSTRACT Objectives The aim of this study was to assess the possible role of HPV in the development of prostate cancer (PCa) and investigate the distribution of the p53 codon 72 polymorphism in PCa in a Turkish population. Materials and methods A total of 96 tissues, which had been obtained using a radical surgery method, formalin-fixed and parafin-embedded, were used in this study. The study group consisted of 60 PCa tissues (open radical prostatectomy) and the control group contained 36 benign prostatic hyperplasia tissues (BPH) (transvesical open prostatectomy). The presence of HPV and the p53 codon 72 polymorphism was investigated in both groups using real-time PCR and pyrosequencing. Results The results of the real-time PCR showed no HPV DNA in any of the 36 BPH tissue samples. HPV-DNA was positive in only 1 of the 60 PCa samples (1.7%). The HPV type of this sample was identified as HPV-57. The distribution of the three genotypes, Arg/Arg, Arg/Pro and Pro/Pro was found to be 45.6, 45.6, and 8.8% in the PCa group and 57.1%, 34.3% and 8.6% in the control group, respectively. Compared with the control group, patients with PCa had a higher frequency of the Arg/Pro genotype and Proline allele (odds ratio (OR)=1.67, 95% confidence interval (CI)=0.68-4.09, p=0.044; OR=1.13, 95% CI=0.76-1.68, p=0.021, respectively). Conclusions The results of the study do not support the hyphothesis that prostate cancer is associated with HPV infection but indicated that Proline allele can be a risk factor in the development of PCa in the Turkish population. PMID:28124524

  8. Lack of evidence of HPV etiology of prostate cancer following radical surgery and higher frequency of the Arg/Pro genotype in turkish men with prostate cancer

    Directory of Open Access Journals (Sweden)

    Merve Aydin

    Full Text Available ABSTRACT Objectives The aim of this study was to assess the possible role of HPV in the development of prostate cancer (PCa and investigate the distribution of the p53 codon 72 polymorphism in PCa in a Turkish population. Materials and methods A total of 96 tissues, which had been obtained using a radical surgery method, formalin-fixed and parafin-embedded, were used in this study. The study group consisted of 60 PCa tissues (open radical prostatectomy and the control group contained 36 benign prostatic hyperplasia tissues (BPH (transvesical open prostatectomy. The presence of HPV and the p53 codon 72 polymorphism was investigated in both groups using real-time PCR and pyrosequencing. Results The results of the real-time PCR showed no HPV DNA in any of the 36 BPH tissue samples. HPV-DNA was positive in only 1 of the 60 PCa samples (1.7%. The HPV type of this sample was identified as HPV-57. The distribution of the three genotypes, Arg/Arg, Arg/Pro and Pro/Pro was found to be 45.6, 45.6, and 8.8% in the PCa group and 57.1%, 34.3% and 8.6% in the control group, respectively. Compared with the control group, patients with PCa had a higher frequency of the Arg/Pro genotype and Proline allele (odds ratio (OR=1.67, 95% confidence interval (CI=0.68-4.09, p=0.044; OR=1.13, 95% CI=0.76-1.68, p=0.021, respectively. Conclusions The results of the study do not support the hyphothesis that prostate cancer is associated with HPV infection but indicated that Proline allele can be a risk factor in the development of PCa in the Turkish population.

  9. Role of Arg228 in the phosphorylation of galactokinase: the mechanism of GHMP kinases by quantum mechanics/molecular mechanics studies.

    Science.gov (United States)

    Huang, Meilan; Li, Xiaozhou; Zou, Jian-Wei; Timson, David J

    2013-07-16

    GHMP kinases are a group of structurally related small molecule kinases. They have been found in all kingdoms of life and are mostly responsible for catalyzing the ATP-dependent phosphorylation of intermediary metabolites. Although the GHMP kinases are of clinical, pharmaceutical, and biotechnological importance, the mechanism of GHMP kinases is controversial. A catalytic base mechanism was suggested for mevalonate kinase that has a structural feature of the γ-phosphate of ATP close to an aspartate residue; however, for one GHMP family member, homoserine kinase, where the residue acting as general base is absent, a direct phosphorylation mechanism was suggested. Furthermore, it was proposed by some authors that all the GHMP kinases function by a similar mechanism. This controversy in mechanism has limited our ability to exploit these enzymes as drug targets and in biotechnology. Here the phosphorylation reaction mechanism of the human galactokinase, a member of the GHMP kinase family, was investigated using molecular dynamics simulations and density functional theory-based quantum mechanics/molecular mechanics calculations (B3LYP-D/AMBER99). The reaction coordinates were localized by potential energy scan using an adiabatic mapping method. Our results indicate that a highly conserved Glu174 captures Arg105 in the proximity of the α-phosphate of ATP, forming a H-bond network; therefore, the mobility of ATP in the large oxyanion hole is restricted. Arg228 functions to stabilize the negative charge developed at the β,γ-bridging oxygen of the ATP during bond cleavage. The reaction occurs via a direct phosphorylation mechanism, and the Asp186 in the proximity of ATP does not directly participate in the reaction pathway. Because Arg228 is not conserved among GHMP kinases, reagents which form interactions with Arg228, and therefore can interrupt its function in phosphorylation, may be developed into potential selective inhibitors for galactokinase.

  10. Using of AFLP to evaluate gamma-irradiated amaranth mutants

    Directory of Open Access Journals (Sweden)

    Labajová Mária

    2013-01-01

    Full Text Available To determine which of several gamma-irradiated mutants of amaranth Ficha cultivar and K-433 hybrid are most genetically similar to their non-irradiated control genotypes, we performed amplified fragment length polymorphism (AFLP based analysis. A total of 40 selective primer combinations were used in reported analyses. First analyses of gamma-irradiated amaranth mutant lines were done used the AFLP. In the study, primers with the differentiation ability for all analysed mutant lines are reported. The very specific changes in the mutant lines´ non-coding regions based on AFLP length polymorphism were analysed. Mutant lines of the Ficha cultivar (C15, C26, C27, C82, C236 shared a genetic dissimilarity of 0,11 and their ISSR profiles are more similar to the Ficha than those of K-433 hybrid mutant lines. The K-433 mutant lines (D54, D279, D282 shared genetic dissimilarity of 0,534 but are more distinct to their control plant as a whole, as those of the Ficha mutant lines. Different AFLP fingerprints patters of the mutant lines when compared to the Ficha cultivar and K-433 hybrid AFLP profiles may be a consequence of the complex response of the intergenic space of mutant lines to the gamma-radiance. Although a genetic polymorphism was detected within accessions, the AFLP markers successfully identified all the accessions. The AFLP results are discussed by a combination of biochemical characteristics of mutant lines and their control genotypes.

  11. Distribution of soluble amino acids in maize endosperm mutants

    Directory of Open Access Journals (Sweden)

    Toro Alejandro Alberto

    2003-01-01

    Full Text Available For human nutrition the main source of vegetable proteins are cereal and legume seeds. The content of total soluble amino acids in mature endosperm of wild-type, opaque and floury maize (Zea mays L. mutants were determined by HPLC. The total absolute concentration of soluble amino acids among the mutants varied depending on the mutant. The o11 and o13 mutants exhibited the highest average content, whereas o10, fl3 and fl1 exhibited the lowest average content. In general, the mutants exhibited similar concentrations of total soluble amino acids when compared to the wild-type lines, with the clear exception of mutants o11 and fl1, with the o11 mutant exhibiting a higher concentration of total soluble amino acids when compared to its wild-type counterpart W22 and the fl1 mutant a lower concentration when compared to its wild-type counterpart Oh43. For methionine, the mutants o2 and o11 and wild-type Oh43 exhibited the highest concentrations of this amino acid. Significant differences were not observed between mutants for other amino acids such as lysine and threonine. The high lysine concentrations obtained originally for these mutants may be due to the amino acids incorporated into storage proteins, but not those present in the soluble form.

  12. Apolar Distal Pocket Mutants of Yeast Cytochrome c Peroxidase: Hydrogen Peroxide Reactivity and Cyanide Binding of the TriAla, TriVal, and TriLeu Variants

    Science.gov (United States)

    Bidwai, Anil K.; Meyen, Cassandra; Kilheeney, Heather; Wroblewski, Damian; Vitello, Lidia B.; Erman, James E.

    2012-01-01

    Three yeast cytochrome c peroxidase (CcP) variants with apolar distal heme pockets have been constructed. The CcP variants have Arg48, Trp51, and His52 mutated to either all alanines, CcP(triAla), all valines, CcP(triVal), or all leucines, CcP(triLeu). The triple mutants have detectable enzymatic activity at pH 6 but the activity is less than 0.02% that of wild-type CcP. The activity loss is primarily due to the decreased rate of reaction between the triple mutants and H2O2 compared to wild-type CcP. Spectroscopic properties and cyanide binding characteristics of the triple mutants have been investigated over the pH stability region of CcP, pH 4 to 8. The absorption spectra indicate that the CcP triple mutants have hemes that are predominantly five-coordinate, high-spin at pH 5 and six-coordinate, low-spin at pH 8. Cyanide binding to the triple mutants is biphasic indicating that the triple mutants have two slowly-exchanging conformational states with different cyanide affinities. The binding affinity for cyanide is reduced at least two orders of magnitude in the triple mutants compared to wild-type CcP and the rate of cyanide binding is reduced by four to five orders of magnitude. Correlation of the reaction rates of CcP and 12 distal pocket mutants with H2O2 and HCN suggests that both reactions require ionization of the reactants within the distal heme pocket allowing the anion to bind the heme iron. Distal pocket features that promote substrate ionization (basic residues involved in base-catalyzed substrate ionization or polar residues that can stabilize substrate anions) increase the overall rate of reaction with H2O2 and HCN while features that inhibit substrate ionization slow the reactions. PMID:23022490

  13. A new factor X defect (factor X Padua 3): a compound heterozygous between true deficiency (Gly(380)-->Arg) and an abnormality (Ser(334)-->Pro).

    Science.gov (United States)

    Vianello, F; Lombardi, A M; Boldrin, C; Luni, S; Girolami, A

    2001-11-15

    We report a novel mutation in Factor X (FX) gene which results in a phenotype without any bleeding tendency. The proband has been found to be a compound heterozygote between a novel FX true deficiency (Gly(380)-->Arg) and a previously reported dysfunctional mutation Ser(334)-->Pro (FX Marsiglia). Prothrombin time (PT) and partial thromboplastin time (PTT) were moderately prolonged and were fully corrected by the addition of normal serum. Her FX activity level varied between 8% and 19% of normal according to the method used whereas the FX antigen level was 40% of the normal control value. All the exons and intron/exon junctions of the FX gene were studied using a combined approach of polymerase chain reaction and conformation sensitive gel electrophoresis. A transversion G to A in exon 8 resulting in the replacement of Gly380 by Arg was found in the proband, in the father and in a proband's brother, whereas heterozygous FX Marsiglia was present in the proband's mother and her sister. Gly380 is strictly linked to Ser379, a component of the catalytic triad. The substitution of Gly for Arg causes the introduction of a large charged amino acid which could affect the catalytic function of FX leading to secretion problem, accounting for the cross-reactive material (CRM) negative phenotype.

  14. The association between the TP53 Arg72Pro polymorphism and colorectal cancer: An updated meta-analysis based on 32 studies

    Science.gov (United States)

    Tian, Xin; Dai, Shundong; Sun, Jing; Jiang, Shenyi; Jiang, Youhong

    2017-01-01

    Several previous studies evaluated the association between the Arg72Pro (rs1042522) polymorphism in the TP53 tumor suppressor gene and colorectal cancer (CRC). However, the results are conflicting. This meta-analysis aimed to shed new light on the precise association between TP53 variants and CRC. We analyzed 32 published case-control studies involving 8,586 cases and 10,275 controls using crude odd ratios (ORs) with 95% confidence intervals (CIs). The meta-analysis was performed using a fixed-effect or random-effects model, as appropriate. We found that the TP53 Arg72Pro polymorphism was not significantly associated with CRC risk in the overall population. However, subgroup analysis based on ethnicity revealed an increased risk of CRC among Asians (CC vs. GC+GG: OR=1.22, 95% CI: 1.02-1.45), and similar results were found for rectal cancer (CC vs. GC+GG: OR=1.34, 95% CI: 1.120-1.62). These results suggest that the TP53 Arg72Pro polymorphism CC genotype may contribute to an increased risk of CRC, especially for rectal cancer and among Asians. PMID:27901479

  15. FGFR4 Gly388Arg polymorphism may affect the clinical stage of patients with lung cancer by modulating the transcriptional profile of normal lung.

    Science.gov (United States)

    Falvella, Felicia S; Frullanti, Elisa; Galvan, Antonella; Spinola, Monica; Noci, Sara; De Cecco, Loris; Nosotti, Mario; Santambrogio, Luigi; Incarbone, Matteo; Alloisio, Marco; Calabrò, Elisa; Pastorino, Ugo; Skaug, Vidar; Haugen, Aage; Taioli, Emanuela; Dragani, Tommaso A

    2009-06-15

    The association of the fibroblast growth factor receptor 4 (FGFR4) Gly388Arg polymorphism with clinical stage and overall survival in a series of 541 Italian lung adenocarcinoma (ADCA) patients indicated a significantly decreased survival in patients carrying the rare Arg388 allele as compared to that in Gly/Gly homozygous patients [hazard ratio (HR) = 1.5; 95% confidence interval (CI) 1.1-1.9], with the decrease related to the association of the same polymorphism with clinical stage (HR = 1.8, 95% CI 1.3-2.6). By contrast, no significant association was detected in small series of either Norwegian lung ADCA patients or Italian lung squamous cell carcinoma (SQCC) patients. Single nucleotide polymorphisms of known FGFR4 ligands expressed in lung (FGF9, FGF18 and FGF19) were not associated with clinical stage or survival and showed no interaction with FGFR4. Analysis of gene expression profile in normal lungs according to FGFR4 genotype indicated a specific transcript pattern associated with the allele carrier status, suggesting a functional role for the FGFR4 polymorphism already detectable in normal lung. These findings confirm the significant association of the FGFR4 Gly388Arg polymorphism with clinical stage and overall survival in an Italian lung ADCA population and demonstrate a FGFR4 genotype-dependent transcriptional profile present in normal lung tissue.

  16. Fibroblast growth factor receptor 4 gene (FGFR4) 388Arg allele predicts prolonged survival and platinum sensitivity in advanced ovarian cancer.

    Science.gov (United States)

    Marmé, Frederik; Hielscher, Thomas; Hug, Sarah; Bondong, Sandra; Zeillinger, Robert; Castillo-Tong, Dan Cacsire; Sehouli, Jalid; Braicu, Ioana; Vergote, Ignace; Isabella, Cadron; Mahner, Sven; Ferschke, Irmgard; Rom, Joachim; Sohn, Christof; Schneeweiss, Andreas; Altevogt, Peter

    2012-08-15

    FGFR4 has been shown to play an important role in the etiology and progression of solid tumors. A single nucleotide polymorphism (SNP) within the FGFR4 gene has previously been linked to prognosis and response to chemotherapy in breast cancer and other malignancies. This study evaluates the relevance of this SNP in advanced ovarian cancer. FGFR4-genotype was analyzed in 236 patients recruited as part of the OVCAD project. Genotyping was performed on germ-line DNA using a TaqMan based genotyping assay. Results were correlated with clinicopathological variables and survival. The FGFR4 388Arg genotype was significantly associated with prolonged progression-free and overall survival (univariate: HR 0.68, p = 0.017; HR 0.49, p = 0.005; multivariate: HR 0.69, p = 0.025; HR 0.49, p = 0.006) though the positive prognostic value was restricted to patients without postoperative residual tumor. Indeed, there was a significant interaction between FGFR4 genotype and residual tumor for overall survival. Furthermore, the FGFR4 388Arg genotype significantly correlated with platinum sensitivity in the same subgroup (multivariate OR 3.81 p = 0.004). FGFR4 Arg388Gly genotype is an independent and strong context specific prognostic factor in patients with advanced ovarian cancer and could be used to predict platinum-sensitivity.

  17. Expression of FOXP3, CD14, and ARG1 in Neuroblastoma Tumor Tissue from High-Risk Patients Predicts Event-Free and Overall Survival

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    Sara Stigliani

    2015-01-01

    Full Text Available The prognosis of children with metastatic neuroblastoma (NB > 18 months at diagnosis is dismal. Since the immune status of the tumor microenvironment could play a role in the history of disease, we evaluated the expression of CD45, CD14, ARG1, CD163, CD4, FOXP3, Perforin-1 (PRF1, Granzyme B (GRMB, and IL-10 mRNAs in primary tumors at diagnosis from children with metastatic NB and tested whether the transcript levels are significantly associated to event-free and overall survival (EFS and OS, resp.. Children with high expression of CD14, ARG1 and FOXP3 mRNA in their primary tumors had significantly better EFS. Elevated expression of CD14, and FOXP3 mRNA was significantly associated to better OS. CD14 mRNA expression levels significantly correlated to all markers, with the exception of CD4. Strong positive correlations were found between PRF1 and CD163, as well as between PFR1 and FOXP3. It is worth noting that the combination of high levels of CD14, FOXP3, and ARG1 mRNAs identified a small group of patients with excellent EFS and OS, whereas low levels of CD14 were sufficient to identify patients with dismal survival. Thus, the immune status of the primary tumors of high-risk NB patients may influence the natural history of this pediatric cancer.

  18. Associations between Dietary Patterns, ADRβ2 Gln27Glu and ADRβ3 Trp64Arg with Regard to Serum Triglyceride Levels: J-MICC Study

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    Hinako Nanri

    2016-09-01

    Full Text Available Interactions between dietary patterns and 2 β-adrenergic receptor (ADRβ gene polymorphisms (ADRβ2 Gln27Glu and ADRβ3 Trp64Arg were examined with regard to the effects on serum triglyceride levels. The cross-sectional study comprised 1720 men and women (aged 35–69 years enrolled in the Japan Multi-Institutional Collaborative Cohort (J-MICC Study. Genotyping was conducted using a multiplex polymerase chain reaction-based invader assay. We used 46 items from a validated short food frequency questionnaire and examined major dietary patterns by factor analysis. We identified four dietary patterns: healthy, Western, seafood and bread patterns. There was no significant association between any dietary pattern and serum triglyceride levels. After a separate genotype-based analysis, significant interactions between ADRβ3 Trp64Arg genotype and the bread pattern (p for interaction = 0.01 were associated with serum triglyceride levels; specifically, after adjusting for confounding factors, Arg allele carriers with the bread pattern had lower serum triglycerides (p for trend = 0.01. However, the Trp/Trp homozygous subjects with the bread pattern showed no association with serum triglycerides (p for trend = 0.55. Interactions between other dietary patterns and ADRβ polymorphisms were not significant for serum triglyceride levels. Our findings suggest that ADRβ3 polymorphism modifies the effects of the bread pattern on triglyceride levels.

  19. Identification of proteinaceous inhibitors of a cysteine proteinase (an Arg-specific gingipain) from Porphyromonas gingivalis in rice grain, using targeted-proteomics approaches.

    Science.gov (United States)

    Taiyoji, Mayumi; Shitomi, Yasuyuki; Taniguchi, Masayuki; Saitoh, Eiichi; Ohtsubo, Sadami

    2009-11-01

    Porphyromonas gingivalis is known to be a major etiologic agent in the onset and progression of chronic periodontitis. Among various virulence factors that this bacterium produces, Arg- and Lys-specific cysteine proteinases (gingipains) are believed to be major determinants of the pathogenicity of P. gingivalis. Here, we report on our finding that there are inhibitors of these cysteine proteinases in a rice protein fraction. Comprehensive affinity chromatography and MS analyses resulted in the identification of 17 Arg-gingipain (Rgp)-interacting proteins in the rice endosperm. Of these, four proteins (i.e., a 26 kDa globulin, a plant lipid transfer/trypsin-alpha amylase inhibitor, the RA17 seed allergen, and an alpha amylase/trypsin inhibitor) were estimated to account for 90% of the Rgp inhibitory activity in the rice protein fraction, using a two-dimensional gel system of double-layer reverse zymography. In addition, a synthetic peptide derived from an Rgp-interacting protein, cyanate hydratase, could inhibit the growth of P. gingivalis and showed inhibitory activity against both the Arg- and Lys-gingipains. These results suggest that these rice proteins may be useful as nutraceutical ingredients for the prevention and management of periodontal diseases.

  20. Novel R-plasmid conjugal transfer inhibitory and antibacterial activities of phenolic compounds from Mallotus philippensis (Lam.) Mull. Arg.

    Science.gov (United States)

    Oyedemi, Blessing O M; Shinde, Vaibhav; Shinde, Kamlesh; Kakalou, Dionysia; Stapleton, Paul D; Gibbons, Simon

    2016-06-01

    Antimicrobial resistance severely limits the therapeutic options for many clinically important bacteria. In Gram-negative bacteria, multidrug resistance is commonly facilitated by plasmids that have the ability to accumulate and transfer refractory genes amongst bacterial populations. The aim of this study was to isolate and identify bioactive compounds from the medicinal plant Mallotus philippensis (Lam.) Mull. Arg. with both direct antibacterial properties and the capacity to inhibit plasmid conjugal transfer. A chloroform-soluble extract of M. philippensis was subjected to bioassay-guided fractionation using chromatographic and spectrometric techniques that led to the isolation of the known compounds rottlerin [5,7-dihydroxy-2,2-dimethyl-6-(2,4,6-trihydroxy-3-methyl-5-acetylbenzyl)-8-cinnamoyl-1,2-chromene] and the red compound (8-cinnamoyl-5,7-dihydroxy-2,2,6-trimethylchromene). Both compounds were characterised and elucidated using one-dimensional and two-dimensional nuclear magnetic resonance (NMR). Rottlerin and the red compound showed potent activities against a panel of clinically relevant Gram-positive bacteria, including meticillin-resistant Staphylococcus aureus (MRSA). No significant direct activities were observed against Gram-negative bacteria. However, both rottlerin and the red compound strongly inhibited conjugal transfer of the plasmids pKM101, TP114, pUB307 and R6K amongst Escherichia coli at a subinhibitory concentration of 100mg/L. Interestingly, despite the planar nature of the compounds, binding to plasmid DNA could not be demonstrated by a DNA electrophoretic mobility shift assay. These results show that rottlerin and the red compound are potential candidates for antibacterial drug lead development. Further studies are needed to elucidate the mode of inhibition of the conjugal transfer of plasmids.

  1. Arg-gly-asp-mannose-6-phosphate inhibits activation and proliferation of hepatic stellate cells in vitro

    Institute of Scientific and Technical Information of China (English)

    Lian-Sheng Wang; Ying-Wei Chen; Ding-Guo Li; Han-Ming Lu

    2006-01-01

    AIM: To investigate the effect of arg-gly-asp-mannose-6phosphate (RGD-M6P) on the activation and proliferation of primary hepatic stellate cells in vitro.METHODS: Hepatic stellate cells (HSCs) were isolated from rats by in situ collagenase perfusion of liver and 18% Nycodenz gradient centrifugation and cultured on uncoated plastic plates for 24 h with DMEM containing 10% fetal bovine serum (FBS/DMEM) before the culture medium was substituted with 2% FBS/DMEM for another 24 h. Then, HSCs were cultured in 2% FBS/DMEM with transforming growth factor β1, M6P, RGD, or RGD-M6P, respectively. Cell morphology was observed under inverted microscope, smooth muscle α-actin (α-SMA)was detected by immunocytochemistry, type Ⅲprocollagen (PCⅢ) in supernatant was determined by radioimmunoassay, and the proliferation rate of HSCs was assessed by flow cytometry.RESULTS: RGD-M6P significantly inhibited the morphological transformation and the α-SMA and PC Ⅲ expressions of HSCs in vitro and also dramatically prevented the proliferation of HSCs in vitro. Such effects were remarkably different from those of RGD or M6P.CONCLUSION: The new compound, RGD-M6P, which has a dramatic effect on primary cultured HSCs in vitro, can inhibit the transformation of HSCs in culture caused by TGFβ1, suppresses the expression of PCⅢand decreases proliferation rate of HSC. RGD-M6P can be applied as a selective drug carrier targeting at HSCs,which may be a new approach to the prevention and treatment of liver fibrosis.

  2. The diversity of the HLA-E-restricted peptide repertoire explains the immunological impact of the Arg107Gly mismatch.

    Science.gov (United States)

    Celik, Alexander A; Kraemer, Thomas; Huyton, Trevor; Blasczyk, Rainer; Bade-Döding, Christina

    2016-01-01

    Human leukocyte antigen (HLA)-E molecules are potent inhibitors of NK cell-mediated killing. Low in polymorphisms, two alleles are widely expressed among diverse populations: HLA-E*01:01 and HLA-E*01:03. Both alleles are distinguished by one SNP resulting in the substitution Arg107Gly. Both alleles present a limited set of peptides derived from class I leader sequences physiologically; however, HLA-E*01:01 presents non-canonical peptides in the absence of HLA class I molecules. To further assess the functional differences between both alleles, we analyzed the peptide repertoire of HLA-E*01:03 by applying soluble HLA technology followed by mass-spectrometric peptide sequencing. HLA-E*01:03 restricted peptides showed a length of 9-17 amino acids and differed in their biophysical properties, no overlap in the peptide repertoire of both allelic variants could be observed; however, both alleles shared marginal peptides from the same proteomic content. Artificial APCs expressing empty HLA-E*01:01 or E*01:03 molecules were generated and stabilized using cognate HLA class I-derived peptide ligands to analyze the impact of residue 107 within the HLA-E heavy chain on the NKG2/CD94 receptor engagement. Differences in peptide stabilization could be translated to the density and half-life time of peptide-HLA-E molecules on the cell surface that subsequently impacted NK cell inhibition as verified by cytotoxicity assays. Taken together, these data illustrate functional differences of HLA-E allelic variants induced by a single amino acid. Furthermore, the function of HLA-E in pathophysiologic situations when the HLA processing machinery is interrupted seems to be more emphasized than previously described, implying a crucial role for HLA-E in tumor or viral immune episodes.

  3. Effects of Condensed Tannins in Mao ( Muell. Arg. Seed Meal on Rumen Fermentation Characteristics and Nitrogen Utilization in Goats

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    P. Gunun

    2016-08-01

    Full Text Available Mao seed is a by-product of the wine and juice industry, which could be used in animal nutrition. The current study was designed to determine the effect of supplementation of mao (Antidesma thwaitesianum Muell. Arg. seed meal (MOSM containing condensed tannins (CT on rumen fermentation, nitrogen (N utilization and microbial protein synthesis in goats. Four crossbred (Thai Native×Anglo Nubian goats with initial body weight (BW 20±2 kg were randomly assigned to a 4×4 Latin square design. The four dietary treatments were MOSM supplementation at 0%, 0.8%, 1.6%, and 2.4% of total dry matter (DM intake, respectively. During the experimental periods, all goats were fed a diet containing roughage to concentrate ratio of 60:40 at 3.0% BW/d and pangola grass hay was used as a roughage source. Results showed that supplementation with MOSM did not affect feed intake, nutrient intakes and apparent nutrient digestibility (p>0.05. In addition, ruminal pH and ammonia nitrogen (NH3-N were not influenced by MOSM supplementation, whilst blood urea nitrogen was decreased quadraticly (p0.05. There were linear decreases in urinary N (p0.05. From the current study, it can be concluded that supplementation of MOSM at 1.6% to 2.4% of total DM intake can be used to modify ruminal fermentation, especially propionate and N utilization in goats, without affecting the nutrient digestibility, microbial populations and microbial protein synthesis.

  4. Pharmacological evaluation of Mallotus philippinensis (Lam. Muell.-Arg. fruit hair extract for Anti-Inflammatory, Analgesic and Hypnotic Activity

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    Mayank Gangwar

    2016-03-01

    Full Text Available Recently we observed wound healing activity of 50% ethanol extract of Mallotus philippinensis Muell. Arg (MP fruit hairs extract (MPE. In several intestinal infections, localized inflammation is of common occurrence and hence we evaluated the anti-inflammatory, analgesic and hypnotic activity of MPE in different rat experimental models. Anti-inflammatory activity was evaluated by carrageenan (acute and turpentine oil induced formalin (subacute induced paw oedema and while, granuloma pouch (subacute in rats. Analgesic and hypnotic activity of MPE was undertaken by tail-flick, hot and ndash;plate and acetic acid-induced writhing tests while pentobarbitone-induced hypnotic potentiation respectively in rats.\tMPE at a dose of 200 mg/kg at 3 hr after their administration showed inhibition of formalin-induced paw oedema by 41.60% (P<0.001 and carrageenan-induced paw oedema by 55.30% (P<0.001. After 7 days of treatments, MPE showed 38.0% (P<0.001 inhibition against formalin-induced paw oedema and reduced weight of turpentine-induced granuloma pouch by 29.6% (P<0.01 and volume of exudates by 26.1% (P<0.01 respectively. MPE (200 mg/kg showed dose-dependent elevation in pain threshold and peak analgesic effect at 120 min as evidenced by increased latency period in tail flick method and increased reaction time in hot-plate test while reduction in the number of acetic acid-induced writhes by 45.7% (P<0.001. The pentobarbitone-induced hypnosis model showed potentiation, as defined by increased duration of sleep in treated group rats as compared to control. Thus, the study revealed MPE is effective in reducing acute and sub-acute inflammation and showed effective and similar analgesic activity. This seemed to be safe in the treatment of pain and inflammation. [J Intercult Ethnopharmacol 2016; 5(1.000: 14-21

  5. Auxin physiology of the tomato mutant diageotropica

    Science.gov (United States)

    Daniel, S. G.; Rayle, D. L.; Cleland, R. E.

    1989-01-01

    The tomato (Lycopersicon esculentum, Mill.) mutant diageotropica (dgt) exhibits biochemical, physiological, and morphological abnormalities that suggest the mutation may have affected a primary site of auxin perception or action. We have compared two aspects of the auxin physiology of dgt and wild-type (VFN8) seedlings: auxin transport and cellular growth parameters. The rates of basipetal indole-3-acetic acid (IAA) polar transport are identical in hypocotyl sections of the two genotypes, but dgt sections have a slightly greater capacity for IAA transport. 2,3,5-Triiodobenzoic acid and ethylene reduce transport in both mutant and wild-type sections. The kinetics of auxin uptake into VFN8 and dgt sections are nearly identical. These results make it unlikely that an altered IAA efflux carrier or IAA uptake symport are responsible for the pleiotropic effects resulting from the dgt mutation. The lack of auxin-induced cell elongation in dgt plants is not due to insufficient turgor, as the osmotic potential of dgt cell sap is less (more negative) than that of VFN8. An auxin-induced increase in wall extensibility, as measured by the Instron technique, only occurs in the VFN8 plants. These data suggest dgt hypocotyls suffer a defect in the sequence of events culminating in auxin-induced cell wall loosening.

  6. Indy mutants: live long and prosper

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    Stewart eFrankel

    2012-02-01

    Full Text Available Indy encodes the fly homologue of a mammalian transporter of di and tricarboxylatecomponents of the Krebs cycle. Reduced expression of fly Indy or two of the C. elegansIndy homologs leads to an increase in life span. Fly and worm tissues that play key roles inintermediary metabolism are also the places where Indy genes are expressed. One of themouse homologs of Indy (mIndy is mainly expressed in the liver. It has been hypothesizedthat decreased INDY activity creates a state similar to caloric restriction (CR. Thishypothesis is supported by the physiological similarities between Indy mutant flies on highcalorie food and control flies on CR, such as increased physical activity and decreases inweight, egg production, triglyceride levels, starvation resistance, and insulin signaling. Inaddition, Indy mutant flies undergo changes in mitochondrial biogenesis also observed inCR animals. Recent findings with mIndy knockout mice support and extend the findingsfrom flies. mIndy-/- mice display an increase in hepatic mitochondrial biogenesis, lipidoxidation and decreased hepatic lipogenesis. When mIndy-/- mice are fed high calorie foodthey are protected from adiposity and insulin resistance. These findings point to INDY as apotential drug target for the treatment of metabolic syndrome, type 2 diabetes and obesity.

  7. Mutants of Saccharomyces cerevisiae with defects in acetate metabolism: isolation and characterization of Acn- mutants.

    Science.gov (United States)

    McCammon, M T

    1996-09-01

    The two carbon compounds, ethanol and acetate, can be oxidatively metabolized as well as assimilated into carbohydrate in the yeast Saccharomyces cerevisiae. The distribution of acetate metabolic enzymes among several cellular compartments, mitochondria, peroxisomes, and cytoplasm makes it an intriguing system to study complex metabolic interactions. To investigate the complex process of carbon catabolism and assimilation, mutants unable to grow on acetate were isolated. One hundred five Acn- ("ACetate Nonutilizing") mutants were sorted into 21 complementation groups with an additional 20 single mutants. Five of the groups have defects in TCA cycle enzymes: MDH1, CIT1, ACO1, IDH1, and IDH2. A defect in RTG2, involved in the retrograde communication between the mitochondrion and the nucleus, was also identified. Four genes encode enzymes of the glyoxylate cycle and gluconeogenesis: ICL1, MLS1, MDH2, and PCK1. Five other genes appear to be defective in regulating metabolic activity since elevated levels of enzymes in several metabolic pathways, including the glyoxylate cycle, gluconeogenesis, and acetyl-CoA metabolism, were detected in these mutants: ACN8, ACN9, ACN17, ACN18, and ACN42. In summary, this analysis has identified at least 22 and as many as 41 different genes involved in acetate metabolism.

  8. Sulfotransferase SULT1A1 Arg213His polymorphism with cancer risk: a meta-analysis of 53 case-control studies.

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    Juanjuan Xiao

    Full Text Available BACKGROUND: The SULT1A1 Arg213His (rs9282861 polymorphism is reported to be associated with many kinds of cancer risk. However, the findings are conflicting. For better understanding this SNP site and cancer risk, we summarized available data and performed this meta-analysis. METHODS: Data were collected from the following electronic databases: PubMed, Web of Knowledge and CNKI. The association was assessed by odd ratio (OR and the corresponding 95% confidence interval (95% CI. RESULTS: A total of 53 studies including 16733 cancer patients and 23334 controls based on the search criteria were analyzed. Overall, we found SULT1A1 Arg213His polymorphism can increase cancer risk under heterozygous (OR  1.09, 95% CI = 1.01-1.18, P = 0.040, dominant (OR = 1.10, 95% CI = 1.01-1.19, P = 0.021 and allelic (OR = 1.08, 95% CI = 1.02-1.16, P = 0.015 models. In subgroup analyses, significant associations were observed in upper aero digestive tract (UADT cancer (heterozygous model: OR = 1.62, 95% CI = 1.11-2.35, P = 0.012; dominant model: OR = 1.63, 95% CI = 1.13-2.35, P = 0.009; allelic model: OR = 1.52, 95% CI = 1.10-2.11, P = 0.012 and Indians (recessive model: OR = 1.93, 95% CI = 1.22-3.07, P = 0.005 subgroups. Hospital based study also showed marginally significant association. In the breast cancer subgroup, ethnicity and publication year revealed by meta-regression analysis and one study found by sensitivity analysis were the main sources of heterogeneity. The association between SULT1A1 Arg213His and breast cancer risk was not significant. No publication bias was detected. CONCLUSIONS: The present meta-analysis suggests that SULT1A1 Arg213His polymorphism plays an important role in carcinogenesis, which may be a genetic factor affecting individual susceptibility to UADT cancer. SULT1A1 Arg213His didn't show any association with breast cancer, but the possible risk in Asian population needs further investigation.

  9. Combined analysis of pri-miR-34b/c rs4938723 and TP53 Arg72Pro with cervical cancer risk.

    Science.gov (United States)

    Yuan, Fang; Sun, Ruifen; Chen, Peng; Liang, Yundan; Ni, Shanshan; Quan, Yi; Huang, Juan; Zhang, Lin; Gao, Linbo

    2016-05-01

    miR-34 family members can form a p53-miR-34 positive feedback loop and induce apoptosis, DNA repair, angiogenesis, and cell cycle arrest. We conducted a case-control study to examine whether two polymorphisms (i.e., rs4938723 in the promoter of pri-miR-34b/c and TP53 Arg72Pro) were linked to the carcinogenesis of cervical cancer among Chinese Han women. Genotypes of the two polymorphisms in 328 cervical cancer patients and 568 control subjects were determined by using a polymerase chain reaction-restriction fragment length polymorphism assay. We found a significantly increased cervical cancer risk in the pri-miR-34b/c rs4938723 under dominant and overdominant model (CT/CC vs. TT: adjusted OR = 1.34, 95 % CI = 1.01-1.77; CT vs. TT/CC: adjusted OR = 1.37, 95 % CI = 1.05-1.80, respectively). Increased cervical cancer risks were also found in the TP53 Arg72Pro under a heterozygous comparison and overdominant model (CG vs. GG: adjusted OR = 1.44, 95 % CI = 1.06-1.95; CG vs. GG/CC: adjusted OR = 1.47, 95 % CI = 1.12-1.94, respectively). Stratification analysis showed that patients carrying the pri-miR-34b/c rs4938723 CT genotype had a significantly increased risk for developing poorly differential status and clinical stage I. Moreover, increased cancer risks were observed for the TP53 Arg72Pro polymorphism in patients with poorly differential status, clinical stage II, and without lymph node metastasis. Combined analysis revealed that the genotypes of rs4938723 CT/CC and TP53 Arg72Pro CG/CC had an increased cervical cancer risk (OR = 2.21, 95 % CI = 1.38-3.53). These findings suggest that the pri-miR-34b/c rs4938723 and TP53 Arg72Pro polymorphisms may contribute to the genesis of cervical cancer.

  10. Elevated ARG1 expression in primary monocytes-derived macrophages as a predictor of radiation-induced acute skin toxicities in early breast cancer patients.

    Science.gov (United States)

    Jung, Karen; Sabri, Siham; Hanson, John; Xu, Yaoxian; Wang, Ying Wayne; Lai, Raymond; Abdulkarim, Bassam S

    2015-01-01

    Radiation therapy (RT) the front-line treatment after surgery for early breast cancer patients is associated with acute skin toxicities in at least 40% of treated patients. Monocyte-derived macrophages are polarized into functionally distinct (M1 or M2) activated phenotypes at injury sites by specific systemic cytokines known to play a key role in the transition between damage and repair in irradiated tissues. The role of M1 and M2 macrophages in RT-induced acute skin toxicities remains to be defined. We investigated the potential value of M1 and M2 macrophages as predictive factors of RT-induced skin toxicities in early breast cancer patients treated with adjuvant RT after lumpectomy. Blood samples collected from patients enrolled in a prospective clinical study (n = 49) were analyzed at baseline and after the first delivered 2Gy RT dose. We designed an ex vivo culture system to differentiate patient blood monocytes into macrophages and treated them with M1 or M2-inducing cytokines before quantitative analysis of their "M1/M2" activation markers, iNOS, Arg1, and TGFß1. Statistical analysis was performed to correlate experimental data to clinical assessment of acute skin toxicity using Common Toxicity Criteria (CTC) grade for objective evaluation of skin reactions. Increased ARG1 mRNA significantly correlated with higher grades of erythema, moist desquamation, and CTC grade. Multivariate analysis revealed that increased ARG1 expression in macrophages after a single RT dose was an independent prognostic factor of erythema (p = 0 .032), moist desquamation (p = 0 .027), and CTC grade (p = 0 .056). Interestingly, multivariate analysis of ARG1 mRNA expression in macrophages stimulated with IL-4 also revealed independent prognostic value for predicting acute RT-induced toxicity factors, erythema (p = 0 .069), moist desquamation (p = 0 .037), and CTC grade (p = 0 .046). To conclude, our findings underline for the first time the biological significance of increased ARG1 m

  11. Contribution of ARLTS1 Cys148Arg (T442C variant with prostate cancer risk and ARLTS1 function in prostate cancer cells.

    Directory of Open Access Journals (Sweden)

    Sanna Siltanen

    Full Text Available ARLTS1 is a recently characterized tumor suppressor gene at 13q14.3, a region frequently deleted in both sporadic and hereditary prostate cancer (PCa. ARLTS1 variants, especially Cys148Arg (T442C, increase susceptibility to different cancers, including PCa. In this study the role of Cys148Arg substitution was investigated as a risk factor for PCa using both genetic and functional analysis. Cys148Arg genotypes and expression of the ARLTS1 were explored in a large set of familial and unselected PCa cases, clinical tumor samples, xenografts, prostate cancer cell lines and benign prostatic hyperplasia (BPH samples. The frequency of the variant genotype CC was significantly higher in familial (OR = 1.67, 95% CI = 1.08-2.56, P = 0.019 and unselected patients (OR = 1.52, 95% CI = 1.18-1.97, P = 0.001 and the overall risk was increased (OR = 1.54, 95% CI = 1.20-1.98, P = 0.0007. Additional analysis with clinicopathological data revealed an association with an aggressive disease (OR = 1.28, 95% CI = 1.05-∞, P = 0.02. The CC genotype of the Cys148Arg variant was also contributing to the lowered ARLTS1 expression status in lymphoblastoid cells from familial patients. In addition significantly lowered ARLTS1 expression was observed in clinical tumor samples compared to BPH samples (P = 0.01. The ARLTS1 co-expression signature based on previously published microarray data was generated from 1587 cancer samples confirming the low expression of ARLTS1 in PCa and showed that ARLTS1 expression was strongly associated with immune processes. This study provides strong confirmation of the important role of ARLTS1 Cys148Arg variant as a contributor in PCa predisposition and a potential marker for aggressive disease outcome.

  12. Defective kernel mutants of maize. I. Genetic and lethality studies.

    Science.gov (United States)

    Neuffer, M G; Sheridan, W F

    1980-08-01

    A planting of 3,919 M(1) kernels from normal ears crossed by EMS-treated pollen produced 3,461 M(1) plants and 3,172 selfed ears. These plants yielded 2,477 (72%) total heritable changes; the selfed ears yielded 2,457 (78%) recessive mutants, including 855 (27%) recessive kernel mutants and 8 (0.23%) viable dominant mutants. The ratio of recessive to dominant mutants was 201:1. The average mutation frequency for four known loci was three per 3,172 genomes analyzed. The estimated total number of loci mutated was 535 and the estimated number of kernel mutant loci mutated was 285. Among the 855 kernel mutants, 432 had a nonviable embryo, and 59 germinated but had a lethal seedling. A sample of 194 of the latter two types was tested for heritability, lethality, chromosome arm location and endosperm-embryo interaction between mutant and nonmutant tissues in special hyper-hypoploid combinations produced by manipulation of B-A translocations. The selected 194 mutants were characterized and catalogued according to endosperm phenotype and investigated to determine their effects on the morphology and development of the associated embryo. The possibility of rescuing some of the lethal mutants by covering the mutant embryo with a normal endosperm was investigated. Ninety of these 194 mutants were located on 17 of the 18 chromosome arms tested. Nineteen of the located mutants were examined to determine the effect of having a normal embryo in the same kernel with a mutant endosperm, and vice versa, as compared to the expression observed in kernels with both embryo and endosperm in a mutant condition. In the first situation, for three of the 19 mutants, the mutant endosperm was less extreme (the embryo helped); for seven cases, the mutant endosperm was more extreme (the embryo hindered); and for nine cases, there was no change. In the reverse situation, for four cases the normal endosperm helped the mutant embryo; for 14 cases there was no change and one case was inconclusive.

  13. Structure-function analysis of CYP27B1 and CYP27A1. Studies on mutants from patients with vitamin D-dependent rickets type I (VDDR-I) and cerebrotendinous xanthomatosis (CTX).

    Science.gov (United States)

    Sawada, N; Sakaki, T; Kitanaka, S; Kato, S; Inouye, K

    2001-12-01

    We have determined eight types of missense mutants of CYP27B1 from Japanese vitamin D-dependent rickets type I (VDDR-I) patients [Kitanaka, S., Takeyama, K., Murayama, A., Sato, T., Okumura, K., Nogami, M., Hasegawa, Y., Niimi, H., Yanagisawa, J., Tanaka, T. & Kato, S. (1998) New England J. Med., 338, 653-661 and Kitanaka, S., Murayama, A., Sakaki, T., Inouye, K., Seino, Y., Fukumoto, S., Shima, M., Yukizane, S., Takayanagi, M., Niimi, H., Takeyama, K. & Kato, S. (1999) J. Clin. Endocrine Metab., 84, 4111-4117]. None of the CYP27B1 mutants showed 1alpha-hydroxylase activity towards 25-hydroxyvitamin D3. Thus, it was assumed that the mutated amino-acid residues play important roles in the 1alpha-hydroxylase activity, such as substrate binding, activation of molecular oxygen, interaction with adrenodoxin, and folding of the cytochrome P450 structure. To examine our hypothesis, we generated various mutants of CYP27B1 and studied their enzymatic properties. In addition, the corresponding mutations were introduced to CYP27A1, which belongs to the same family as CYP27B1. As CYP27A1 showed much higher expression level than CYP27B1 in Escherichia coli, further analysis including heme-binding and substrate-binding was performed with CYP27A1 in place of CYP27B1. Western blot analysis, spectral analysis including reduced CO-difference spectra and substrate-induced difference spectra, and enzymatic analysis of the mutant CYP27A1 gave information on the structure-function relationships of both CYP27A1 and CYP27B1. Although the sequence alignment suggested that Arg107, Gly125, and Pro497 of CYP27B1 might be involved in substrate binding, the experimental data strongly suggested that mutations of these amino-acid residues destroyed the tertiary structure of the substrate-heme pocket. It was also suggested that Arg389 and Arg453 of CYP27B1 were involved in heme-propionate binding, and Asp164 stabilized the four-helix bundle consisting of D, E, I and J helices, possibly by forming

  14. Mutants of downy mildew resistance in Lactuca sativa (lettuce).

    Science.gov (United States)

    Okubara, P A; Anderson, P A; Ochoa, O E; Michelmore, R W

    1994-07-01

    As part of our investigation of disease resistance in lettuce, we generated mutants that have lost resistance to Bremia lactucae, the casual fungus of downy mildew. Using a rapid and reliable screen, we identified 16 distinct mutants of Latuca sativa that have lost activity of one of four different downy mildew resistance genes (Dm). In all mutants, only a single Dm specificity was affected. Genetic analysis indicated that the lesions segregated as single, recessive mutations at the Dm loci. Dm3 was inactivated in nine of the mutants. One of five Dm 1 mutants was selected from a population of untreated seeds and therefore carried a spontaneous mutation. All other Dm1, Dm3, Dm5/8 and Dm7 mutants were derived from gamma- or fast neutron-irradiated seed. In two separate Dm 1 mutants and in each of the eight Dm3 mutants analyzed, at least one closely linked molecular marker was absent. Also, high molecular weight genomic DNA fragments that hybridized to a tightly linked molecular marker in wild type were either missing entirely or were truncated in two of the Dm3 mutants, providing additional evidence that deletions had occurred in these mutants. Absence of mutations at loci epistatic to the Dm genes suggested that such loci were either members of multigene families, were critical for plant survival, or encoded components of duplicated pathways for resistance; alternatively, the genes determining downy mildew resistance might be limited to the Dm loci.

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

    Science.gov (United States)

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

    2015-07-06

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

  16. Neurobehavioral Mutants Identified in an ENU Mutagenesis Project

    Energy Technology Data Exchange (ETDEWEB)

    Cook, Melloni N. [University of Memphis; Dunning, Jonathan P [University of Memphis; Wiley, Ronald G [Vanderbilt University and Veterans Administration, Nashville, TN; Chesler, Elissa J [ORNL; Johnson, Dabney K [ORNL; Goldowitz, Daniel [University of Tennessee Health Science Center, Memphis

    2007-01-01

    We report on a behavioral screening test battery that successfully identified several neurobehavioral mutants among a large-scale ENU-mutagenized mouse population. Large numbers of ENU mutagenized mice were screened for abnormalities in central nervous system function based on abnormal performance in a series of behavior tasks. We developed and employed a high-throughput screen of behavioral tasks to detect behavioral outliers. Twelve mutant pedigrees, representing a broad range of behavioral phenotypes, have been identified. Specifically, we have identified two open field mutants (one displaying hyper-locomotion, the other hypo-locomotion), four tail suspension mutants (all displaying increased immobility), one nociception mutant (displaying abnormal responsiveness to thermal pain), two prepulse inhibition mutants (displaying poor inhibition of the startle response), one anxiety-related mutant (displaying decreased anxiety in the light/dark test), and one learning and memory mutant (displaying reduced response to the conditioned stimulus) These findings highlight the utility of a set of behavioral tasks used in a high throughput screen to identify neurobehavioral mutants. Further analysis (i.e., behavioral and genetic mapping studies) of mutants is in progress with the ultimate goal of identification of novel genes and mouse models relevant to human disorders as well as the identification of novel therapeutic targets.

  17. Structure of the thermolabile mutant aldolase B, A149P: molecular basis of hereditary fructose intolerance.

    Science.gov (United States)

    Malay, Ali D; Allen, Karen N; Tolan, Dean R

    2005-03-18

    Hereditary fructose intolerance (HFI) is a potentially lethal inborn error in metabolism caused by mutations in the aldolase B gene, which is critical for gluconeogenesis and fructose metabolism. The most common mutation, which accounts for 53% of HFI alleles identified worldwide, results in substitution of Pro for Ala at position 149. Structural and functional investigations of human aldolase B with the A149P substitution (AP-aldolase) have shown that the mutation leads to losses in thermal stability, quaternary structure, and activity. X-ray crystallography is used to reveal the structural basis of these perturbations. Crystals of AP-aldolase are grown at two temperatures (4 degrees C and 18 degrees C), and the structure solved to 3.0 angstroms resolution, using the wild-type structure as the phasing model. The structures reveal that the single residue substitution, A149P, causes molecular disorder around the site of mutation (residues 148-159), which is propagated to three adjacent beta-strand and loop regions (residues 110-129, 189-199, 235-242). Disorder in the 110-129-loop region, which comprises one subunit-subunit interface, provides an explanation for the disrupted quaternary structure and thermal instability. Greater structural perturbation, particularly at a Glu189-Arg148 salt bridge in the active-site architecture, is observed in the structure determined at 18 degrees C, which could explain the temperature-dependent loss in activity. The disorder revealed in these structures is far greater than that predicted by homology modeling and underscores the difficulties in predicting perturbations of protein structure and function by homology modeling alone. The AP-aldolase structure reveals the molecular basis of a hereditary disease and represents one of only a few structures known for mutant proteins at the root of the thousands of other inherited disorders.

  18. Plant genetics: increased outcrossing in hothead mutants.

    Science.gov (United States)

    Peng, Peng; Chan, Simon W-L; Shah, Govind A; Jacobsen, Steve E

    2006-09-28

    Arising from: S. J. Lolle, J. L. Victor, J. M. Young & R. E. Pruitt 434, 505-509 (2005); Lolle et al. reply. Lolle et al. report that loss-of-function alleles of the HOTHEAD (HTH) gene in Arabidopsis thaliana are genetically unstable, giving rise to wild-type revertants. On the basis of the reversion of many other genetic markers in hth plants, they suggested a model in which a cache of extragenomic information could cause genes to revert to the genotype of previous generations. In our attempts to reproduce this phenomenon, we discovered that hth mutants show a marked tendency to outcross (unlike wild-type A. thaliana, which is almost exclusively self-fertilizing). Moreover, when hth plants are grown in isolation, their genetic inheritance is completely stable. These results may provide an alternative explanation for the genome wide non-mendelian inheritance reported by Lolle et al.

  19. Google: a narrativa de uma marca mutante

    Directory of Open Access Journals (Sweden)

    Elizete de Azevedo Kreutz

    2010-01-01

    Full Text Available As marcas mutantes já fazem parte de nossa realidade, embora ainda não totalmente percebidas e/ou aceitas como tal. O presente artigo busca refletir sobre a relevância dessas novas estratégias de comunicação e branding, identificando suas principais características. Para isso, utilizamos o método de estudo de caso, o Google, ancorado nos métodos de pesquisa bibliográfica e de internet. A escolha foi intencional, posto que a organização é referência em sua categoria, mecanismo de busca, e reflete essa estratégia comunicacional contemporânea. Como resultado, as informações obtidas nos possibilitam compreender essa tendência de comportamento de marca que busca a interação com seus públicos.

  20. Mutants of Cercospora kikuchii altered in cercosporin synthesis and pathogenicity

    Energy Technology Data Exchange (ETDEWEB)

    Upchurch, R.G.; Walker, D.C.; Rollins, J.A.; Ehrenshaft, M.; Daub, M.E. (North Carolina State Univ., Raleigh (United States))

    1991-10-01

    The authors have obtained spontaneous and UV-induced stable mutants, altered in the synthesis of cercosporin, of the fungal soybean pathogen Cercospora kikuchii. The mutants were isolated on the basis of colony color on minimal medium. The UV-induced mutants accumulated, at most, 2% of wild-type cercosporin levels on all media tested. In contrast, cercosporin accumulation by the spontaneous mutants was strongly medium regulated, occurring only on potato dextrose medium but at concentrations comparable to those produced by the wild-type strain. UV-induced mutants unable to synthesize cercosporin on any medium were unable to incite lesions when inoculated onto the soybean host. Cercosporin was reproducibly isolated from all inoculated leaves showing lesions. Although cercosporin involvement in disease has been indirectly suggested by many previous studies, this is the first report in which mutants blocked in cercosporin synthesis have been used to demonstrate that cercosporin is a crucial pathogenicity factor for this fungal genus.

  1. Mutants of Cercospora kikuchii Altered in Cercosporin Synthesis and Pathogenicity.

    Science.gov (United States)

    Upchurch, R G; Walker, D C; Rollins, J A; Ehrenshaft, M; Daub, M E

    1991-10-01

    We have obtained spontaneous and UV-induced stable mutants, altered in the synthesis of cercosporin, of the fungal soybean pathogen Cercospora kikuchii. The mutants were isolated on the basis of colony color on minimal medium. The UV-induced mutants accumulated, at most, 2% of wild-type cercosporin levels on all media tested. In contrast, cercosporin accumulation by the spontaneous mutants was strongly medium regulated, occurring only on potato dextrose medium but at concentrations comparable to those produced by the wild-type strain. UV-induced mutants unable to synthesize cercosporin on any medium were unable to incite lesions when inoculated onto the soybean host. Cercosporin was reproducibly isolated from all inoculated leaves showing lesions. Although cercosporin involvement in disease has been indirectly suggested by many previous studies, this is the first report in which mutants blocked in cercosporin synthesis have been used to demonstrate that cercosporin is a crucial pathogenicity factor for this fungal genus.

  2. Colony mutants of compatible nocardiae displaying variations in recombining capacity.

    Science.gov (United States)

    Brownell, G H; Walsh, R S

    1972-03-01

    Colonial morphology mutants of Nocardia erythropolis were isolated following ultraviolet (UV) irradiation. The alleles rou-1/smo-1 were located by recombinant analysis and found to be linked to previously mapped characters. On the basis of recombinant class type patterns obtained from various selective characters it was postulated that the rou-1 allele may span a region of unique nucleotides in the Mat-Ce genome. Recombination frequencies of rou-1 and smo-2 bearing mutants of the Mat-Ce mating type were found to differ by over 1000 fold. Attempts to demonstrate that low recombination frequencies produced by the Smo mutants were due to Rec(-) genes were unsuccessful. No increased sensitivity to either UV or X irradiation was observed by the Smo mutants. Acriflavine treatment of either Rou or Smo colony mutants failed to accelerate reversion or to alter the recombining potentials of the mutants.

  3. Screening and identification of mutants of Magnaporthe grisea by REMI

    Institute of Scientific and Technical Information of China (English)

    XIONG Ruyi; LIU Juan; ZHOU Yijun; FAN Yongjian; ZHENG Xiaobo

    2007-01-01

    The plasmid pUCATPH was used to establish a transformation system in wild-type isolate M131 of Magnaporthe grisea.Six hundred and thirty-nine transformants were obtained by restriction enzyme-mediated integration (REMI) with hygromycin B (hyg B) resistance as a tag.Morphological analysis of two of the REMI mutants confirmed that they produced little melanin under black light and continued for three generations.Pathogenicity identification of six mutants screened proved that they made pathogenicity changes on three sets of differential varieties with different resistance genes.Rep-PCR analyses showed that two morphological mutants and two pathogenicity mutants differed from wild-type isolate M131 at the molecular level.RFLP analyses were performed to study the four mutants at the molecular level and the integration sites of the plasmid DNA.The results showed that the plasmid was inserted into all four mutants and that the insertion sites were random.

  4. 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.

  5. Variantes polimórficas Ala513Pro y Gly972Arg del gen IRS-1 no se asocian a la diabetes mellitus tipo 2 en un grupo de la población cubana The Ala513Pro and Gly972ARg polymorphous variants of IRS-1 gen are not associated with type diabetes mellitus in a group of the Cuban population

    Directory of Open Access Journals (Sweden)

    Luis Miguel Pérez

    2011-08-01

    Full Text Available Introducción: la diabetes mellitus tipo 2 es una enfermedad heterogénea y multifactorial, que está determinada por factores genéticos y no genéticos. El sustrato 1 del receptor de la insulina (IRS-1 cumple una función fundamental en la transmisión de la señal insulínica, por tanto sus variantes génicas constituyen blancos importantes en el estudio de la susceptibilidad genética a esta enfermedad en las diferentes poblaciones. Objetivo: explorar el papel de las variantes polimórficas Gly972Arg y Ala513Pro del gen IRS-1 en la susceptibilidad genética de la diabetes mellitus tipo 2 en un grupo de la población cubana. Métodos: se determinó la frecuencia de los polimorfismos Gly972Arg y Ala513Pro del IRS-1 en 499 ciudadanos cubanos, con un índice de masa corporal entre 22-30, con edades comprendidas entre los 40 y 70 años: de ellos 272 (54,5 % diabéticos y 227 (45,5 % no diabéticos. Resultados: la frecuencia del alelo Pro513 fue baja (1,2 % y similar para ambos grupos (1,1 % vs. 1,3 % para el grupo de diabéticos y el grupo control, respectivamente. La frecuencia del polimorfismo Gly972Arg fue de 16,2%, superior a la reportada para la mayoría de las poblaciones estudiadas. No se encontraron diferencias significativas en la frecuencia del alelo Arg972 entre el grupo de diabéticos y el grupo control (15,4 % vs. 17,3 %, ni cambios en los niveles de glucemia e insulinemia asociados a la presencia del alelo polimórfico Arg972. Conclusiones: en este grupo de sujetos de la población cubana, las variantes polimórficas Ala513Pro y Gly972Arg del gen IRS-1 no participan en la etiología de la diabetes mellitus tipo 2.Introduction: the type 2 diabetes mellitus is a heterogeneous and multifactor disease determined by genetic and no-genetic factors. The substrate 1 of insulin receptor (IRS-1 has a fundamental function in transmission of insulin signal, thus its genic variants are significant targets in study of genetic susceptibility to

  6. Preliminary study on a gravity-insensitive rice mutant

    Institute of Scientific and Technical Information of China (English)

    金静; 朱诚; 张红心; 孙宗修

    2004-01-01

    A gravity-insensitive mutant was isolated from rice (Oryza sativa L. Cv. Zhonghua 11) transformed by Agrobacterium tumefaciens. The mutant's shoot growth (prostrate growth) was insensitive to gravity; whereas root growth displayed a normal positive gravitropism.Histological observation of root caps and leaf sheaths indicated that there was no significant difference in the number and size of amyloplasts in cells of the mutant and cells of the wild type

  7. Preliminary study on a gravity-insensitive rice mutant

    Institute of Scientific and Technical Information of China (English)

    金静; 朱诚; 张红心; 孙宗修

    2004-01-01

    A gravity-insensitive mutant was isolated from rice (Oryza sativa L. cv. Zhonghua 11) transformed by Agrobacterium tumefaciens. The mutant's shoot growth (prostrate growth) was insensitive to gravity; whereas root growth displayed a normal positive gravitropism. Histological observation of root caps and leaf sheaths indicated that there was no significant difference in the number and size of amyloplasts in cells of the mutant and cells of the wild type.

  8. [Eremothecium ashbyii mutants resistant to 2,6-diaminopurine].

    Science.gov (United States)

    Stepanov, A I; Beburov, M Iu; Zhdanov, V G

    1975-01-01

    3 groups of Eremothecium ashbyii mutants resistant to 5-10(-3) M 2,6-diaminopurine (DAP) ahve been obtained. The mutants of the 1st group (Dap-r) are selected from the initial susceptible strain by the ability to grow in the presence of 5-10(-3) M DAP. The mutants of the 2nd group (Azg-Dap-r) are selected in the selective background of two analogues of 5-10(-3) M DAP and 10(-4) M 8-azaguanine (AG). The mutants of the 3rd group (Azg-r - DAP-r) are isolated from the mutant Azg-r 34 resistant to 10(-4) M AG. The results of studying cross-resistance of mutants to DAP, AG and 8-azaadenine (AA) show that Dap-r and Azg-Dap-r mutants in contrast to Azg-r - Dap-r, have common phenotypic properties and can grow only on the analogues of adenine. DAP, but not AA, eliminates the inhibitory effect of AG on the growth of these mutants. This effect is probably due to deaminating DAP to guanine. Mutants Azg-r - Dap-r retain the initial resistance to 10(-4) M AG, but are susceptible to higher concentrations of AG and in this case DAP does not eliminate the inhibitory effect of AG. In all mutants obtained the effectiveness of the incorporation of 14C-adenine (but not 14C-guanine) is sharply reduced, thus indicating the absence of adenosine-monophosphate pyrophosphorylase activity. The mutants do not excrete purine-like compounds into the medium. In the course of the continuous growth of mutants in the presence of DAP but not of guanine the red intracellular pigment is formed which seems to be a complex of riboflavin with DAP. A disturbance in the synthesis of adenosine monophosphate pyrophosphorylase does not influence practically the level of the synthesis of riboflavin in E. ashbyii.

  9. Fatty acid biosynthesis in novel ufa mutants of Neurospora crassa.

    Science.gov (United States)

    Goodrich-Tanrikulu, M; Stafford, A E; Lin, J T; Makapugay, M I; Fuller, G; McKeon, T A

    1994-10-01

    New mutants of Neurospora crassa having the ufa phenotype have been isolated. Two of these mutants, like previously identified ufa mutants, require an unsaturated fatty acid for growth and are almost completely blocked in the de novo synthesis of unsaturated fatty acids. The new mutations map to a different chromosomal location than previously characterized ufa mutations. This implies that at least one additional genetic locus controls the synthesis of unsaturated fatty acids in Neurospora.

  10. GABAB R活性水平对致痫大鼠认知功能及Arc/Arg3.1表达的影响%Effects of GABAB receptor expression level on cognitive impairment and Arc/Arg3.1 expression in induced epileptic rats model

    Institute of Scientific and Technical Information of China (English)

    兰彦平; 孙涛; 张春; 袁聪聪; 杨征; 王峰

    2016-01-01

    目的 探讨GABABR活性变化对癫痫大鼠认知功能及Arc/Arg3.1的影响.方法 建立氯化锂-匹罗卡品致痫模型,随机分成正常组、巴氯酚组、CGP组、单纯点燃组.避暗、水迷宫实验观察大鼠认知情况,免疫组化、荧光定量PCR、免疫印迹检测海马组织内GABABR(GB1、GB2)、Arc/Arg3.1蛋白及mRNA表达情况.结果 避暗实验:4组大鼠穿梭次数为:6.8±0.6、1.2土0.2、5.4±0.5及3.6±0.3,潜伏期为:26.1 ±3.9、152.2±12.9、65.8 ±7.0、91.2±9.1,与水迷宫行为学变化趋势一致,显示致痫大鼠认知功能减退,巴氯酚进一步抑制致痫大鼠学习和记忆获取能力,CGP35348可改善致痫大鼠认知功能.Arc/Arg3.1及GB1、GB2相对表达量检测显:致痫大鼠较正常大鼠Arc/Arg3.1及GB1、GB2表达量明显增高,致痫组大鼠相对比,巴氯酚组Arc/Arg3.1表达量下降,GB1、GB2增高;而CGP35348组Arc/Arg3.1表达量增高,GB1、GB2降低.结论 GABABR活性水平可以调控Arc/Arg3.1表达,并影响致痫大鼠认知功能.%Objective To investigate the effects of GABAB receptor on cognitive impairment by using pilocarpine induced kindled rats model and also to check early gene (Arc/Arg3.1) expression.Methods Pilocarpine induced kindled rats were divided into four groups (Group normal,Baclofen,CGP and Kindled) randomly,and every group included 20 rats.We checked their cognitive impairment by using passive avoidance test and water maze test.The expression of GABAB receptor (GB1,GB2) and Arc/Arg3.1 was tested by immunohistochemical staining,RT-PCR and Western blot.Results Passive avoidance test showed four Group rats shuttle times were 6.8 ± 0.6,1.2 ± 0.2,5.4 ± 0.5,3.6 ± 0.3,incubation period were 26.1 ±3.9,152.2 ± 12.9,65.8 ±7.0,91.2 ±9.1,and water maze test had the same trend,with values in epilepsy groups significantly lower than the normal group of rats,which meant cognitive dysfunction.The above results also showed Baclofen further inhibited the learning

  11. Characterization of host-range mutants of cyanophage N-1.

    Science.gov (United States)

    Sarma, T A; Kaur, B

    1997-10-01

    Fifteen host-range (h) mutants of cyanophage N-1 were characterized with reference to their efficiency of plating, time of appearance, morphology and size of plaques on Nostoc muscorum and its three phage-resistant (Nm 1/N-1, Nm 2/N-1 and Nm 8/N-1) mutants. While phage N-1 did not adsorb to the three phage-resistant mutants, the h mutants differed one from the other in having lower or higher adsorption rate constants on N. muscorum or the phage-resistant mutants. The inability of majority of h mutants isolated on Nm 1/N-1 to grow in Nm 8/N-1 was shown to be due to a failure of adsorption. The h mutants also differed one from the other in their reversion (back mutation) frequencies. The lethal doses (LD37) required to kill 37% of free phage particles after UV-irradiation, heating and ethylenediamine tetraacetate (EDTA) treatment greatly varied. Most of the h mutants were found to be considerably more sensitive to UV and thermic inactivation than N-1 while they were resistant to EDTA. The h mutants except five of them were unable to multiply at 40 degrees C. The significance of these features is discussed.

  12. plenty, a novel hypernodulation mutant in Lotus japonicus.

    Science.gov (United States)

    Yoshida, Chie; Funayama-Noguchi, Sachiko; Kawaguchi, Masayoshi

    2010-09-01

    Nitrogen fixation in nodules that contain symbiotic rhizobial bacteria enables legumes to thrive in nitrogen-poor soils. However, this symbiosis is energy consuming. Therefore, legumes strictly control nodulation at both local and systemic levels. Mutants deficient in such controls exhibit a range of phenotypes from non-nodulation to hypernodulation. Here, we isolated a novel hypernodulation mutant from the M(2) progeny derived from Lotus japonicus MG-20 seeds mutagenized by irradiation with a carbon ion beam. We named the mutant 'plenty' because it formed more nodules than the wild-type MG-20. The nodulation zone in the plenty mutant was wider than that in the wild type, but not as enhanced as those in other previously reported hypernodulation mutants such as har1, klv or tml of L. japonicus. Unlike these hypernodulation mutants, the plenty mutant developed nodules of the same size as MG-20. Overall, the plenty mutant exhibited a unique phenotype of moderate hypernodulation. However, a biomass assay indicated that this unique pattern of hypernodulation was a hindrance to host plant growth. The plenty mutant displayed some tolerance to external nitrates and a normal triple response to ethylene. Grafting experiments demonstrated that the root of plenty was responsible for its hypernodulation phenotype. Genetic mapping indicated that the PLENTY gene was located on chromosome 2.

  13. Induction and selection of citrus mutant by gamma-irradiation

    Energy Technology Data Exchange (ETDEWEB)

    Kim, In Jung; Oh, Seung Kyu; Lee, Hyo Yeon [Jeju National University, Jeju (Korea, Republic of)

    2010-09-15

    We have subjected to gamma-irradiation to citrus buds and then grafted onto mature citrus tree. Mutant citrus branch lines have been induced. As a result of first selection, we found the several mutant lines showing interesting phenotypes such as higher sugar content. We have selected several branches showing good qualities such as higher sweetness and/or lower acidity. Some branch lines showed over 13 .deg. Brix sugar content and below 0.9% acidity. Other mutant branch lines showed the changes of shape, size, peel thickness, and fiber contents or distribution of fruits. The results suggest that gamma-irradiation is an effective tool for induction of citrus mutant lines.

  14. Neutron structure of the T26H mutant of T4 phage lysozyme provides insight into the catalytic activity of the mutant enzyme and how it differs from that of wild type.

    Science.gov (United States)

    Hiromoto, Takeshi; Meilleur, Flora; Shimizu, Rumi; Shibazaki, Chie; Adachi, Motoyasu; Tamada, Taro; Kuroki, Ryota

    2017-07-13

    T4 phage lysozyme is an inverting glycoside hydrolase that degrades the murein of bacterial cell walls by cleaving the β-1,4-glycosidic bond. The substitution of the catalytic Thr26 residue to a histidine converts the wild type from an inverting to a retaining enzyme, which implies that the original general acid Glu11 can also act as an acid/base catalyst in the hydrolysis. Here, we have determined the neutron structure of the perdeuterated T26H mutant to clarify the protonation states of Glu11 and the substituted His26, which are key in the retaining reaction. The 2.09-Å resolution structure shows that the imidazole group of His26 is in its singly protonated form in the active site, suggesting that the deprotonated Nɛ2 atom of His26 can attack the anomeric carbon of bound substrate as a nucleophile. The carboxyl group of Glu11 is partially protonated and interacts with the unusual neutral state of the guanidine moiety of Arg145, as well as two heavy water molecules. Considering that one of the water-binding sites has the potential to be occupied by a hydronium ion, the bulk solvent could be the source for the protonation of Glu11. The respective protonation states of Glu11 and His26 are consistent with the bond lengths determined by an unrestrained refinement of the high-resolution X-ray structure of T26H at 1.04-Å resolution. The detail structural information, including the coordinates of the deuterium atoms in the active site, provides insight into the distinctively different catalytic activities of the mutant and wild type enzymes. © 2017 The Authors Protein Science published by Wiley Periodicals, Inc. on behalf of The Protein Society.

  15. A relevance study on uterine leiomyoma and gene polymorphisms of CYP1A1 MspⅠand SULT1A1 Arg213His%CYP1A1基因MspⅠ位点和SULT1A1基因Arg213His位点多态性与子宫肌瘤的关联性研究

    Institute of Scientific and Technical Information of China (English)

    周超; 林林; 张英姿; 徐天和; 张磊磊

    2011-01-01

    目的 探讨细胞色素P450(cytochrome P450,CYP)1A1基因MspⅠ位点和硫酸氨基转移酶(sulfotransferase,SULT)1A1基因Arg213His位点多态性与鲁北地区汉族女性子宫肌瘤的关系.方法 采用聚合酶链式反应-限制性片段长度多态性(PCR-RFLP)方法检测123例子宫肌瘤患者和123例健康对照组的CYP1A1基因MspⅠ位点的基因型和SULT1A1基因Arg213His位点的基因型,分析基因多态性与子宫肌瘤的关系.结果 子宫肌瘤组CYP1A1基因MspⅠ位点的基因型与对照组中的分布比较,差异无统计学意义(P=0.927);而子宫肌瘤组SULT1A1基因Arg213His位点的基因型与对照组中的分布比较,差异有统计学意义(P=0.011).CYP1A1基因MspⅠ位点和SULT1A1基因Arg213His位点多态性在子宫肌瘤的发生过程中的交互作用比较,差异有统计学意义(P=0.024).结论 CYP1A1基因MspⅠ位点多态性与鲁北地区汉族女性子宫肌瘤的易感性无显著相关;SULT1A1基因Arg213His位点多态性与鲁北地区汉族女性子宫肌瘤的发生有关,并增加了子宫肌瘤的患病风险;CYP1A1基因MspⅠ位点和SULT1A1基因Arg213His位点多态性在子宫肌瘤的发生过程中具有交互作用.

  16. Investigation of Antibiotic Resistance Genes (ARGs) in Landfill%垃圾填埋场抗生素抗性基因初探

    Institute of Scientific and Technical Information of China (English)

    李蕾; 徐晶; 赵由才; 宋立岩

    2015-01-01

    Antibiotic resistant genes (ARGs), an emerging contaminant, have been detected worldwide in various environments such as sediments and river. However, little is known about ARGs distribution in landfill. In this study, we investigated five ARGs [sulfonamides resistant genes (sul and sul ), chloramphenicols resistant gene ( cat), β-lactams resistant gene ( bla-SHV), and tetracyclines resistant gene (tetW)] in refuse samples collected from jiangcungou landfill (Xi’an, China) by real-time PCR. We then correlated the ARGs and physiochemical properties of refuse to examine the link between them. Results showed that all tested ARGs have been detected in all samples, suggesting that landfill served as ARGs reservoir. The highest copies numbers of sul , sul , tetW, bla-SHV, and cat were (3. 70 ± 0. 06) × 108 copies•g - 1 (dry refuse), (9. 33 ± 0. 06) × 106 copies•g - 1 (dry refuse), (2. 27 ± 0. 08) × 105 copies•g - 1 ( dry refuse), (3. 68 ± 0. 09) × 104 copies•g - 1 ( dry refuse), and (1. 39 ± 0. 10) × 104 copies•g - 1 ( dry refuse), respectively. Further, sul , sul , and cat positively correlated to moisture and sul and cat negatively correlated to pH.%不同环境介质中抗生素抗性基因普遍存在,但是在垃圾填埋场中抗生素抗性基因尚无相关报道.本实验以西安江村沟垃圾填埋场为研究对象,采集不同方位不同深度垃圾样品,分析垃圾理化性质,用荧光定量 PCR 检测磺胺类抗生素抗性基因(sul 和sul )、抗氯霉素类抗生素抗性基因(cat)、β-内酰胺类抗生素抗性基因(bla-SHV),以及四环素类抗生素抗性基因(tetW)等5种抗生素抗性基因的含量,以相关性分析垃圾理化性质与抗性基因的关联.结果表明,5种抗生素抗性基因均存在于垃圾中,基因拷贝数(以干土计)最大值分别为:(3.70±0.06)×108 copies•g -1( sul )、(9.33±0.06)×106 copies•g -1(sul )、(2.27±0.08)×105 copies•g -1( tetW)、(3.68±0.09)×104 copies

  17. Influence of ADRB2 Gln27Glu and ADRB3 Trp64Arg polymorphisms on body weight and body composition changes after a controlled weight-loss intervention.

    Science.gov (United States)

    Szendrei, Barbara; González-Lamuño, Domingo; Amigo, Teresa; Wang, Guan; Pitsiladis, Yannis; Benito, Pedro J; Gomez-Candela, Carmen; Calderón, Francisco J; Cupeiro, Rocío

    2016-03-01

    The β-2 and β-3 adrenergic receptors (ADRB2 and ADRB3) are thought to play a role in energy expenditure and lipolysis. However, the effects of the ADRB2 glutamine (Gln) 27 glutamic acid (glutamate) (Glu) and ADRB3 tryptophan (Trp) 64 arginine (Arg) polymorphisms on weight loss remain controversial. The aim of this study was to investigate the effect of these polymorphisms on changes in weight and body composition during a controlled weight-loss program. One hundred seventy-three healthy overweight and obese participants (91 women, 82 men) aged 18-50 years participated in a 22-week-long intervention based on a hypocaloric diet and exercise. They were randomly assigned to 1 of 4 groups: strength, endurance, strength and endurance combined, and physical activity recommendations only. Body weight, body mass index (BMI), and body composition variables were assessed before and after the intervention. Genetic analysis was carried out according to standard protocols. No effect of the ADRB2 gene was shown on final weight, BMI, or body composition, although in the supervised male group, Glu27 carriers tended to have greater weight (p = 0.019, 2.5 kg) and BMI (p = 0.019, 0.88 kg/m(2)) reductions than did noncarriers. There seems to be an individual effect of the ADRB3 polymorphism on fat mass (p = 0.004) and fat percentage (p = 0.036), in addition to an interaction with exercise for fat mass (p = 0.038). After the intervention, carriers of the Arg64 allele had a greater fat mass and fat percentage than did noncarriers (p = 0.004, 2.8 kg). In conclusion, the ADRB2 Gln27Glu and ADRB3 Trp64Arg polymorphisms may influence weight loss and body composition, although the current evidence is weak; however, further studies are necessary to clarify their roles.

  18. Algunas observaciones sobre la sinterización del acero austenítico 316L en atmósfera de argón

    Directory of Open Access Journals (Sweden)

    Gómez, F.

    1998-05-01

    Full Text Available PM high speed steels are prone to higher corrosion rates due to residual porosity as well as chromium depletion of the matrix during sintering AISI 316L powders have been cold compacted (100-1,000 MPa and sintered (1,000-1,250°C, 15-240 min under several argon containing or vacuum atmospheres. Better densification rates can be achieved as sintering time or temperature increase in the presence of argon. No surface oxidation has been observed with the use of low pressure argon atmospheres, as long as samples are protected in a stainless steel partially sealed container.

    La utilización de acero inoxidable pulvimetalúrgico está limitada, entre otras razones, por la presencia de porosidad que pueda favorecer procesos corrosivos, así como a posibles pérdidas de cromo en la matriz durante los procesos de sinterización. Se han realizado experiencias de procesado de polvos de acero 316L con distintas presiones de compactación (100-1.000 MPa, temperaturas de sinterización (1.100-1.250°C, tiempos de sinterización (15-240 min y en distintas atmósferas de sinterización, de argón y vacío. Se obtiene una mejor densificación con elevada temperatura y tiempos de sinterización en atmósferas de argón a baja presión. En dicha atmósfera, no se produce oxidación superficial aislando parcialmente las muestras del flujo directo del argón.

  19. A ribosomal misincorporation of Lys for Arg in human triosephosphate isomerase expressed in Escherichia coli gives rise to two protein populations.

    Directory of Open Access Journals (Sweden)

    Beatriz Aguirre

    Full Text Available We previously observed that human homodimeric triosephosphate isomerase (HsTIM expressed in Escherichia coli and purified to apparent homogeneity exhibits two significantly different thermal transitions. A detailed exploration of the phenomenon showed that the preparations contain two proteins; one has the expected theoretical mass, while the mass of the other is 28 Da lower. The two proteins were separated by size exclusion chromatography in 3 M urea. Both proteins correspond to HsTIM as shown by Tandem Mass Spectrometry (LC/ESI-MS/MS. The two proteins were present in nearly equimolar amounts under certain growth conditions. They were catalytically active, but differed in molecular mass, thermostability, susceptibility to urea and proteinase K. An analysis of the nucleotides in the human TIM gene revealed the presence of six codons that are not commonly used in E. coli. We examined if they were related to the formation of the two proteins. We found that expression of the enzyme in a strain that contains extra copies of genes that encode for tRNAs that frequently limit translation of heterologous proteins (Arg, Ile, Leu, as well as silent mutations of two consecutive rare Arg codons (positions 98 and 99, led to the exclusive production of the more stable protein. Further analysis by LC/ESI-MS/MS showed that the 28 Da mass difference is due to the substitution of a Lys for an Arg residue at position 99. Overall, our work shows that two proteins with different biochemical and biophysical properties that coexist in the same cell environment are translated from the same nucleotide sequence frame.

  20. A salt bridge between Arg-20 on parathyroid hormone (PTH) and Asp-137 on the PTH1 receptor is essential for full affinity.

    Science.gov (United States)

    Weaver, Richard E; Wigglesworth, Mark J; Donnelly, Dan

    2014-11-01

    Parathyroid hormone (PTH) acts via the receptor PTH1 and plays an important role in calcium homeostasis. PTH's interaction with the N-terminal domain of PTH1 is mediated in part by Arg-20 on the peptide which forms a number of interactions with the receptor: a charge-charge interaction with Asp-137; hydrogen bonds with the backbone of Asp-29 and Met-32; and hydrophobic interactions with Met-32 and Gln-37. The aim of this work was to establish the importance of the charge-charge interaction through the combined use of modified peptide ligands, site-directed mutations of the receptor, and pharmacological assays. The substitution of Arg-20 with norleucine resulted in a 50-fold reduction in potency at PTH1 and Asp-137-Glu while, in contrast, both Asp-137-Asn and Asp-137-Ala receptors were largely insensitive to this ligand modification. The effect of this removal of the positive charge as position 20 could be partially rescued at PTH1 and Asp-137-Glu, but not Asp-137-Asn and Asp-137-Ala, through a substitution of peptide position 20 with ornithine. The latter two receptors, which have no negative charge at position 137, displayed potency for PTH that was reduced by 40- and 117-fold, respectively. These data demonstrate that a negative charge at residue-137 is important for interacting with ligands containing a positive charge at residue-20, and that the Arg-20 interaction with Asp-137, observed in the crystal structure of the isolated N-terminal domain of PTH1, is likely to be present in the full length receptor where it provides an important affinity- and potency-generating interaction through a salt bridge. Copyright © 2014 Elsevier Inc. All rights reserved.

  1. Mechanistic and Structural Analyses of the Roles of Arg409 and Asp402 in the Reaction of the Flavoprotein Nitroalkane Oxidase

    Energy Technology Data Exchange (ETDEWEB)

    Fitzpatrick,P.; Bozinovski, D.; Heroux, A.; Shaw, P.; Valley, M.; Orville, A.

    2007-01-01

    The flavoprotein nitroalkane oxidase (NAO) catalyzes the oxidation of primary and secondary nitroalkanes to the corresponding aldehydes and ketones. The enzyme is a homologue of acyl-CoA dehydrogenase. Asp402 in NAO has been proposed to be the active site base responsible for removing the substrate proton in the first catalytic step; structurally it corresponds to the glutamate which acts as the base in medium chain acyl-CoA dehydrogenase. In the active site of NAO, the carboxylate of Asp402 forms an ionic interaction with the side chain of Arg409. The R409K enzyme has now been characterized kinetically and structurally. The mutation results in a decrease in the rate constant for proton abstraction of 100-fold. Analysis of the three-dimensional structure of the R409K enzyme, determined by X-ray crystallography to a resolution of 2.65 Angstroms, shows that the critical structural change is an increase in the distance between the carboxylate of Asp402 and the positively charged nitrogen in the side chain of the residue at position 409. The D402E mutation results in a smaller decrease in the rate constant for proton abstraction of 18-fold. The structure of the D402E enzyme, determined at 2.4 Angstroms resolution, shows that there is a smaller increase in the distance between Arg409 and the carboxylate at position 402, and the interaction of this residue with Ser276 is perturbed. These results establish the critical importance of the interaction between Asp402 and Arg409 for proton abstraction by nitroalkane oxidase.

  2. Synthesis and tritium labeling of the highly potent mast cell-degranulating substance P analog H-Arg-Pro-Lys-Pro-NH-C sub 12 H sub 25

    Energy Technology Data Exchange (ETDEWEB)

    Bienert, M.; Oehlke, J.; Niedrich, H. (Academy of Sciences of GDR, Berlin (Germany, F.R.). Inst. of Drug Research); Mittag, E. (Zentralinstitut fuer Kernforschung, Rossendorf (Germany, F.R.))

    1990-12-01

    Tritium labeling of the mast cell degranulating substance P analog H-Arg-Pro-Lys(3,4-{sup 3}H-Pro)-NH-C{sub 12}H{sub 25} by catalytic saturation of the dehydroproline (Dhp{sup 1}) double bond is described. Catalytic tritiation in water afforded the radioactive analog with a specific activity of 1.07 TBq/mmol. Tenfold enhancement of the catalyst-to-substrate ratio resulted in a reduced specific activity of 0.74 TBq/mmol. (author).

  3. Análisis multielemental secuencial de aleaciones mediante plasma de argón generado por acoplamiento inductivo de radiofrecuencia

    OpenAIRE

    Farías de Funes, Silvia

    1996-01-01

    En el presente trabajo se describe la caracterización espectroscópica y analítica de un espectrómetro óptico constituido por el acoplamiento de una fuente de plasma inductivo de argón (IPC) y un monocromador programable de alta resolución, aplicable a la determinación secuencial de Mg, Mn, Cr, Mo en aleaciones de aluminio y de base Cu-Ni. Después de una discusión general sobre las características fundamentales del dispositivo instrumental empleado, se consideran aspectos básicos relacionados ...

  4. Cellular localization of kinin B1 receptor in the spinal cord of streptozotocin-diabetic rats with a fluorescent [Nα-Bodipy]-des-Arg9-bradykinin

    Directory of Open Access Journals (Sweden)

    Gaudreau Pierrette

    2009-03-01

    Full Text Available Abstract Background The kinin B1 receptor (B1R is upregulated by pro-inflammatory cytokines, bacterial endotoxins and hyperglycaemia-induced oxidative stress. In animal models of diabetes, it contributes to pain polyneuropathy. This study aims at defining the cellular localization of B1R in thoracic spinal cord of type 1 diabetic rats by confocal microscopy with the use of a fluorescent agonist, [Nα-Bodipy]-des-Arg9-BK (BdABK and selective antibodies. Methods Diabetes was induced by streptozotocin (STZ; 65 mg/kg, i.p.. Four days post-STZ treatment, B1R expression was confirmed by quantitative real-time PCR and autoradiography. The B1R selectivity of BdABK was determined by assessing its ability to displace B1R [125I]-HPP-desArg10-Hoe140 and B2R [125I]-HPP-Hoe 140 radioligands. The in vivo activity of BdABK was also evaluated on thermal hyperalgesia. Results B1R was increased by 18-fold (mRNA and 2.7-fold (binding sites in the thoracic spinal cord of STZ-treated rats when compared to control. BdABK failed to displace the B2R radioligand but displaced the B1R radioligand (IC50 = 5.3 nM. In comparison, IC50 values of B1R selective antagonist R-715 and B1R agonist des-Arg9-BK were 4.3 nM and 19 nM, respectively. Intraperitoneal BdABK and des-Arg9-BK elicited dose-dependent thermal hyperalgesia in STZ-treated rats but not in control rats. The B1R fluorescent agonist was co-localized with immunomarkers of microglia, astrocytes and sensory C fibers in the spinal cord of STZ-treated rats. Conclusion The induction and up-regulation of B1R in glial and sensory cells of the spinal cord in STZ-diabetic rats reinforce the idea that kinin B1R is an important target for drug development in pain processes.

  5. Convergent evidence of the contribution of TP53 genetic variation (Pro72Arg) to metabolic activity and white matter volume in the frontal lobe in schizophrenia patients.

    Science.gov (United States)

    Molina, Vicente; Papiol, Sergi; Sanz, Javier; Rosa, Araceli; Arias, Bárbara; Fatjó-Vilas, Mar; Calama, Julia; Hernández, Ana I; Bécker, Joemir; Fañanás, Lourdes

    2011-05-01

    Abnormalities in white matter (WM) volumes and integrity in schizophrenia, together with post-mortem studies showing reduced expression of oligodendrocyte/myelination genes and apoptotic processes taking place in oligodendrocytes, suggest the interest of major regulators of apoptosis as candidate genes for some features related to myelin integrity in schizophrenia. Protein p53, encoded by TP53 gene, has a central role in the control of apoptosis and is involved in oligodendrocyte development. TP53 gene polymorphisms may account for variability in WM features, metabolic activity and biochemical markers of neuronal integrity and membrane turnover. Pro72Arg and Ins16bp polymorphisms at TP53 gene were analyzed in 20 DSM-IV schizophrenia patients. T1/T2-weighted sequences of these patients were acquired using a 1.5T Philips Gyroscan system. Scans were transformed into Talairach space and segmented into gray matter (GM), WM and cerebrospinal fluid (CSF) using Statistical Parametric Mapping under a ROI approach. Likewise dorsolateral prefrontal cortex (DLPFC) metabolic activity was measured using a procedure based on MRI/PET image fusion. In 13 of these patients proton magnetic resonance spectroscopy was used to examine N-acetylaspartate (NAA), creatine (Cr) and choline (Cho) levels in dorsolateral-medial prefrontal cortex (DLMPFC). MRI data were adjusted for age and brain volume using regression parameters from a healthy control group (n=45). Patients Pro/Arg heterozygous (Pro72Arg polymorphism) showed a generalized deficit in whole-brain WM that was especially prominent in frontal lobe and a lower metabolic activity in the DLPFC as compared to Pro/Pro homozygous. Pro/Arg subjects also showed decreased NAA/Cho and increased Cho/Cr ratios in right DLMPFC. TP53 genetic variability influences WM volumes in frontal lobes and it seems to modulate the metabolic activity in this region. Our results suggest that TP53 might influence aspects of myelin and white matter integrity

  6. Association of Gln27Glu and Arg16Gly polymorphisms in Beta2-adrenergic receptor gene with obesity susceptibility: a meta-analysis.

    Directory of Open Access Journals (Sweden)

    Hongxiu Zhang

    Full Text Available BACKGROUND: The beta2-adrenergic receptor (ADRB2 gene polymorphism has been implicated in susceptibility to obesity, but study results are still controversial. OBJECTIVE: The present meta-analysis is performed to determine whether there are any associations between the Gln27Glu (rs1042714 or the Arg16Gly (rs1042713 polymorphisms in ADRB2 and obesity susceptibility. METHODS: The PubMed (1950-2014, Embase (1974-2014, and China National Knowledge Infrastructure (CNKI, 1994-2014 databases were searched using the search terms ("Beta2-adrenergic receptor", "β2-adrenergic receptor" or "ADRB2", "polymorphism," and "obesity". Fixed- or random-effects pooled measures were determined on the bias of heterogeneity tests across studies. Publication bias was examined by Egger's test and the modified Begg's test. RESULTS: Eighteen published articles were selected for meta-analysis. Overall analyses showed that rs1042714 (Gln27Glu was associated with significantly increased obesity risk in the heterozygote model (Gln/Glu vs. Gln/Gln: OR: 1.16, 95% CI: 1.04-1.30, I2 = 49%, P = 0.009 and the dominant model (Gln/Glu + Glu/Glu vs. Gln/Gln: OR: 1.2, 95% CI: 1.00-1.44, I2 = 55%, P = 0.04, whereas no significant association was found in the other models for rs1042714. Also, no significant association was found between the rs1042713 (Arg16Gly gene polymorphism and the risk of obesity in all genetic models. In addition, neither rs1042713 (Arg16Gly nor rs1042714 (Gln27Glu showed any significant association with obesity susceptibility when the population were stratified based on gender. CONCLUSION: Our meta-analysis revealed that the rs1042714 (Gln27Glu polymorphism is associated with obesity susceptibility. However, our results do not support an association between rs1042713 (Arg16Gly polymorphisms and obesity in the populations investigated. This conclusion warrants confirmation by more case-control and cohort studies.

  7. Integration of regulatory signals through involvement of multiple global regulators: control of the Escherichia coli gltBDF operon by Lrp, IHF, Crp, and ArgR

    Directory of Open Access Journals (Sweden)

    Mishra Pankaj K

    2007-01-01

    Full Text Available Abstract Background The glutamate synthase operon (gltBDF contributes to one of the two main pathways of ammonia assimilation in Escherichia coli. Of the seven most-global regulators, together affecting expression of about half of all E. coli genes, two were previously shown to exert direct, positive control on gltBDF transcription: Lrp and IHF. The involvement of Lrp is unusual in two respects: first, it is insensitive to the usual coregulator leucine, and second, Lrp binds more than 150 bp upstream of the transcription starting point. There was indirect evidence for involvement of a third global regulator, Crp. Given the physiological importance of gltBDF, and the potential opportunity to learn about integration of global regulatory signals, a combination of in vivo and in vitro approaches was used to investigate the involvement of additional regulatory proteins, and to determine their relative binding positions and potential interactions with one another and with RNA polymerase (RNAP. Results Crp and a more local regulator, ArgR, directly control gltBDF transcription, both acting negatively. Crp-cAMP binds a sequence centered at -65.5 relative to the transcript start. Mutation of conserved nucleotides in the Crp binding site abolishes the Crp-dependent repression. ArgR also binds to the gltBDF promoter region, upstream of the Lrp binding sites, and decreases transcription. RNAP only yields a defined DNAse I footprint under two tested conditions: in the presence of both Lrp and IHF, or in the presence of Crp-cAMP. The DNAse I footprint of RNAP in the presence of Lrp and IHF is altered by ArgR. Conclusion The involvement of nearly half of E. coli's most-global regulatory proteins in the control of gltBDF transcription is striking, but seems consistent with the central metabolic role of this operon. Determining the mechanisms of activation and repression for gltBDF was beyond the scope of this study. However the results are consistent with a

  8. Pathogenic Variant in ACTB, p.Arg183Trp, Causes Juvenile-Onset Dystonia, Hearing Loss, and Developmental Delay without Midline Malformation

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    Erin Conboy

    2017-01-01

    Full Text Available ACTB encodes the β-actin, and pathogenic variations in this gene have typically been associated with Baraitser-Winter cerebrofrontofacial syndrome, a congenital malformation syndrome characterized by short stature, craniofacial anomalies, and cerebral anomalies. Here, we describe the third case with the p.Arg183Trp variant in ACTB causing juvenile-onset dystonia. Our patient has severe, intractable dystonia, developmental delay, and sensorineural hearing loss, besides hyperintensities in the caudate nuclei and putamen on the brain MRI, which is a distinct but overlapping phenotype with the previously reported case of identical twins with the same alteration in ACTB.

  9. Synthesis of the C-terminal octapeptide of pig oxyntomodulin. Lys-Arg-Asn-Lys-Asn-Asn-Ile-Ala: a potent inhibitor of pentagastrin-induced acid secretion.

    Science.gov (United States)

    Audousset-Puech, M P; Jarrousse, C; Dubrasquet, M; Aumelas, A; Castro, B; Bataille, D; Martinez, J

    1985-10-01

    The synthesis of Lys-Arg-Asn-Lys-Asn-Asn-Ile-Ala representing the C-terminal octapeptide of oxyntomodulin isolated from pig intestine is described. Its structure was confirmed by its 360-MHz 1H NMR spectra. The octapeptide was tested for its ability to inhibit pentagastrin-induced acid secretion, in the anaesthetized rat, in the conscious rat with chronic gastric fistula, and in the conscious cat with gastric chronic fistula. The octapeptide inhibits pentagastrin-induced acid secretion in all three models. Compared to oxyntomodulin, the parent hormone, the synthetic peptide was approximately 150 times less potent but has the same efficacy. Biological data are presented and discussed.

  10. High-resolution X-ray structure of the unexpectedly stable dimer of the [Lys(-2)-Arg(-1)-des(17-21)]endothelin-1 peptide.

    Science.gov (United States)

    Hoh, François; Cerdan, Rachel; Kaas, Quentin; Nishi, Yoshinori; Chiche, Laurent; Kubo, Shigeru; Chino, Naoyoshi; Kobayashi, Yuji; Dumas, Christian; Aumelas, André

    2004-12-07

    Previous structural studies on the [Lys((-2))-Arg((-1))]endothelin-1 peptide (KR-ET-1), 540-fold less potent than ET-1, strongly suggested the presence of an intramolecular Arg(-1)-Asp(8) (R(-1)-D(8)) salt bridge that was also observed in the shorter [Lys((-2))-Arg((-1))-des(17-21)]endothelin-1 derivative (KR-CSH-ET). In addition, for these two analogues, we have shown that the Lys-Arg dipeptide, which belongs to the prosequence, significantly improves the formation of the native disulfide bonds (>or=96% instead of approximately 70% for ET-1). In contrast to what was inferred from NMR data, molecular dynamics simulations suggested that such an intramolecular salt bridge would be unstable. The KR-CSH-ET peptide has now been crystallized at pH 5.0 and its high-resolution structure determined ab initio at 1.13 A using direct methods. Unexpectedly, KR-CSH-ET was shown to be a head-to-tail symmetric dimer, and the overall interface involves two intermolecular R(-1)-D(8) salt bridges, a two-stranded antiparallel beta-sheet, and hydrophobic contacts. Molecular dynamics simulations carried out on this dimer clearly showed that the two intermolecular salt bridges were in this case very stable. Sedimentation equilibrium experiments unambiguously confirmed that KR-ET-1 and KR-CSH-ET also exist as dimers in solution at pH 5.0. On the basis of the new dimeric structure, previous NMR data were reinterpreted. Structure calculations were performed using 484 intramolecular and 38 intermolecular NMR-derived constraints. The solution and the X-ray structures of the dimer are very similar (mean rmsd of 0.85 A). Since the KR dipeptide at the N-terminus of KR-CSH-ET is present in the prosequence, it can be hypothesized that similar intermolecular salt bridges could be involved in the in vivo formation of the native disulfide bonds of ET-1. Therefore, it appears to be likely that the prosequence does assist the ET-1 folding in a chaperone-like manner before successive cleavages that

  11. Chemotyping of yeast mutants using robotics.

    Science.gov (United States)

    Rieger, K J; El-Alama, M; Stein, G; Bradshaw, C; Slonimski, P P; Maundrell, K

    1999-07-01

    By now, the EUROFAN programme for the functional analysis of genes from the yeast genome has attained its cruising speed. Indeed, several hundreds of yeast mutants with no phenotype as tested by growth on standard media and no significant sequence similarity to proteins of known function are available through the efforts of various laboratories. Based on the methodology initiated during the pilot project on yeast chromosome III (Yeast 13, 1547-1562, 1997) we adapted it to High Throughput Screening (HTS), using robotics. The first 100 different gene deletions from EUROSCARF, constructed in an FY1679 strain background, were run against a collection of about 300 inhibitors. Many of these inhibitors have not been reported until now to interfere in vivo with growth of Saccharomyces cerevisiae. In the present paper we provide a list of novel growth conditions and a compilation of 49 yeast deletants (from chromosomes II, IV, VII, X, XIV, XV) corresponding to 58% of the analysed genes, with at least one clear and stringent phenotype. The majority of these deletants are sensitive to one or two compounds (monotropic phenotype) while a distinct subclass of deletants displays a hyper-pleiotropic phenotype with sensitivities to a dozen or more compounds. Therefore, chemotyping of unknown genes with a large spectrum of drugs opens new vistas for a more in-depth functional analysis and a more precise definition of molecular targets.

  12. Pharmacological correctors of mutant CFTR mistrafficking

    Directory of Open Access Journals (Sweden)

    Nicoletta ePedemonte

    2012-10-01

    Full Text Available The lack of phenylalanine 508 (∆F508 mutation in the CFTR Cl- channel represents the most frequent cause of cystic fibrosis (CF, a genetic disease affecting multiple organs such lung, pancreas, and liver. ∆F508 causes instability and misfolding of CFTR protein leading to early degradation in the endoplasmic reticulum and accelerated removal from the plasma membrane. Pharmacological correctors of mutant CFTR protein have been identified by high-throughput screening of large chemical libraries, by in silico docking of virtual compounds on CFTR structure models, or by using compounds that affect the whole proteome (e.g. histone deacetylase inhibitors or a single CFTR-interacting protein. The presence of multiple defects caused at the CFTR protein level by ∆F508 mutation and the redundancy of quality control mechanisms detecting ∆F508-CFTR as a defective protein impose a ceiling to the maximal effect that a single compound (corrector may obtain. Therefore, treatment of patients with the most frequent CF mutation may require the optimized combination of two drugs having additive or synergic effects.

  13. New types of Escherichia coli recombination-deficient mutants.

    Science.gov (United States)

    Freifelder, D

    1976-11-01

    A set of Escherichia coli mutants deficient in intramolecular recombination and different from those previously found is described. All have temperature-sensitive lethal mutations. The mutants have been characterized with respect to the following properties: the Pap phenotype, deoxyribonucleic acid synthesis, sensitivity to ultraviolet light, ability to support the growth of phage lambda, filament formation, and mutation frequency.

  14. Characterization of Gibberellin Receptor Mutants of Barley (Hordeum vulgare L.)

    Institute of Scientific and Technical Information of China (English)

    Peter M.Chandler; Carol A.Harding; Anthony R.Ashton; Mark D.Mulcair; Nicholas E.Dixon; Lewis N.Mander

    2008-01-01

    The sequence of Gidl (a gene for a gibberellin (GA) receptor from rice) was used to identify a putative orthoIogue from barley.This was expressed in E.coil,and produced a protein that was able to bind GA in vitro with both structural specificity and saturability.Its potential role in GA responses was investigated using barley mutants with reduced GA sensitivity (gsel mutants).Sixteen different gsel mutants each carried a unique nucleotide substitution in this sequence.In all but one case,these changes resulted in single amino acid substitutions,and,for the remaining mutant,a substitution in the 5' untranslated region of the mRNA is proposed to interfere with translation initiation.There was perfect linkage in segregating populations between new mutant alleles and the gsel phenotype,leading to the conclusion that the putative GID1 GA receptor sequence in barley corresponds to the Gsel locus.Determination of endogenous GA contents in one of the mutants revealed enhanced accumulation of bioactive GA1,and a deficit of C20 GA precursors.All of the gsel mutants had reduced sensitivity to exogenous GA3,and to AC94377 (a GA analogue) at concentrations that are normally 'saturating',but,at much higher concentrations,there was often a considerable response.The comparison between barley and rice mutants reveals interesting differences between these two cereal species in GA hormonal physiology.

  15. Mutants of Pseudomonas putida affected in poly-3-hydroxyalkanoate synthesis

    NARCIS (Netherlands)

    Ren, Q; Kessler, B; van der Leij, F; Witholt, B.

    1998-01-01

    The generation and characterization of Pseudomonas putida KT2442 mutants affected in poly-3-hydroxyalkanoate (PHA) synthesis are reported. The mutants from P. putida KT2442 carrying several copies of the PHA-polymerase-encoding gene (phaC) were isolated via N-methyl-N'-nitro-N-nitrosoguanidine chemi

  16. A Mutant Hunt Using the C-Fern (Ceratopteris Richardii)

    Science.gov (United States)

    Calie, Patrick J.

    2005-01-01

    A modification of the popular C-Fern system, the tropical fern Ceratopteris richardii is developed in which students plate out a genetically mixed set of fern spores and then select for specific mutants. This exercise can provide students with an experience in plant mutant selection and can be used as a platform to expose students to a diverse…

  17. Characterization of peroxisome-deficient mutants of Hansenula polymorpha

    NARCIS (Netherlands)

    Tan, Xuqiu; Titorenko, Vladimir I.; Klei, Ida J. van der; Sulter, Grietje J.; Haima, Peter; Waterham, Hans R.; Evers, Melchior; Harder, Willem; Veenhuis, Marten; Cregg, James M.

    1995-01-01

    In the methylotrophic yeast Hansenula polymorpha, approximately 25% of all methanol-utilization-defective (Mut(-)) mutants are affected in genes required for peroxisome biogenesis (PER genes). Previously, we reported that one group of pel mutants, termed Pim(-), are characterized by the presence of

  18. Absence of Pneumocystis dihydropteroate synthase mutants in Brittany, France.

    Science.gov (United States)

    Le Gal, Solène; Robert-Gangneux, Florence; Perrot, Maëla; Rouillé, Amélie; Virmaux, Michèle; Damiani, Céline; Totet, Anne; Gangneux, Jean-Pierre; Nevez, Gilles

    2013-05-01

    Archival Pneumocystis jirovecii specimens from 84 patients monitored at Rennes University Hospital (Rennes, France) were assayed at the dihydropteroate synthase (DHPS) locus. No patient was infected with mutants. The results provide additional data showing that P. jirovecii infections involving DHPS mutants do not represent a public health issue in Brittany, western France.

  19. Poliovirus Mutants Resistant to Neutralization with Soluble Cell Receptors

    Science.gov (United States)

    Kaplan, Gerardo; Peters, David; Racaniello, Vincent R.

    1990-12-01

    Poliovirus mutants resistant to neutralization with soluble cellular receptor were isolated. Replication of soluble receptor-resistant (srr) mutants was blocked by a monoclonal antibody directed against the HeLa cell receptor for poliovirus, indicating that the mutants use this receptor to enter cells. The srr mutants showed reduced binding to HeLa cells and cell membranes. However, the reduced binding phenotype did not have a major impact on viral replication, as judged by plaque size and one-step growth curves. These results suggest that the use of soluble receptors as antiviral agents could lead to the selection of neutralization-resistant mutants that are able to bind cell surface receptors, replicate, and cause disease.

  20. Determinants of lipoprotein(a) assembly: a study of wild-type and mutant apolipoprotein(a) phenotypes isolated from human and rhesus monkey lipoprotein(a) under mild reductive conditions.

    Science.gov (United States)

    Edelstein, C; Mandala, M; Pfaffinger, D; Scanu, A M

    1995-12-19

    We previously observed that rhesus monkey lipoprotein(a) [Lp(a)], is lysine-binding defective (Lys-) and attributed this deficiency to the presence of Arg72 in the lysine-binding site (LBS) of kringle IV-10 of apolipoprotein(a) [apo(a)] [Scanu, A.M., Miles, L.A., Fless, G.M., Pfaffinger, D., Eisenbart, J., Jackson, E., Hoover-Plow, J.L., Brunck, T., & Plow, E.F. (1993) J. Clin. Invest. 91, 283-291]. We also identified human mutants having Arg72 instead of Trp72 (wild type) in the LBS of kringle IV-10 [Scanu, A M., Pfaffinger, D., lEE, J.C., & Hinman, J. (1994) Biochim. Biophys. Acta 1227, 41-45]. Unique to the human mutant phenotype were the very low levels of plasma Lp(a), suggesting structural differences between human and rhesus apo(a) and a possible divergent mode of Lp(a) assembly. In order to explore the possibility of a relationship between apo(a) LBS and Lp(a) assembly, we developed a novel method for isolating wild-type and mutant apo(a) phenotypes in a free form by subjecting each parent Lp(a) to mild reductive conditions using 2 mM dithioerythritol (DTE) and 100 mM of the lysine analogue, epsilon-aminocaproic acid (EACA). The application of this method to the study of wild-type and mutant apo(a) species showed that regardless of the source of Lp(a), i.e., positive lysine binding (Lys+) or negative lysine binding (Lys-), all of the isolated free apo(a)s were Lys+. Moreover, incubation of free apo(a)s with their autologous human or rhesus low-density lipoproteins (LDL) generated Lp(a) complexes which were structurally and functionally indistinguishable from their parent native Lp(a). In each instance, the reassembly process was inhibited by the presence of either EACA or proline. These two reagents had a minimal effect on either Lp(a) or reassembled Lp(a) [RLp(a)]. Free apo(a) bound to apoB100 of very low density lipoproteins (VLDL) to form a triglyceride-rich Lp(a). These results show that (1) both human and rhesus Lp(a) are amenable to dissassembly and

  1. Phenotypic Characterization of a Female Sterile Mutant in Rice

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    A female sterile mutant, derived from a spontaneous mutation, wasfirst discovered in rice (Oryza sativa L. ssp.indica) restorer line 202R. With normal flowering, the mutant exhibits an extremely Iow seed-setting rate. When the mutant is crossed as a pollen donor, the seeds set normally; whereas when it is used as a pollen receiver,no seeds are obtained even with mixed pollen grains of different varieties sprinkled over the stigmas. The floret of the mutant, consisting of six stamens and one pistil, looks the same as that of the wild type in the malefemale organs, except that less than 10% of the mutant florets have three stigmas on the ovary. Although the mutant has a low seed-setting rate, Its pollen fertility is approximately 87.1%, which is equal to that of the wild type. In addition, more than 90% of the mature embryo sacs of the mutant have complete inner structures. At every stage after pollination, the sperm, embryo, and endosperm are not found in the mutant embryo sac,whereas the disintegration of the egg cell that does not accomplish fertilization is visible. Through observations with a fluorescence microscope, we have found that the pollen grains germinate normally, whereas the pollen tube abnormally elongates in the style-transmitting tissue. The mutant pollen tubes display various defects in the style, such as slower elongation, conversed elongation, distorted elongation, swollen tips, or branched tips. As a result, the growth of the pollen tubes ceases in the style, and, therefore, the pollen tubes cannot reach the embryo sac and the process of double fertilization is blocked. Based on these observations,we conclude that this mutant, designated as fs-202R, is a novel type of female sterile mutation in rice, which causes the arrest of the elongation of the pollen tube.

  2. Isolation and Characterization of mAMSA-hypersensitive Mutants

    Science.gov (United States)

    Rogojina, Anna T.; Nitiss, John L.

    2008-01-01

    Topoisomerase II (Top2) is the primary target for active anti-cancer agents. We developed an efficient approach for identifying hypersensitive Top2 mutants and isolated a panel of mutants in yeast Top2 conferring hypersensitivity to the intercalator N-[4-(9-acridinylamino)-3-methoxyphenyl]methanesulphonanilide (mAMSA). Some mutants conferred hypersensitivity to etoposide as well as mAMSA, whereas other mutants exhibited hypersensitivity only to mAMSA. Two mutants in Top2, changing Pro473 to Leu and Gly737 to Val, conferred extraordinary hypersensitivity to mAMSA and were chosen for further characterization. The mutant proteins were purified, and their biochemical activities were assessed. Both mutants encode enzymes that are hypersensitive to inhibition by mAMSA and other intercalating agents and exhibited elevated levels of mAMSA-induced Top2:DNA covalent complexes. While Gly737 → Val Top2p generated elevated levels of Top2-mediated double strand breaks in vitro, the Pro473 → Leu mutant protein showed only a modest increase in Top2-mediated double strand breaks but much higher levels of Top2-mediated single strand breaks. In addition, the Pro473 → Leu mutant protein also generated high levels of mAMSA-stabilized covalent complexes in the absence of ATP. We tested the role of single strand cleavage in cell killing with alleles of Top2 that could generate single strand breaks, but not double strand breaks. Expression in yeast of a Pro473 → Leu mutant that could only generate single strand breaks conferred hypersensitivity to mAMSA. These results indicate that generation of single strand breaks by Top2-targeting agents can be an important component of cell killing by Top2-targeting drugs. PMID:18723844

  3. Ophthalmic findings in a family with early-onset isolated ectopia lentis and the p.Arg62Cys mutation of the fibrillin-1 gene (FBN1).

    Science.gov (United States)

    Zhao, Jun-Hong; Jin, Tian-Bo; Liu, Qing-Bo; Chen, Chao; Hu, Hai-Tao

    2013-01-01

    The purpose of this paper is to describe ophthalmic findings in a family with isolated ectopia lentis (EL) caused by a specific FBN1 mutation. Detailed family histories and clinical data were recorded for six isolated EL patients of 11 family members. The ophthalmological and systematic examinations were performed on patients and unaffected members of the investigated family. The detailed ocular examinations included visual acuity, anterior chamber depth, pupil size, lens location, optometry, central corneal thickness, keratometry, slitlamp examination, fundus examination, axial length, ocular B-ultrasound, gonioscope checking, ultrasound biomicroscopy (UBM) and intraocular pressure (IOP; Goldmann applanation tonometer). Systematic examinations included the measurement of echocardiogram, height, arm span, skull, face, jaw, tooth, breast bone, spinal column, and skin. Genomic DNA was extracted using the phenol-chloroform extraction method for all subjects, and sequencing was carried out on an ABI Prism 3730 Genetic Analyzer. A heterozygous mutation, c.184C>T (p.Arg62Cys) in exon 2 of FBN1 was identified in all affected members but was not found in any unaffected member of the family. Our study presented detailed clinical manifestations, including some novel ophthalmic findings, such as pupillary abnormality, different types of glaucoma, and progressive hyperopia. Ophthalmic findings and the p.Arg62Cys mutation of FBN1 gene were reported in a family with early-onset isolated ectopia lentis.

  4. ARGE Weser (Working Group for Water Pollution Abatement in the Weser River). Weser water quality report 1996; Arbeitsgemeinschaft zur Reinhaltung der Weser. Weserguetebericht 1996

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    1997-08-01

    The ARGE Weser (Arbeitsgemeinschaft zur Reinhaltung der Weser), which comprises representatives of the German states of Bremen, Hessen, Niedersachsen, Nordrhein-Westfalen and Thueringen, publishes a yearly water quality report based on the measuring programmes for the Weser river and for the Werra/Ulster rivers (the latter is a joint project of the states of Hessen and Thrueringen). The Weser river today has a water quality of II-III after the successful implementation of the 1989 action programme. Chloride concentrations were reduced as well, but efforts will still be made by communal authorities, agriculture, industry and administrative bodies if the goals set for 2000 are to be reached. (orig/AJ) [Deutsch] Die Arbeitsgemeinschaft zur Reinhaltung der Weser (ARGE Weser), der die Bundeslaender Bremen, Hessen, Niedersachsen, Nordrhein-Westfalen und Thueringen angehoeren, dokumentiert jaehrlich im Weserguetebericht die Ergebnisse des Weser- und des von Hessen und Thueringen gemeinsam gefuehrten Messprogrammes Werra/Ulster. Die Weser weist heute ueberwiegend die Gewaesserguete II-III auf. Dies ist ein Ergebnis des 1989 beschlossenen Aktionsprogrammes. Deutliche Erfolge konnten auch im Hinblick auf die Reduzierung der Chloridbelastung erzielt werden. Die volle Verwirklichung des Aktionsprogrammes Weser bis zum Jahr 2000 wird allerdings noch erhebliche Anstrengungen auf Seiten der Kommunen, der Landwirtschaft, der Industrie und der Verwaltungen erfordern. (orig./AJ)

  5. Un arma no solo de prestigio: la espada argárica de Peñalosa (Baños de la Encina, Jaén

    Directory of Open Access Journals (Sweden)

    Moreno Onorato, Auxilio

    2015-12-01

    Full Text Available The sword found in a house at the Bronze Age settlement of Peñalosa (Baños de la Encina, province of Jaén is the only example from a domestic context in the El Argar culture. We analize the circumstances of the discovery, and discuss the results of the various analyses that have been performed. Finally, we compare the Peñalosa find to other contemporaneous swords from the Iberian Peninsula and discuss whether the role of Argaric weapons was symbolic or functional.Se estudia una espada singular de la cultura argárica tanto por su tipología, estado de conservación (hoja, piezas de la empuñadura y metal (bronce y plata como por proceder de un contexto doméstico del poblado de la Edad del Bronce de Peñalosa (Baños de la Encina, Jaén. Se analizan las circunstancias del hallazgo, presentándose para su discusión los resultados de los análisis FRX y ICP-SFMS realizados. Se compara con otras espadas contemporáneas aparecidas en la Península Ibérica y se debate el papel simbólico o funcional de las armas argáricas.

  6. Hippocampal Arc (Arg3.1) expression is induced by memory recall and required for memory reconsolidation in trace fear conditioning.

    Science.gov (United States)

    Chia, Chester; Otto, Tim

    2013-11-01

    Mounting evidence suggests that long-lasting, protein synthesis-dependent changes in synaptic strength accompany both the initial acquisition and subsequent recall of specific memories. Within brain areas thought to be important for learning and memory, including the hippocampus, learning-related plasticity is likely mediated in part by NMDA receptor activation and experience-dependent changes in gene expression. In the present study, we examined the role of activity-regulated cytoskeletal-associated protein (Arc/Arg3.1) expression in the acquisition, recall, and reconsolidation of memory in a trace fear conditioning paradigm. First, we show that the expression of Arc protein in ventral hippocampus (VH) is dramatically enhanced by memory recall 24h after the acquisition of trace fear conditioning, and that both memory recall and the associated recall-induced enhancement of Arc expression are blocked by pre-training administration of 2-amino-5-phosphonovaleric acid (APV). Next, we show that while infusion of Arc antisense oligodeoxynucleotides (ODNs) into VH prior to testing had little effect on memory recall, it significantly reduced both Arc protein expression and freezing behavior during subsequent testing sessions. Collectively, these results suggest that Arc/Arg3.1 protein plays an important functional role in both the initial acquisition of hippocampal-dependent memory and the reconsolidation of these memories after recall. Copyright © 2013 Elsevier Inc. All rights reserved.

  7. Change in microbial community in landfill refuse contaminated with antibiotics facilitates denitrification more than the increase in ARG over long-term

    Science.gov (United States)

    Wu, Dong; Chen, Guanzhou; Zhang, Xiaojun; Yang, Kai; Xie, Bing

    2017-01-01

    In this study, the addition of sulfamethazine (SMT) to landfill refuse decreased nitrogen intermediates (e.g. N2O and NO) and dinitrogen (N2) gas fluxes to antibiotic resistance gene) levels in the refuse increased tenfold after long-term exposure to antibiotics, followed by a fourfold increase in the N2 flux, but SMT-amended samples with the largest resistome facilitated the denitrification (the nitrogen accumulated as NO gas at ~6 μg-N/kg-refuse·h‑1) to a lesser extent than OTC-amended samples. Further, deep sequencing results show that long-term OTC exposure partially substituted Hyphomicrobium, Fulvivirga, and Caldilinea (>5%) for the dominant bacterial hosts (Rhodothermus, ~20%) harboring nosZ and norB genes that significantly correlated with nitrogen emission pattern, while sulfamethazine amendment completely reduced the relative abundance of the “original inhabitants” functioning to produce NOx gas reduction. The main ARG carriers (Pseudomonas) that were substantially enriched in the SMT group had lower levels of denitrifying functional genes, which could imply that denitrification is influenced more by bacterial dynamics than by abundance of ARGs under antibiotic pressures.

  8. Leptin Receptor Gene Gln223Arg Polymorphism Is Not Associated with Hypertension: A Preliminary Population-Based Cross-Sectional Study

    Directory of Open Access Journals (Sweden)

    Geórgia das Graças Pena

    2014-01-01

    Full Text Available Hypertension is responsible for high morbidity and mortality as one of the most important cardiometabolic risk factors. The aim of the study was to investigate whether the Gln223Arg in the leptin receptor (LEPR influences the prevalence of hypertension. A cross-sectional study was carried out in individuals aged ≥ 18 years. Polymorphism identification was performed using PCR-RFLP analysis. Participants with blood pressure ≥ 140/90 mmHg or medication use were considered hypertensive. Frequencies, means, cross-tabulations, and multivariate models were produced to study differences in hypertension prevalence by genotypes. The study includes 470 participants. The frequency of GG polymorphism variant was 10.43%, 46.81% AG, and 42.77% AA. The distribution of hypertension frequency by LEPR genotypes was the following: AA 43.8%, AG 40.4%, and GG 40.8%; there were no significant differences between groups. Comparative analysis which used multivariate Poisson regression adjusted by many potential confounders (age, sex, schooling, smoking, alcohol intake, obesity, and family history of parental obesity did not modify this result. In this large sample of population-based study, the association of the LEPR Gln223Arg gene polymorphism with hypertension was not observed.

  9. TÜRKİYE’DE AR-GE & İNOVASYON HARCAMALARININ GELİŞİM SÜRECİ VE EKONOMİK ETKİLERİ

    Directory of Open Access Journals (Sweden)

    Murat DEMİR

    2014-10-01

    Full Text Available Bu çalışmanın amacı, özellikle son yıllarda gittikçe önem kazanan İnovasyon kavramını açıklamak; bilim, teknoloji ve inovasyon faaliyetleriyle önemli başarılar elde etmiş Doğu Asya ülkelerini ve başarılarını değerlendirmek ve de bu faaliyetlerin Türkiye’deki gelişim sürecini incelemektir. Çalışmanın bir diğer amacı da Türkiye’de Ar-Ge & inovasyon yatırımlarının doğal sonucu olan patent başvuru rakamlarını ve patentlemedeki mevcut durumu ele almaktır. Çalışmadan elde edilen bulgular, Türkiye’de Ar-Ge ve inovasyon harcamalarının yeterli düzeyde olmadığını, bunun yansıması olan patent başvuru ve kabul sayılarının çok düşük seviyelerde olduğu sonucunu göstermektedir.

  10. Polimorfisme Gly972Arg Gen IRS-1 dan Cys981Tyr Gen PTPN1 sebagai Faktor Risiko pada Sindrom Metabolik dengan Riwayat Berat Bayi Lahir Rendah

    Directory of Open Access Journals (Sweden)

    Hikmat Permana

    2012-09-01

    Full Text Available Increasing prevalence of obesity occurs worldwide and can affect all ages levels. Risk factors of increasing metabolic syndrome were associated with changes in lifestyle, obesity and low birth weight (LBW. This study was aimed to analyze the genotype IRS-1, genotype PTPN1 and LBW in metabolic syndrome risk factors. In 2009, this restrospective cohort study was comprised of 97 LBW and 100 normal birth weight (NBW, aged 20–21 years old, from the previous cohort study in District Tanjung Sari since 1989. The role of IRS-1 gene, PTPN1 gene and LBW in the metabolic syndrome factors were analyzed using multiple logistic regressions. The result showed there were no significant relationship between IRS-1 and PTPN1 genotype with metabolic syndrome factors but only IRS-1 genotype was significantly associated with trygliceride (p=0.006. Gly972Arg IRS-1 gene polymorphism in LBW has predictive value of increasing diastolic blood pressure, waist circumference and trygliceride. Cys981 Tyr PTPN1 gene polymorphism showed a risk factor for systolic blood pressure and high density lipoprotein (HDL cholesterol. Gly972Arg IRS-1 gene polymorphism was a dominant factor to increase diastolic blood pressure, waist circumference and trygliceride. In conclusions, Cys981 Tyr PTPN1 gene polymorphism is a risk factor of increased systolic blood pressure and HDL cholesterol, and LBW is the risk factor increasing fasting glucose level.

  11. Molecular mechanisms mediating the beneficial metabolic effects of [Arg4]tigerinin-1R in mice with diet-induced obesity and insulin resistance.

    Science.gov (United States)

    Ojo, Opeolu O; Srinivasan, Dinesh K; Owolabi, Bosede O; McGahon, Mary K; Moffett, R Charlotte; Curtis, Tim M; Conlon, J Michael; Flatt, Peter R; Abdel-Wahab, Yasser H A

    2016-08-01

    The frog skin host-defense peptide tigerinin-1R stimulates insulin release in vitro and improves glucose tolerance and insulin sensitivity in animal models of type 2 diabetes. This study extends these observations by investigating the molecular mechanisms of action underlying the beneficial metabolic effects of the analogue [Arg4]tigerinin-1R in mice with diet-induced obesity, glucose intolerance and insulin resistance. The study also investigates the electrophysiological effects of the peptide on KATP and L-type Ca2+ channels in BRIN-BD11 clonal β cells. Non-fasting plasma glucose and glucagon concentrations were significantly (pinsulin increased by twice daily treatment with [Arg4]tigerinin-1R (75 nmol/kg body weight) for 28 days. Oral and intraperitoneal glucose tolerance were significantly (pinsulin. The peptide blocked KATP channels and, consistent with this, improved beta cell responses of isolated islets to a range of secretagogues. Peptide administration resulted in up-regulation of key functional genes in islets involved insulin secretion (Abcc8, Kcnj11, Cacna1c and Slc2a2) and in skeletal muscle involved with insulin action (Insr, Irs1, Pdk1, Pik3ca, and Slc2a4). These observations encourage further development of tigerinin-1R analogues for the treatment of patients with type 2 diabetes.

  12. Severe respiratory phenotype caused by a de novo Arg528Gly mutation in the CACNA1S gene in a patient with hypokalemic periodic paralysis.

    Science.gov (United States)

    Kil, Tae-Hwan; Kim, June-Bum

    2010-05-01

    Hypokalemic periodic paralysis (HOKPP) is a rare disorder characterized by episodic muscle weakness with hypokalemia. Mutations in the CACNA1S gene, which encodes the alpha 1-subunit of the skeletal muscle L-type voltage-dependent calcium channel, have been reported to be mainly responsible for HOKPP. The paralytic attacks generally spare the respiratory muscles and the heart. Here, we report the case of a 16-year-old boy who presented with frequent respiratory insufficiency during the severe attacks. Mutational analysis revealed a heterozygous c.1582C>G substitution in the CACNA1S gene, leading to an Arg528Gly mutation in the protein sequence. The parents were clinically unaffected and did not show a mutation in the CACNA1S gene. A de novo Arg528Gly mutation has not previously been reported. The patient described here presents the unique clinical characteristics, including a severe respiratory phenotype and a reduced susceptibility to cold exposure. The patient did not respond to acetazolamide and showed a marked improvement of the paralytic symptoms on treatment with a combination of spironolactone, amiloride, and potassium supplements. Copyright 2009 European Paediatric Neurology Society. Published by Elsevier Ltd. All rights reserved.

  13. Mapping pathological phenotypes in reelin mutant mice.

    Science.gov (United States)

    Michetti, Caterina; Romano, Emilia; Altabella, Luisa; Caruso, Angela; Castelluccio, Paolo; Bedse, Gaurav; Gaetani, Silvana; Canese, Rossella; Laviola, Giovanni; Scattoni, Maria Luisa

    2014-01-01

    Autism Spectrum Disorders (ASD) are neurodevelopmental disorders with multifactorial origin characterized by social communication deficits and the presence of repetitive behaviors/interests. Several studies showed an association between the reelin gene mutation and increased risk of ASD and a reduced reelin expression in some brain regions of ASD subjects, suggesting a role for reelin deficiency in ASD etiology. Reelin is a large extracellular matrix glycoprotein playing important roles during development of the central nervous system. To deeply investigate the role of reelin dysfunction as vulnerability factor in ASD, we assessed the behavioral, neurochemical, and brain morphological features of reeler male mice. We recently reported a genotype-dependent deviation in the ultrasonic vocal repertoire and a general delay in motor development of reeler pups. We now report that adult male heterozygous (Het) reeler mice did not show social behavior and communication deficits during male-female social interactions. Wildtype and Het mice showed a typical light/dark locomotor activity profile, with a peak during the central interval of the dark phase. However, when faced with a mild stressful stimulus (a saline injection) only Het mice showed an over response to stress. In addition to the behavioral studies, we conducted high performance liquid chromatography and magnetic resonance imaging and spectroscopy to investigate whether reelin mutation influences brain monoamine and metabolites levels in regions involved in ASD. Low levels of dopamine in cortex and high levels of glutamate and taurine in hippocampus were detected in Het mice, in line with clinical data collected on ASD children. Altogether, our data detected subtle but relevant neurochemical abnormalities in reeler mice supporting this mutant line, particularly male subjects, as a valid experimental model to estimate the contribution played by reelin deficiency in the global ASD neurobehavioral phenotype.

  14. Ethanol production using nuclear petite yeast mutants

    Energy Technology Data Exchange (ETDEWEB)

    Hutter, A.; Oliver, S.G. [Department of Biomolecular Sciences, UMIST, Manchester (United Kingdom)

    1998-12-31

    Two respiratory-deficient nuclear petites, FY23{Delta}pet191 and FY23{Delta}cox5a, of the yeast Saccharomyces cerevisiae were generated using polymerase-chain-reaction-mediated gene disruption, and their respective ethanol tolerance and productivity assessed and compared to those of the parental grande, FY23WT, and a mitochondrial petite, FY23{rho}{sup 0}. Batch culture studies demonstrated that the parental strain was the most tolerant to exogenously added ethanol with an inhibition constant. K{sub i}, of 2.3% (w/v) and a specific rate of ethanol production, q{sub p}, of 0.90 g ethanol g dry cells{sup -1} h{sup -1}. FY23{rho}{sup 0} was the most sensitive to ethanol, exhibiting a K{sub i} of 1.71% (w/v) and q{sub p} of 0.87 g ethanol g dry cells{sup -1} h{sup -1}. Analyses of the ethanol tolerance of the nuclear petites demonstrate that functional mitochondria are essential for maintaining tolerance to the toxin with the 100% respiratory-deficient nuclear petite, FY23{Delta}pet191, having a K{sub i} of 2.14% (w/v) and the 85% respiratory-deficient FY23{Delta}cox5a, having a K{sub i} of 1.94% (w/v). The retention of ethanol tolerance in the nuclear petites as compared to that of FY23{rho}{sup 0} is mirrored by the ethanol productivities of these nuclear mutants, being respectively 43% and 30% higher than that of the respiratory-sufficient parent strain. This demonstrates that, because of their respiratory deficiency, the nuclear petites are not subject of the Pasteur effect and so exhibit higher rates of fermentation. (orig.)

  15. Mapping pathological phenotypes in Reelin mutant mice

    Directory of Open Access Journals (Sweden)

    Caterina eMichetti

    2014-09-01

    Full Text Available Autism Spectrum Disorders (ASD are neurodevelopmental disorders with multifactorial origin characterized by social communication and behavioural perseveration deficits. Several studies showed an association between the reelin gene mutation and increased risk of ASD and a reduced reelin expression in some brain regions of ASD subjects, suggesting a role for reelin deficiency in ASD etiology. Reelin is a large extracellular matrix glycoprotein playing important roles during development of the central nervous system. To deeply investigate the role of reelin dysfunction as vulnerability factor in ASD, we investigated the behavioural, neurochemical and brain morphological features of reeler male mice. We recently reported a genotype-dependent deviation in ultrasonic vocal repertoire and a general delay in motor development in reeler pups. We now report that adult male heterozygous reeler mice did not show social behaviour and communication deficits during male-female social interactions. Wildtype and heterozygous mice also showed a typical light/dark locomotor activity profile, with a peak during the central interval of the dark phase. However, when faced with a mild stressful stimulus (a saline injection only heterozygous mice showed an over response to stress. At the end of the behavioural studies, we conducted high performance liquid chromatography and magnetic resonance imaging and spectroscopy to investigate whether reelin mutation influences brain monoamine and metabolites levels in regions involved in ASD. Low levels of dopamine in cortex and high levels of glutamate and taurine in hippocampus were detected in heterozygous mice, in line with clinical data collected on ASD children. Altogether, our data detected subtle but relevant neurochemical abnormalities in reeler mice supporting this mutant line, particularly male subjects, as a valid experimental model to estimate the contribution played by reelin deficiency in the global ASD

  16. Proliferator-activated receptor gamma Pro12Ala interacts with the insulin receptor substrate 1 Gly972Arg and increase the risk of insulin resistance and diabetes in the mixed ancestry population from South Africa.

    Science.gov (United States)

    Vergotine, Zelda; Yako, Yandiswa Y; Kengne, Andre P; Erasmus, Rajiv T; Matsha, Tandi E

    2014-01-21

    The peroxisome proliferator-activated receptor gamma (PPARG), Pro12Ala and the insulin receptor substrate (IRS1), Gly972Arg confer opposite effects on insulin resistance and type 2 diabetes mellitus (T2DM). We investigated the independent and joint effects of PPARG Pro12Ala and IRS1 Gly972Arg on markers of insulin resistance and T2DM in an African population with elevated risk of T2DM. In all 787 (176 men) mixed-ancestry adults from the Bellville-South community in Cape Town were genotyped for PPARG Pro12Ala and IRS1 Gly972Arg by two independent laboratories. Glucose tolerance status and insulin resistance/sensitivity were assessed. Genotype frequencies were 10.4% (PPARG Pro12Ala) and 7.7% (IRS1 Gly972Arg). Alone, none of the polymorphisms predicted prevalent T2DM, but in regression models containing both alleles and their interaction term, PPARG Pro12 conferred a 64% higher risk of T2DM. Furthermore PPARG Pro12 was positively associated in adjusted linear regressions with increased 2-hour post-load insulin in non-diabetic but not in diabetic participants. The PPARG Pro12 is associated with insulin resistance and this polymorphism interacts with IRS1 Gly972Arg, to increase the risk of T2DM in the mixed-ancestry population of South Africa. Our findings require replication in a larger study before any generalisation and possible application for risk stratification.

  17. Mutant prevention concentrations of pradofloxacin for susceptible and mutant strains of Escherichia coli with reduced fluoroquinolone susceptibility.

    Science.gov (United States)

    Marcusson, Linda L; Komp Lindgren, Patricia; Olofsson, Sara K; Hughes, Diarmaid; Cars, Otto

    2014-10-01

    Pharmacodynamic and mutant prevention properties of the fluoroquinolone pradofloxacin (PRA) were measured against a set of 17 Escherichia coli strains carrying no, one or two known mutations conferring reduced fluoroquinolone susceptibility. The strains included susceptible wild-types, isogenic constructed mutants, isogenic selected mutants and clinical isolates. The effectiveness of PRA was determined with regard to preventing the selection of resistant mutants, using static and changing concentrations of drug. Ciprofloxacin was used as a reference drug. Minimum inhibitory concentrations (MICs) and mutant prevention concentrations (MPCs) of PRA for the susceptible wild-type strains were in the range 0.012-0.016mg/L and 0.2-0.3mg/L, respectively, giving a mean±standard deviation mutant prevention index (MPI=MPC/MIC) of 17.7±1.1. The mean MPI PRA of the 14 mutant strains was 19.2±12, and the mean MPI across all 17 strains was 18.9±10.8. In an in vitro kinetic model in which PRA was diluted with a half-life of 7h to mimic in vivo conditions, an initial concentration of PRA of 1.6-2.4mg/L (8-10× MPC), giving a PRA AUC/MPC ratio of 73-92, and a T>MPC of 21-23h was sufficient to prevent the selection of resistant mutants from the three susceptible wild-type strains. Dosing to reduce selection for antibiotic resistance in veterinary therapy has a role in reducing the reservoir of resistant mutants. We conclude that a level of dosing that prevents the selection of resistant mutants during therapy should be achievable in vivo. Copyright © 2014 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved.

  18. Effect of different immunosuppressive drugs on calcineurin and its mutants

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    Several mutants in Loop7 region and near Loop7 region of calcineurin A (CN A) subunit have been constructed and purified using site-directed mutagenesis.Their phosphatase activity and the corresponding solution conformation were examined.Their phosphatase activities between wild-type CN and mutants were compared to identify the interaction of different immunosuppressive drugs with CN.The results showed that the phosphatase activities of the mutants at Loop7 were much higher than the one of wild-type CN.Furthermore,circular dichroism spectra of the mutants revealed that their solution conformations gave rise in changes in native structure of the protein.Cyclophilin-CyclosporinA (CyP-CsA) significantly inhibited the phosphatase activity of wild-type CN,and had no effects on the phosphatase activity of mutants in Loop7 region,which indicates that the site-directed mutagenesis at Loop7 region made a significant change in the interaction between CyP-CsA and CN.Examination of the activities of these mutants resulted in the presence of immunosuppressive component from traditional Chinese drugs.The component of Chinese drug,ZIP1,could directly inhibit both CN and CN mutants without drug binding protein.These results suggest that the Loop7 region is an important structural area involved in the inhibition by CyP-CsA.It is valuable to further study the inhibition by ZIP1.

  19. Methods of producing protoporphyrin IX and bacterial mutants therefor

    Energy Technology Data Exchange (ETDEWEB)

    Zhou, Jizhong; Qiu, Dongru; He, Zhili; Xie, Ming

    2016-03-01

    The presently disclosed inventive concepts are directed in certain embodiments to a method of producing protoporphyrin IX by (1) cultivating a strain of Shewanella bacteria in a culture medium under conditions suitable for growth thereof, and (2) recovering the protoporphyrin IX from the culture medium. The strain of Shewanella bacteria comprises at least one mutant hemH gene which is incapable of normal expression, thereby causing an accumulation of protoporphyrin IX. In certain embodiments of the method, the strain of Shewanella bacteria is a strain of S. loihica, and more specifically may be S. loihica PV-4. In certain embodiments, the mutant hemH gene of the strain of Shewanella bacteria may be a mutant of shew_2229 and/or of shew_1140. In other embodiments, the presently disclosed inventive concepts are directed to mutant strains of Shewanella bacteria having at least one mutant hemH gene which is incapable of normal expression, thereby causing an accumulation of protoporphyrin IX during cultivation of the bacteria. In certain embodiments the strain of Shewanella bacteria is a strain of S. loihica, and more specifically may be S. loihica PV-4. In certain embodiments, the mutant hemH gene of the strain of Shewanella bacteria may be a mutant of shew_2229 and/or shew_1140.

  20. Epigenetic Suppression of T-DNA Insertion Mutants in Arabidopsis

    Institute of Scientific and Technical Information of China (English)

    Yangbin Gao; Yunde Zhao

    2013-01-01

    T-DNA insertion mutants have been widely used to define gene functions in Arabidopsis and in other plants.Here,we report an unexpected phenomenon of epigenetic suppression of T-DNA insertion mutants in Arabidopsis.When the two T-DNA insertion mutants,yucl-1 and ag-TD,were crossed together,the defects in all of the ag-TD plants in the F2 population were partially suppressed regardless of the presence of yucl-1.Conversion of ag-TD to the suppressed ag-TD (named as ag-TD*) did not follow the laws of Mendelian genetics.The ag-TD* could be stably transmitted for many generations without reverting to ag-TD,and ag-TD* had the capacity to convert ag-TD to ag-TD*.We show that epigenetic suppression of T-DNA mutants is not a rare event,but certain structural features in the T-DNA mutants are needed in order for the suppression to take place.The suppressed T-DNA mutants we observed were all intronic T-DNA mutants and the T-DNA fragments in both the trigger T-DNA as well as in the suppressed T-DNA shared stretches of identical sequences.We demonstrate that the suppression of intronic T-DNA mutants is mediated by trans-interactions between two ToDNA insertions.This work shows that caution is needed when intronic T-DNA mutants are used.

  1. Sensorimotor learning in Dab1(scm) (scrambler) mutant mice.

    Science.gov (United States)

    Lalonde, R; Strazielle, C

    2011-04-15

    Homozygous Dab1(scm) mouse mutants with cell ectopias in cerebellar cortex and neocortex were compared with non-ataxic controls on two tests of motor coordination: rotorod and grid climbing. Even at the minimal speed of 4 rpm and unlike controls, none of the Dab1(scm) mutants reached criterion on the constant speed rotorod. In contrast, Dab1(scm) mutants improved their performances on the vertical grid over the course of the same number of trials. Thus, despite massive cerebellar degeneration, sensorimotor learning for equilibrium is still possible, indicating the potential usefulness of the grid-climbing test in determining residual functions in mice with massive cerebellar damage.

  2. Fusion genetic analysis of jasmonate-signalling mutants in Arabidopsis

    DEFF Research Database (Denmark)

    Jensen, Anders Bøgh; Raventos, D.; Mundy, John Williams

    2002-01-01

    Jasmonates induce plant-defence responses and act to regulate defence-related genes including positive feedback of the lipoxygenase 2 (LOX2) gene involved in jasmonate synthesis. To identify jasmonate-signalling mutants, we used a fusion genetic strategy in which the firefly luciferase (FLUC...... as two recessive mutants, designated joe1 and 2, that overexpress the reporter. Genetic analysis indicated that reporter overexpression in the joe mutants requires COI. joe1 responded to MeJA with increased anthocyanin accumulation, while joe2 responded with decreased root growth inhibition. In addition...

  3. Structural and functional study of the GlnB22-insulin mutant responsible for maturity-onset diabetes of the young.

    Directory of Open Access Journals (Sweden)

    Květoslava Křížková

    Full Text Available The insulin gene mutation c.137G>A (R46Q, which changes an arginine at the B22 position of the mature hormone to glutamine, causes the monogenic diabetes variant maturity-onset diabetes of the young (MODY. In MODY patients, this mutation is heterozygous, and both mutant and wild-type (WT human insulin are produced simultaneously. However, the patients often depend on administration of exogenous insulin. In this study, we chemically synthesized the MODY mutant [GlnB22]-insulin and characterized its biological and structural properties. The chemical synthesis of this insulin analogue revealed that its folding ability is severely impaired. In vitro and in vivo tests showed that its binding affinity and biological activity are reduced (both approximately 20% that of human insulin. Comparison of the solution structure of [GlnB22]-insulin with the solution structure of native human insulin revealed that the most significant structural effect of the mutation is distortion of the B20-B23 β-turn, leading to liberation of the B chain C-terminus from the protein core. The distortion of the B20-B23 β-turn is caused by the extended conformational freedom of the GlnB22 side chain, which is no longer anchored in a hydrogen bonding network like the native ArgB22. The partially disordered [GlnB22]-insulin structure appears to be one reason for the reduced binding potency of this mutant and may also be responsible for its low folding efficiency in vivo. The altered orientation and flexibility of the B20-B23 β-turn may interfere with the formation of disulfide bonds in proinsulin bearing the R46Q (GlnB22 mutation. This may also have a negative effect on the WT proinsulin simultaneously biosynthesized in β-cells and therefore play a major role in the development of MODY in patients producing [GlnB22]-insulin.

  4. Mutant γPKC that causes spinocerebellar ataxia type 14 upregulates Hsp70, which protects cells from the mutant's cytotoxicity.

    Science.gov (United States)

    Ogawa, Kota; Seki, Takahiro; Onji, Tomoya; Adachi, Naoko; Tanaka, Shigeru; Hide, Izumi; Saito, Naoaki; Sakai, Norio

    2013-10-11

    Several missense mutations in the protein kinase Cγ (γPKC) gene have been found to cause spinocerebellar ataxia type 14 (SCA14), an autosomal dominant neurodegenerative disease. We previously demonstrated that the mutant γPKC found in SCA14 is misfolded, susceptible to aggregation and cytotoxic. Molecular chaperones assist the refolding and degradation of misfolded proteins and prevention of the proteins' aggregation. In the present study, we found that the expression of mutant γPKC-GFP increased the levels of heat-shock protein 70 (Hsp70) in SH-SY5Y cells. To elucidate the role of this elevation, we investigated the effect of siRNA-mediated knockdown of Hsp70 on the aggregation and cytotoxicity of mutant γPKC. Knockdown of Hsp70 exacerbated the aggregation and cytotoxicity of mutant γPKC-GFP by inhibiting this mutant's degradation. These findings suggest that mutant γPKC increases the level of Hsp70, which protects cells from the mutant's cytotoxicity by enhancing its degradation.

  5. Association between essential hypertension and polymorphisms of beta 1 adrenergic receptor gene G1165C (Gly389Arg) in Chinese Mongolian population

    Institute of Scientific and Technical Information of China (English)

    Rile Hu; Shigang Zhao; Guangming Niu; Chunyu Zhang; Zhiguang Wang; Mingfang Jiang

    2006-01-01

    BACKGROUND: The prevalences of hypertension, cerebrovascular diseases, etc. are higher in Mongolian population because of the influence of various factors including genetics, geography, diet, etc. Therefore, it is helpful to develop researches on the genetics of various diseases including hypertension in Mongolian population.OBJECTIVE: To analyze the association between the polymorphism of beta1 adrenergic receptor (β1-AR)gene G1165C (Arg389Gly), an important candidate gene for various diseases of cardiovascular system, and essential hypertension in Mongolian population.DESIGN: A cross-sectional study.SETTINGS: Department of Neurology, the First Affiliated Hospital of Inner Mongolia Medical College; Wulate Houqi Red Cross Society.PARTICIPANTS: The survey was carried out from February 2003 to March 2005. Totally 239 Mongolian residents, whose blood relations of 3 generations were all Mongolians, were selected from Wulate Houqi, Inner Mongolia, and they were all informed with the survey and detected items. Based on the diagnostic standard of hypertension set by WHO in 1999, the subjects were divided into two groups according to the level blood pressure: ① Normal blood pressure group (n=117): systolic blood pressure (SBP) < 140 mm Hg (1 mm Hg =0.133 kPa), diastolic blood pressure (DBP) < 90 mm Hg, and those having histories of cerebrovascular disease, heart disease, diseases of liver, kidney and tiroides, and diabetes mellitus were excluded. ② Essential hypertension group (n=122): including 51 patients with simple high SBP. All the enrolled subjects had no blood relationship with each other, and had no history of miscegenation.METHODS: The body height, body mass, waist circumference and blood lipids were measured routinely, and their habits of smoking and drinking were also investigated. Peripheral venous blood (5 mL) was drawn, the genome DNA was extracted, and the polymorphisms of the β1-AR G1165C (Gly389Arg) genotype were detected with the Sequenom system

  6. The Gly16 Allele of the Gly16Arg Single-Nucleotide Polymorphism in the β2-Adrenergic Receptor Gene Augments Perioperative Use of Vasopressors

    DEFF Research Database (Denmark)

    Nielsen, Morten; Staalsø, Jonatan Myrup; Ullum, Henrik

    2016-01-01

    receptor (ADRB2) influences perioperative arterial blood pressure and consequently the use of vasopressors. METHODS: Five hundred seventy-one Danish Caucasians undergoing neurosurgery were genotyped for 5 marker single-nucleotide polymorphisms (SNPs) within ADRB2 (Gly16Arg, Gln27Glu, Thr164Ile, Arg175Arg......, and Gly351Gly). A pairwise tagging principle was used to identify ADRB2 haplotypes. Mean arterial blood pressure (MAP) was recorded in the supine awake state and, together with administration of vasopressors (ephedrine and/or phenylephrine), for 30 minutes after induction of general anesthesia...... (sevoflurane/remifentanil or propofol/remifentanil). RESULTS: Four hundred thirteen (72%) patients received ephedrine and/or phenylephrine. Only baseline MAP (P

  7. Development and dissection of diagnostic SNP markers for the downy mildew resistance genes Pl Arg and Pl 8 and maker-assisted gene pyramiding in sunflower (Helianthus annuus L.).

    Science.gov (United States)

    Qi, L L; Talukder, Z I; Hulke, B S; Foley, M E

    2017-02-03

    Diagnostic DNA markers are an invaluable resource in breeding programs for successful introgression and pyramiding of disease resistance genes. Resistance to downy mildew (DM) disease in sunflower is mediated by Pl genes which are known to be effective against the causal fungus, Plasmopara halstedii. Two DM resistance genes, Pl Arg and Pl 8 , are highly effective against P. halstedii races in the USA, and have been previously mapped to the sunflower linkage groups (LGs) 1 and 13, respectively, using simple sequence repeat (SSR) markers. In this study, we developed high-density single nucleotide polymorphism (SNP) maps encompassing the Pl arg and Pl 8 genes and identified diagnostic SNP markers closely linked to these genes. The specificity of the diagnostic markers was validated in a highly diverse panel of 548 sunflower lines. Dissection of a large marker cluster co-segregated with Pl Arg revealed that the closest SNP markers NSA_007595 and NSA_001835 delimited Pl Arg to an interval of 2.83 Mb on the LG1 physical map. The SNP markers SFW01497 and SFW06597 delimited Pl 8 to an interval of 2.85 Mb on the LG13 physical map. We also developed sunflower lines with homozygous, three gene pyramids carrying Pl Arg , Pl 8 , and the sunflower rust resistance gene R 12 using the linked SNP markers from a segregating F2 population of RHA 340 (carrying Pl 8 )/RHA 464 (carrying Pl Arg and R 12 ). The high-throughput diagnostic SNP markers developed in this study will facilitate marker-assisted selection breeding, and the pyramided sunflower lines will provide durable resistance to downy mildew and rust diseases.

  8. Research method and progress on antibiotics resistance genes(ARGs) in air%空气中抗性基因(ARGs)的研究方法及研究进展

    Institute of Scientific and Technical Information of China (English)

    贺小萌; 曹罡; 邵明非; 李继

    2014-01-01

    The long-term overuse of antibiotics lead to the emergence of antibiotic resistant bacteria and accelerate the transport and the spread of antibiotic resistance genes ( ARGs ) in different environmental matrix, which poses potential health risks to humans and animals. As a new category of environmental contaminants, ARGs have been one of the hot topics in the area of environmental research in recent years. Most research, however, focuses on AGRs in water, soil and sediment. There have been only a few studies on ARGs in air. This paper summarizes the research progress of ARGs in air and discusses the methods that can be used for collecting and detecting ARGs samples in air. The aim of this paper is to provide the scientific basis and technical strategies to support the development of studies on ARGs in air.%抗生素的长期滥用导致大量耐药菌的出现,并加剧抗生素抗性基因( ARGs)在不同环境介质中传播扩散,对人类和动物健康造成潜在威胁。作为一种新型污染物,ARGs已经成为近年来环境研究领域的热点之一。然而目前多数研究关注的是水、土壤和沉积物中的ARGs,国内外对空气中ARGs的研究相对较少且零散。本文综述了空气中ARGs的国内外研究现状,并探讨了空气中ARGs样品的采集和检测方法,旨在为空气中ARGs的研究提供科学依据和技术策略。

  9. Selection of mutants of capsicum annuum induced by gamma ray

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Y. I.; Lee, Y. B. [Korea Atomic Energy Research Institute, Taejeon (Korea, Republic of); Lee, E. K. [Chungnam National Univ., Taejeon (Korea, Republic of)

    1998-06-01

    For induction and selection of mutations of Capsicum annuum L., dry seeds of pure lines No.1 and No.2 were irradiated with gamma ray of 150Gy, 200Gy and 250Gy. Various mutants were selected such as showing early maturity, short plant height, long fruit and chlorophyll mutations. Mutation frequency of No.1 line was 3.4% in the dose of 150Gy, while the frequency of No.2 line was 2.7% in the dose of 250Gy. For selection of resistant mutant to amino acid analog, the optimum concentration of 5-methyltryptophan (5-MT) and S-(2-aminoethyl)-L-cysteine were 25 ppm and 30 ppm, respectively. Four resistant mutant lines to 5-MT were selected among 400 mutant lines.

  10. Characterization of a Salmonella typhimurium mutant defective in phosphoribosylpyrophosphate synthetase

    DEFF Research Database (Denmark)

    Jochimsen, Bjarne; Hove-Jensen, Bjarne; Garber, Bruce B.;

    1985-01-01

    This study describes the isolation and characterization of a mutant (strain GP122) of Salmonella typhimurium with a partial deficiency of phosphoribosylpyrophosphate (PRPP) synthetase activity. This strain was isolated in a purE deoD gpt purine auxotroph by a procedure designed to select guanosine......-utilizing mutants. Strain GP122 had roughly 15% of the PRPP synthetase activity and 25% of the PRPP pool of its parent strain. The mutant exhibited many of the predicted consequences of a decreased PRPP pool and a defective PRPP synthetase enzyme, including: poor growth on purine bases; decreased accumulation of 5...... phosphoribosyltransferase, enzymes involved in the pyrimidine de novo biosynthetic pathway; growth stimulation by PRPP-sparing compounds (e.g. guanosine, histidine); poor growth in low phosphate medium; and increased heat lability of the defective enzyme. This mutant strain also had increased levels of guanosine 5...

  11. Generation of Peroxisome-Deficient Somatic Animal Cell Mutants.

    Science.gov (United States)

    Okumoto, Kanji; Fujiki, Yukio

    2017-01-01

    Cell mutants with a genetic defect affecting various cellular phenotypes are widely utilized as a powerful tool in genetic, biochemical, and cell biological research. More than a dozen complementation groups of animal somatic mutant cells defective in peroxisome biogenesis have been successfully isolated in Chinese hamster ovary (CHO) cells and used as a model system reflecting fatal human severe genetic disorders named peroxisome biogenesis disorders (PBD). Isolation and characterization of peroxisome-deficient CHO cell mutants has allowed the identification of PEX genes and the gene products peroxins, which directly leads to the accomplishment of isolation of pathogenic genes responsible for human PBDs, as well as elucidation of their functional roles in peroxisome biogenesis. Here, we describe the procedure to isolate peroxisome-deficient mammalian cell mutants from CHO cells, by making use of an effective, photo-sensitized selection method.

  12. Status and Perspectives on the Researches of Rice Glutelin Mutants

    Institute of Scientific and Technical Information of China (English)

    ZHENG Tian-qing; SHEN Wen-biao; ZHU Su-song; ZHAI Hu-qu; WAN Jian-min

    2003-01-01

    Rice (Oryza sativa L. ) is one of the model plants for genomics research. As the raising offunctional rice breeding for special usage, glutelin mutants play a more and more important role in the func-tional rice breeding as well as eukaryotic gene expression and regulation research materials. For example, therice cultivar special for the patients suffering from kidney disease and diabetes could be developed from the riceglutelin mutants. In this paper, current researches on characterization, mutation mechanism and breeding us-age of various rice glutelin mutants, especially the low glutelin content cultivars, were all discussed with per-spectives on the trends of the glutelin mutant researches in the era of post-genomics.

  13. Phosphoribosylpyrophosphate synthetase of Escherichia coli, Identification of a mutant enzyme

    DEFF Research Database (Denmark)

    Hove-Jensen, Bjarne; Nygaard, Per

    1982-01-01

    , stimulated the mutant enzyme. The activity of PRib-PP synthetase in crude extract was higher in the mutant than in the parent. When starved for purines an accumulation of PRib-PP was observed in the parent strain, while the pool decreased in the mutant. During pyrimidine starvation derepression of PRib....... Kinetic analysis of the mutant PRib-PP synthetase revealed an apparent Km for ATP and ribose 5-phosphate of 1.0 mM and 240 μM respectively, compared to 60 μM and 45 μM respectively for the wild-type enzyme. ADP, which inhibits the wild-type enzyme at a concentration of 0.5 mM ribose 5-phosphate...

  14. Assessment of Genetic diversity in mutant cowpea lines using ...

    African Journals Online (AJOL)

    FKOLADE

    2016-11-09

    Nov 9, 2016 ... option of NTSYS, a rooted tree was also generated from the .... Dellarporta SF, Wood J, Hicks JB (1983). ... genetic diversity in Pigeon Pea (Cajanus sp). ... diversity in somatic mutants of grape (Vitis vinifera) cultivar Italia.

  15. Analysis of the aspartic acid metabolic pathway using mutant genes.

    Science.gov (United States)

    Azevedo, R A

    2002-01-01

    Amino acid metabolism is a fundamental process for plant growth and development. Although a considerable amount of information is available, little is known about the genetic control of enzymatic steps or regulation of several pathways. Much of the information about biochemical pathways has arisen from the use of mutants lacking key enzymes. Although mutants were largely used already in the 60's, by bacterial and fungal geneticists, it took plant research a long time to catch up. The advance in this area was rapid in the 80's, which was followed in the 90's by the development of techniques of plant transformation. In this review we present an overview of the aspartic acid metabolic pathway, the key regulatory enzymes and the mutants and transgenic plants produced for lysine and threonine metabolism. We also discuss and propose a new study of high-lysine mutants.

  16. Phosphoribosylpyrophosphate synthetase of Escherichia coli, Identification of a mutant enzyme

    DEFF Research Database (Denmark)

    Hove-Jensen, Bjarne; Nygaard, Per

    1982-01-01

    . Kinetic analysis of the mutant PRib-PP synthetase revealed an apparent Km for ATP and ribose 5-phosphate of 1.0 mM and 240 μM respectively, compared to 60 μM and 45 μM respectively for the wild-type enzyme. ADP, which inhibits the wild-type enzyme at a concentration of 0.5 mM ribose 5-phosphate......, stimulated the mutant enzyme. The activity of PRib-PP synthetase in crude extract was higher in the mutant than in the parent. When starved for purines an accumulation of PRib-PP was observed in the parent strain, while the pool decreased in the mutant. During pyrimidine starvation derepression of PRib...

  17. Characterization of Glutamine-Requiring Mutants of Pseudomonas aeruginosa

    NARCIS (Netherlands)

    Janssen, Dick B.; Joosten, Han M.L.J.; Herst, Patricia M.; Drift, Chris van der

    1982-01-01

    Revertants were isolated from a glutamine-requiring mutant of Pseudomonas aeruginosa PAO. One strain showed thermosensitive glutamine requirement and formed thermolabile glutamine synthetase, suggesting the presence of a mutation in the structural gene for glutamine synthetase. The mutation conferri

  18. Resistant mechanism study of benzalkonium chloride selected Salmonella Typhimurium mutants.

    Science.gov (United States)

    Guo, Wei; Cui, Shenghui; Xu, Xiao; Wang, Haoyan

    2014-02-01

    Benzalkonium chloride is one of the invaluable biocides that is extensively used in healthcare settings as well as in the food processing industry. After exposing wild-type Salmonella Typhimurium 14028s or its AcrAB inactivation mutant to gradually increasing levels of benzalkonium chloride, resistance mutants S-41, S-150, S-AB-23, S-AB-38, and S-AB-73 were selected and these mutants also showed a 2-64-fold stable minimum inhibitory concentration (MIC) increase to chloramphenicol, ciprofloxacin, nalidixic acid, and tetracycline. In S-41 and S-150, the expression of acrB was increased 2.7- and 7.6-fold, and ΔtolC or ΔacrAB mutants of S-41 and S-150 showed the same MICs to all tested antimicrobials as the equivalent Salmonella Typhimurium 14028s mutants. However, in S-AB-23, S-AB-38, and S-AB-73, the expression of acrF was increased 96-, 230-, and 267-fold, respectively, and ΔtolC or ΔacrEF mutants of S-AB-23, S-AB-38, and S-AB-73 showed the similar MICs to all tested antimicrobials as the ΔtolC mutant of Salmonella Typhimurium 14028s. Our data showed that constitutively over-expressed AcrAB working through TolC was the main resistance mechanism in ST14028s benzalkonium chloride resistance mutants. However, after AcrAB had been inactivated, benzalkonium chloride-resistant mutants could still be selected and constitutively over-expressed, AcrEF became the dominant efflux pump working through TolC and being responsible for the increasing antimicrobial resistance. These data indicated that different mechanisms existed for acrB and acrF constitutive over-expression. Since exposure to benzalkonium chloride may lead to Salmonella mutants with a decreased susceptibility to quinolones, which is currently one of the drugs of choice for the treatment of life-threatening salmonelosis, research into the pathogenesis and epidemiology of the benzalkonium chloride resistance mutants will be of increasing importance.

  19. Targeting Palmitoyl Acyltransferases in Mutant NRAS-Driven Melanoma

    Science.gov (United States)

    2014-08-01

    regulation of synaptic and neuronal functions.17 A point mutation in DHHC21 was identified in the depilated (dep) mouse mutant, resulting in hair follicle ...and hair follicle differentiation. PLoS Genet. 5, e1000748. (19) Mansilla, F., Birkenkamp-Demtroder, K., Kruhoffer, M., Sorensen, F. B., Andersen, C...AWARD NUMBER: W81XWH-13-1-0203 TITLE: Targeting Palmitoyl Acyltransferases in Mutant NRAS-Driven Melanoma PRINCIPAL INVESTIGATOR: Xu Wu

  20. A relaxed mutant with an altered ribosomal protein L11.

    Science.gov (United States)

    Parker, J; Watson, R J; Friesen, J D

    1976-02-27

    Relaxed mutants of Escherichia coli have been isolated which have an altered electrophoretic mobility of ribosomal protein L11. It can be shown that reversion to stringency in one of these mutants occurs simultaneously with a reversion of L11 protein to tis normal mobility. The L11 structural gene, rplK, maping near rif, is carried by the bacteriophage lambdacI857S7drifd18, and is most likely identical with relC.

  1. Multiple defects in Escherichia coli mutants lacking HU protein.

    OpenAIRE

    Huisman, O; Faelen, M; Girard, D; Jaffé, A; Toussaint, A; Rouvière-Yaniv, J

    1989-01-01

    The HU protein isolated from Escherichia coli, composed of two partially homologous subunits, alpha and beta, shares some of the properties of eucaryotic histones and is a major constituent of the bacterial nucleoid. We report here the construction of double mutants totally lacking both subunits of HU protein. These mutants exhibited poor growth and a perturbation of cell division, resulting in the formation of anucleate cells. In the absence of HU, phage Mu was unable to grow, to lysogenize,...

  2. Survival bias and drug interaction can attenuate cross-sectional case-control comparisons of genes with health outcomes. An example of the kinesin-like protein 6 (KIF6 Trp719Arg polymorphism and coronary heart disease

    Directory of Open Access Journals (Sweden)

    Pendyala Lakshmana

    2011-03-01

    Full Text Available Abstract Background Case-control studies typically exclude fatal endpoints from the case set, which we hypothesize will substantially underestimate risk if survival is genotype-dependent. The loss of fatal cases is particularly nontrivial for studies of coronary heart disease (CHD because of significantly reduced survival (34% one-year fatality following a coronary attack. A case in point is the KIF6 Trp719Arg polymorphism (rs20455. Whereas six prospective studies have shown that carriers of the KIF6 Trp719Arg risk allele have 20% to 50% greater CHD risk than non-carriers, several cross-sectional case-control studies failed to show that carrier status is related to CHD. Computer simulations were therefore employed to assess the impact of the loss of fatal events on gene associations in cross-sectional case-control studies, using KIF6 Trp719Arg as an example. Results Ten replicates of 1,000,000 observations each were generated reflecting Canadian demographics. Cardiovascular disease (CVD risks were assigned by the Framingham equation and events distributed among KIF6 Trp719Arg genotypes according to published prospective studies. Logistic regression analysis was used to estimate odds ratios between KIF6 genotypes. Results were examined for 33%, 41.5%, and 50% fatality rates for incident CVD. In the absence of any difference in percent fatalities between genotypes, the odds ratios (carriers vs. noncarriers were unaffected by survival bias, otherwise the odds ratios were increasingly attenuated as the disparity between fatality rates increased between genotypes. Additional simulations demonstrated that statin usage, shown in four clinical trials to substantially reduce the excess CHD risk in the KIF6 719Arg variant, should also attenuate the KIF6 719Arg odds ratio in case-control studies. Conclusions These computer simulations show that exclusions of prior CHD fatalities attenuate odds ratios of case-control studies in proportion to the difference

  3. [Polymorphic markers Ala455Val of the THBD gene and Arg353Gln of the F7 gene and association with unfavorable outcomes of coronary atherosclerosis in patients with a history of acute ischemic heart disease].

    Science.gov (United States)

    Pushkov, A A; Blagodatskikh, K A; Nikitin, A G; Agapkina, Iu V; Brovkin, A N; Chudakova, D A; Evdokimova, M A; Aseĭcheva, O Iu; Osmolovskaia, V S; Minushkina, L O; Baklanova, T N; Talyzin, P A; Donetskaia, O P; Tereshchenko, S N; Dzhaiani, N A; Akatova, E A; Glezer, M G; Galiavich, A S; Zakirova, V B; Koziolova, N A; Iagoda, A V; Boeva, O I; Horolets, E V; Shlyk, S V; Volkova, E G; Margarian, M P; Guz', I O; Konstantinov, V O; Sidorenko, B A; Zateĭshchikov, D A; Nosikov, V V

    2011-10-01

    The polymorphic markers Ala455Val of the THBD gene and Arg353Gln of the F7 gene were tested for association with the frequency of unfavorable outcomes in patients with a history of acute ischemic heart disease. The study involved 1145 patients hospitalized in cardiology clinics of Moscow, St. Petersburg, Kazan, Chelyabinsk, Perm, Stavropol, and Rostov-on-Don because of acute ischemic heart disease. The patients were followed up for up to 62.5 months. None of the markers displayed a significant association with the time to an endpoint. The patients were then grouped by sex. In females, the frequency of unfavorable outcomes (fatal or nonfatal myocardial infarction and fatal or nonfatal stroke) was higher in carriers of allele Val of the Ala344Val polymorphic marker of the THBD gene and carriers of genotype Arg/Arg of the Arg353Gln polymorphic marker of the F7 gene, but the difference was not statistically significant. Such an increase in frequency was not observed in males. To study the combined effect of the polymorphic markers of the THBD and F7 genes, the course of ischemic heart disease was compared for two female subgroups. One included carriers of allele Val of the Ala344Val polymorphic marker of the THBD gene and genotype Arg/Arg of the Arg353Gln polymorphic marker of the F7 gene; the other subgroup included carriers ofgenotype Ala/Ala of the Ala455Val polymorphic marker of the THBD gene and allele Gln of the Arg353Gln polymorphic marker of the F7 gene. The frequency of unfavorable outcomes in the first subgroup was higher than in the second one. The time to an endpoin was 40.5 months (95% confidence interval (CI) 33.5-47.6) in the first subgroup and 51.6 months (95% CI 45.0-58.1) in the second subgroup (chi2 = 4.15, P = 0.042). The results made it possible to assume that the F7 and THBD genes play an important role in genetic predisposition to unfavorable outcomes in patients with a history of acute ischemic heart disease.

  4. From the potent and selective mu opioid receptor agonist H-Dmt-d-Arg-Phe-Lys-NH(2) to the potent delta antagonist H-Dmt-Tic-Phe-Lys(Z)-OH.

    Science.gov (United States)

    Balboni, Gianfranco; Cocco, Maria Teresa; Salvadori, Severo; Romagnoli, Romeo; Sasaki, Yusuke; Okada, Yoshio; Bryant, Sharon D; Jinsmaa, Yunden; Lazarus, Lawrence H

    2005-08-25

    H-Dmt-d-Arg-Phe-Lys-NH(2) ([Dmt(1)]DALDA) binds with high affinity and selectivity to the mu opioid receptor and is a potent and long-acting analgesic. Substitution of d-Arg in position 2 with Tic and masking of the lysine amine side chain by Z protection and of the C-terminal carboxylic function instead of the amide function transform a potent and selective mu agonist into a potent and selective delta antagonist H-Dmt-Tic-Phe-Lys(Z)-OH. Such a delta antagonist could be used as a pharmacological tool.

  5. Isolation of new gravitropic mutants under hypergravity conditions

    Directory of Open Access Journals (Sweden)

    Akiko Mori

    2016-09-01

    Full Text Available Forward genetics is a powerful approach used to link genotypes and phenotypes, and mutant screening/analysis has provided deep insights into many aspects of plant physiology. Gravitropism is a tropistic response in plants, in which hypocotyls and stems sense the direction of gravity and grow upwards. Previous studies of gravitropic mutants have suggested that shoot endodermal cells in Arabidopsis stems and hypocotyls are capable of sensing gravity (i.e., statocytes. In the present study, we report a new screening system using hypergravity conditions to isolate enhancers of gravitropism mutants, and we also describe a rapid and efficient genome mapping method, using Next-Generation Sequencing (NGS and Single Nucleotide Polymorphism (SNP-based markers. Using the endodermal-amyloplast less 1 (eal1 mutant, which exhibits defective development of endodermal cells and gravitropism, we found that hypergravity (10 g restored the reduced gravity responsiveness in eal1 hypocotyls and could, therefore, be used to obtain mutants with further reduction in gravitropism in the eal1 background. Using the new screening system, we successfully isolated six ene (enhancer of eal1 mutants that exhibited little or no gravitropism under hypergravity conditions, and using NGS and map-based cloning with SNP markers, we narrowed down the potential causative genes, which revealed a new genetic network for shoot gravitropism in Arabidopsis.

  6. Isolation of New Gravitropic Mutants under Hypergravity Conditions

    Science.gov (United States)

    Mori, Akiko; Toyota, Masatsugu; Shimada, Masayoshi; Mekata, Mika; Kurata, Tetsuya; Tasaka, Masao; Morita, Miyo T.

    2016-01-01

    Forward genetics is a powerful approach used to link genotypes and phenotypes, and mutant screening/analysis has provided deep insights into many aspects of plant physiology. Gravitropism is a tropistic response in plants, in which hypocotyls and stems sense the direction of gravity and grow upward. Previous studies of gravitropic mutants have suggested that shoot endodermal cells in Arabidopsis stems and hypocotyls are capable of sensing gravity (i.e., statocytes). In the present study, we report a new screening system using hypergravity conditions to isolate enhancers of gravitropism mutants, and we also describe a rapid and efficient genome mapping method, using next-generation sequencing (NGS) and single nucleotide polymorphism (SNP)-based markers. Using the endodermal-amyloplast less 1 (eal1) mutant, which exhibits defective development of endodermal cells and gravitropism, we found that hypergravity (10 g) restored the reduced gravity responsiveness in eal1 hypocotyls and could, therefore, be used to obtain mutants with further reduction in gravitropism in the eal1 background. Using the new screening system, we successfully isolated six ene (enhancer of eal1) mutants that exhibited little or no gravitropism under hypergravity conditions, and using NGS and map-based cloning with SNP markers, we narrowed down the potential causative genes, which revealed a new genetic network for shoot gravitropism in Arabidopsis.

  7. Molecular analysis of mutants of the Neurospora adenylosuccinate synthetase locus

    Indian Academy of Sciences (India)

    A. Wiest; A. J. McCarthy; R. Schnittker; K. McCluskey

    2012-08-01

    The ad-8 gene of Neurospora crassa, in addition to being used for the study of purine biology, has been extensively studied as a model for gene structure, mutagenesis and intralocus recombination. Because of this there is an extensive collection of well-characterized N. crassa ad-8 mutants in the Fungal Genetics Stock Center collection. Among these are spontaneous mutants and mutants induced with X-ray, UV or chemical mutagens. The specific lesions in these mutants have been genetically mapped at high resolution. We have sequenced the ad-8 locus from 13 of these mutants and identified the molecular nature of the mutation in each strain. We compare the historical fine-structure map to the DNA and amino acid sequence of each allele. The placement of the individual lesions in the fine-structure map was more accurate at the 5′ end of the gene and no mutants were identified in the 3′ untranslated region of this gene. We additionally analysed ad-8+ alleles in 18 N. crassa strains subjected to whole-genome sequence analysis and describe the variability among Neurospora strains and among fungi and other organisms.

  8. Non-opioid nociceptive activity of human dynorphin mutants that cause neurodegenerative disorder spinocerebellar ataxia type 23

    NARCIS (Netherlands)

    Watanabe, Hiroyuki; Mizoguchi, Hirokazu; Verbeek, Dineke S.; Kuzmin, Alexander; Nyberg, Fred; Krishtal, Oleg; Sakurada, Shinobu; Bakalkin, Georgy

    2012-01-01

    We previously identified four missense mutations in the prodynorphin gene that cause human neurodegenerative disorder spinocerebellar ataxia type 23 (SCA23). Three mutations substitute Leu(5), Arg(6), and Arg(9) to Ser (L5S), Trp (R6W) and Cys (R9C) in dynorphin A(1-17) (Dyn A), a peptide with both

  9. Non-opioid nociceptive activity of human dynorphin mutants that cause neurodegenerative disorder spinocerebellar ataxia type 23

    NARCIS (Netherlands)

    Watanabe, Hiroyuki; Mizoguchi, Hirokazu; Verbeek, Dineke S.; Kuzmin, Alexander; Nyberg, Fred; Krishtal, Oleg; Sakurada, Shinobu; Bakalkin, Georgy

    2012-01-01

    We previously identified four missense mutations in the prodynorphin gene that cause human neurodegenerative disorder spinocerebellar ataxia type 23 (SCA23). Three mutations substitute Leu(5), Arg(6), and Arg(9) to Ser (L5S), Trp (R6W) and Cys (R9C) in dynorphin A(1-17) (Dyn A), a peptide with both

  10. No association of the neuropeptide Y (Leu7Pro) and ghrelin gene (Arg51Gln, Leu72Met, Gln90Leu) single nucleotide polymorphisms with eating disorders.

    Science.gov (United States)

    Kindler, Jochen; Bailer, Ursula; de Zwaan, Martina; Fuchs, Karoline; Leisch, Friedrich; Grün, Bettina; Strnad, Alexandra; Stojanovic, Mirjana; Windisch, Julia; Lennkh-Wolfsberg, Claudia; El-Giamal, Nadja; Sieghart, Werner; Kasper, Siegfried; Aschauer, Harald

    2011-06-01

    Genetic factors likely contribute to the biological vulnerability of eating disorders. Case-control association study on one neuropeptide Y gene (Leu7Pro) polymorphism and three ghrelin gene (Arg51Gln, Leu72Met and Gln90Leu) polymorphisms. 114 eating disorder patients (46 with anorexia nervosa, 30 with bulimia nervosa, 38 with binge eating disorder) and 164 healthy controls were genotyped. No differences were detected between patients and controls for any of the four polymorphisms in allele frequency and genotype distribution (P > 0.05). Allele frequencies and genotypes had no significant influence on body mass index (P > 0.05) in eating disorder patients. Positive findings of former case-control studies of associations between ghrelin gene polymorphisms and eating disorders could not be replicated. Neuropeptide Y gene polymorphisms have not been investigated in eating disorders before.

  11. Cabezo Pardo. Análisis instrumental de materiales de construcción de barro del yacimiento argárico

    OpenAIRE

    Martínez Mira, I.; Vilaplana Ortego, E.; Such Basañez, I.; Juan Juan, J.; García del Cura, M. Ángeles

    2014-01-01

    Partiendo del análisis instrumental de cuatro pequeños fragmentos constructivos se intenta, en lo posible, contestar a una serie de cuestiones básicas señaladas por algunos autores (De Chazelles y Poupet, 1989: 10-11; Sánchez García, 1995: 350; 1997: 142) para poder comprender el nivel técnico de una época o cultura y la evolución de sus técnicas constructivas, en este caso aplicadas a la fase argárica del yacimiento del Cabezo Pardo. MARQ, Museo Arqueológico de Alicante, Diputación de Ali...

  12. Cabezo Pardo. Análisis instrumental de materiales de construcción de barro del yacimiento argárico

    OpenAIRE

    2014-01-01

    Partiendo del análisis instrumental de cuatro pequeños fragmentos constructivos se intenta, en lo posible, contestar a una serie de cuestiones básicas señaladas por algunos autores (De Chazelles y Poupet, 1989: 10-11; Sánchez García, 1995: 350; 1997: 142) para poder comprender el nivel técnico de una época o cultura y la evolución de sus técnicas constructivas, en este caso aplicadas a la fase argárica del yacimiento del Cabezo Pardo. MARQ, Museo Arqueológico de Alicante, Diputación de Ali...

  13. 乌桕菌用林栽培技术%Cultivation techniques of Sapium discolor ( Champ. ) Mfiell - Arg for fungus-producing forest

    Institute of Scientific and Technical Information of China (English)

    彭彪; 陈孝丑; 高文; 雷莹; 林雄平

    2011-01-01

    Sapium discolor (Champ.) Mtiell - Arg with a long cultivation history and versatility is a unique arbor species China, which is a versatile tree used as energy, herb, wood, fungus and ornament. The paper reviews and summarizes the local distribution, Biol%乌桕是我国特有的乔木树种,栽培历史悠久、用途广泛,是一种集能源、药用、材用、菌用、观赏为一体的多用途树种.阐述了乌桕的地区分布、适生环境和生物学特性,并就乌桕菌用林栽培技术进行了实验,为乌桕菌用林播种育苗、人工林培育提供借鉴经验.

  14. SPATIAL PATTERN OF REGENERATION OF Aspidosperma polyneuron Müll. Arg. IN A REMNANT SECTOR SUBMITTED TO SELECTIVE USE IN THE NORTH OF MISIONES PROVINCE

    Directory of Open Access Journals (Sweden)

    Martín Martín Pinazo

    2009-10-01

    Full Text Available The spatial pattern of regeneration of Aspidosperma polyneuron Müll.Arg. was studied in a remnant sector of the Paranaense forest in the Misiones province, Argentina. All individuals were mapped in a 210 m x 100 m area (2,1 ha. Univariate and bivariate point pattern analyses were used by means of Wiegand and Moloney O ring index. Regeneration showed clumped pattern to scales up to10 m and clusters of 30 m of diameter. No association was found between regeneration and both adult trees of A. polyneuron and standing death trees, whereas a small scale relationship between regeneration and adult trees of all species was observed. These results revealed that the regeneration of Aspidosperma polyneuron is strongly restricted to the presence of conspecific adult trees. On the other hand, the recruitments are associated with individuals from different species with DBH > 10 cm and infrequent in open sites.

  15. A non-synonymous coding change in the CYP19A1 gene Arg264Cys (rs700519 does not affect circulating estradiol, bone structure or fracture

    Directory of Open Access Journals (Sweden)

    Wang Jenny Z

    2011-12-01

    Full Text Available Abstract Background The biosynthesis of estrogens from androgens is catalyzed by aromatase P450 enzyme, coded by the CYP19A1 gene on chromosome 15q21.2. Genetic variation within the CYP19A1 gene sequence has been shown to alter the function of the enzyme. The aim of this study is to investigate whether a non-synonymous Arg264Cys (rs700519 single nucleotide polymorphism (SNP is associated with altered levels of circulating estradiol, areal bone mineral density or fracture. Methods This population- based study of 1,022 elderly Caucasian women (mean age 74.95 ± 2.60 years was genotyped for the rs700519 SNP were analyzed to detect any association with endocrine and bone phenotypes. Results The genotype frequencies were 997 wildtype (97.6%, 24 heterozygous (2.3% and 1 homozygous (0.1%. When individuals were grouped by genotype, there was no association between the polymorphism and serum estradiol (wildtype 27.5 ± 16.0; variants 31.2 ± 18.4, P = 0.27. There was also no association seen on hip bone mineral density (wildtype 0.81 ± 0.12; 0.84 ± 0.14 for variants, P = 0.48 or femoral neck bone mineral density (0.69 ± 0.10 for wildtype; 0.70 ± 0.12 for variants, P = 0.54 before or after correction of the data with age, height, weight and calcium therapy. There were also no associations with quantitative ultrasound measures of bone structure (broadband ultrasound attenuation, speed of sound and average stiffness. Conclusions In a cohort of 1,022 elderly Western Australian women, the presence of Arg264Cys (rs700519 polymorphism was not found to be associated with serum estradiol, bone structure or phenotypes.

  16. The conserved residue Arg46 in the N-terminal heptad repeat domain of HIV-1 gp41 is critical for viral fusion and entry.

    Directory of Open Access Journals (Sweden)

    Xiaoyi Wang

    Full Text Available During the process of HIV-1 fusion with the target cell, the N-terminal heptad repeat (NHR of gp41 interacts with the C-terminal heptad repeat (CHR to form fusogenic six-helix bundle (6-HB core. We previously identified a crucial residue for 6-HB formation and virus entry--Lys63 (K63 in the C-terminal region of NHR (aa 54-70, which forms a hydrophobic cavity. It can form an important salt bridge with Asp121 (D121 in gp41 CHR. Here, we found another important conserved residue for virus fusion and entry, Arg46 (R46, in the N-terminal region of NHR (aa 35-53, which forms a hydrogen bond with a polar residue, Asn43 (N43, in NHR, as a part of the hydrogen-bond network. R46 can also form a salt bridge with a negatively charged residue, Glu137 (E137, in gp41 CHR. Substitution of R46 with the hydrophobic residue Ala (R46A or the negatively charged residue Glu (R46E resulted in disruption of the hydrogen bond network, breakage of the salt bridge and reduction of 6-HB's stability, leading to impairment of viral fusion and decreased inhibition of N36, an NHR peptide. Similarly, CHR peptide C34 with substitution of E137 for Ala (E137A or Arg (E137R also exhibited reduced inhibitory activity against HIV-1 infection and HIV-1-mediated cell-to-cell fusion. These results suggest that the positively charged residue R46 and its hydrogen bond network, together with the salt bridge between R46 and E137, are important for viral fusion and entry and may therefore serve as a target for designing novel HIV fusion/entry inhibitors.

  17. Covalent attachment of cell-adhesive peptide Gly-Arg-Gly-Asp (GRGD) to poly(etheretherketone) surface by tailored silanization layers technique

    Energy Technology Data Exchange (ETDEWEB)

    Zheng, Yanyan [Chengdu Institute of Organic Chemistry, Chinese Academy of Sciences, Chengdu 610041 (China); University of Chinese Academy of Sciences, Beijing 100049 (China); Xiong, Chengdong [Chengdu Institute of Organic Chemistry, Chinese Academy of Sciences, Chengdu 610041 (China); Li, Xiaoyu [State Key Laboratory of Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu 610041 (China); Zhang, Lifang, E-mail: zhanglfcioc@163.com [Chengdu Institute of Organic Chemistry, Chinese Academy of Sciences, Chengdu 610041 (China)

    2014-11-30

    Highlights: • The carbonyl groups on PEEK surface were effectively reduced to hydroxyl groups using sodium borohydride. • Silanization layers technique was employed to immobilize the cell-adhesive peptide Gly-Arg-Gly-Asp (GRGD) on hydroxylation-pretreated PEEK sheet surface by covalent chemical attachment. • XPS, surface profiler and water contact angle measurements proved the presence of GRGD on PEEK surface. • Osteoblast-like cells (MC3T3-E1) attachment and proliferation were improved effectively on GRGD-modified PEEK surface. - Abstract: Poly(etheretherketone) (PEEK) is a rigid semicrystalline polymer that combines excellent mechanical properties, broad chemical resistance and bone-like stiffness and is widely used in biomedical fields. However, PEEK is naturally bioinert, leading to limited biomedical applications, especially when a direct bone-implant osteointegration is desired. In this study, a three-step reaction procedure was employed to immobilize the cell-adhesive peptide Gly-Arg-Gly-Asp (GRGD) on the surface of PEEK sheet by covalent chemical attachment to favor cell adhesion and proliferation. First, hydroxylation-pretreated PEEK surfaces were silanized with 7-Oct-1-enyltrichlorosilane (OETS) in dry cyclohexane, resulting in a silanization layer with terminal ethenyl. Second, the terminal ethylenic double bonds of the silanization layer on PEEK surface were converted to carboxyl groups through acidic potassium manganate oxidation. Finally, GRGD was covalently attached by carbodiimide mediated condensation between the carboxyl on PEEK surface and amine presents in GRGD. X-ray photoelectron spectroscopy (XPS), attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy, surface profiler and water contact angle measurements were applied to characterize the modified surfaces. The effect of cells attachment and proliferation on each specimen was investigated. Pre-osteoblast cells (MC3T3-E1) attachment, spreading and proliferation

  18. The alpha-galactosidase A p.Arg118Cys variant does not cause a Fabry disease phenotype: data from individual patients and family studies

    Science.gov (United States)

    Ferreira, Susana; Ortiz, Alberto; Germain, Dominique P.; Viana-Baptista, Miguel; Gomes, António Caldeira; Camprecios, Marta; Fenollar-Cortés, Maria; Gallegos-Villalobos, Ángel; Garcia, Diego; García-Robles, José Antonio; Egido, Jesús; Gutiérrez-Rivas, Eduardo; Herrero, José Antonio; Mas, Sebastián; Oancea, Raluca; Péres, Paloma; Salazar-Martín, Luis Manuel; Solera-Garcia, Jesús; Alves, Helena; Garman, Scott C.; Oliveira, João Paulo

    2015-01-01

    Summary Lysosomal α-galactosidase A (α-Gal) is the enzyme deficient in Fabry disease (FD), an X-linked glycosphingolipidosis caused by pathogenic mutations affecting the GLA gene. The early-onset, multi-systemic FD classical phenotype is associated with absent or severe enzyme deficiency, as measured by in vitro assays, but patients with higher levels of residual α-Gal activity may have later-onset, more organ-restricted clinical presentations. A change in the codon 118 of the wild-type α-Gal sequence, replacing basic arginine by a potentially sulfhydryl-binding cysteine residue – GLA p.(Arg118Cys) –, has been recurrently described in large FD screening studies of high-risk patients. Although the Cys118 allele is associated with high residual α-Gal activity in vitro, it has been classified as a pathogenic mutation, mainly on the basis of theoretical arguments about the chemistry of the cysteine residue. However its pathogenicity has never been convincingly demonstrated by pathology criteria. We reviewed the clinical, biochemical and histopathology data obtained from 22 individuals of Portuguese and Spanish ancestry carrying the Cys118 allele, including 3 homozygous females. Cases were identified either on the differential diagnosis of possible FD manifestations and on case-finding studies (n=11; 4 males), or on unbiased cascade screening of probands’ close relatives (n=11; 3 males). Overall, those data strongly suggest that the GLA p.(Arg118Cys) variant does not segregate with FD clinical phenotypes in a Mendelian fashion, but might be a modulator of the multifactorial risk of cerebrovascular disease, since the allelic frequency in stroke patients was 0.0087 (p=0.0185 vs the general population). The Cys118 allelic frequency in healthy Portuguese adults (n=696) has been estimated as 0.001, therefore not qualifying for “rare” condition. PMID:25468652

  19. Spontaneous chlorophyll mutants of Pennisetum americanum: Genetics and chlorophyll quantities.

    Science.gov (United States)

    Koduru, P R; Rao, M K

    1980-05-01

    Thirteen spontaneously occurring chlorophyll deficient phenotypes have been described and their genetic basis was established. Ten of these - 'white', 'white tipped green', 'patchy white', 'white virescent', 'white striping 1', 'white striping 2', 'white striping 4', 'fine striping', 'chlorina' and 'yellow virescent' showed monogenic recessive inheritance and the remaining three - 'yellow striping', 'yellow green' and 'light green' seedling phenotypes showed digenic recessive inheritance. The genes for (i) 'white tipped green' (wr) and 'yellow virescent' (yv) and (ii) 'patchy white' (pw) and 'white striping 1' (wst 1) showed independent assortment. Further, the genes for 'white' (w), 'white tipped green' (wr) and 'yellow virescent' (yv) were inherited independently of the gene for hairy leaf margin (Hm).In the mutants - 'white tipped green', 'patchy white', 'white striping 1', 'white striping 2', 'fine striping', 'chlorina', 'yellow virescent', 'yellow striping', 'yellow green' and 'light green' phenotypes total quantity of chlorophyll was significantly less than that in the corresponding controls, while in 'white virescent' there was no reduction in the mature stage. For nine of the mutants the quantity of chlorophyll was also estimated in F1's (mutant x control green). In F1's of six of the mutants - 'white tip', 'patchy white', 'chlorina', 'yellow virescent', 'fine striping' and 'yellow striping' the quantity of chlorophyll was almost equal to the wild type. In the F1's of three of the mutants - 'white striping 1', 'white striping 2' and 'light green' an intermediate value between the mutant and wild types was observed. In 'yellow virescent' retarded synthesis of chlorophyll, particularly chlorophyll a was observed in the juvenile stage. Reduced quantity of chlorophyll was associated with defective chloroplasts. In the mutants - 'white tipped green, 'white virescent', 'fine striping', 'chlorina', 'yellow striping', 'yellow green' and 'light green' defective

  20. Isolation and characterization of selenate resistant mutants of Acremonium chrysogenum

    Directory of Open Access Journals (Sweden)

    Airton Vialta

    1999-01-01

    Full Text Available Mutants unable to convert exogenous sulfate to sulfite were isolated using the toxic analogue selenate. Three of twenty-eight isolated mutants were chromate sensitive. They showed a possible lesion in the gene that codes the ATP sulfurylase. The others were chromate resistant, and probably had a lesion in one or both of the genes that code the sulfate permease. Methionine increased the resistance levels to selenate. In addition, the frequency of spontaneous mutants obtained in a medium containing methionine was higher (between 2.4 x 10-6 and 18.0 x 10-6 than that obtained using a medium without any intentional source of sulfur (between 0.7 x 10-6 and 5.0 x 10-6. The original strain, as well as the mutants, were able to grow in a sulfur-free liquid medium even after 4 consecutive inoculation procedures. These results indicated the existence of sulfur traces in the medium and/or an efficient intracellular storage system. There was no significant difference between cephalosporin C production in mutants and the original strain.Mutantes incapazes de converter o sulfato extracelular em sulfito foram isolados utilizando o análogo tóxico selenato. De 28 mutantes isolados, apenas 3 foram sensíveis ao cromato, provavelmente apresentando lesão no gene que codifica a ATP sulfurilase. Os demais foram resistentes ao cromato e devem conter lesão no gene sB ou também no gene sC. A metionina elevou os níveis de resistência ao selenato e a freqüência de mutantes espontâneos obtida em meio contendo este aminoácido foi maior (entre 2,42 x 10-6 e 18,04 x 10-6 do que a obtida no meio sem a adição de qualquer fonte intencional de enxofre (entre 0,71 x 10-6 e 5,0 x 10-6. A linhagem original e os mutantes foram capazes de crescer, mesmo depois de quatro etapas de inóculo, fato que pode ser explicado pela existência de traços do referido elemento no meio e/ou a presença de um sistema eficiente de estocagem intracelular. A produção de cefalosporina C

  1. Isolation of Escherichia coli mutants defective in uptake of molybdate.

    Science.gov (United States)

    Hemschemeier, S; Grund, M; Keuntje, B; Eichenlaub, R

    1991-10-01

    For the study of molybdenum uptake by Escherichia coli, we generated Tn5lac transposition mutants, which were screened for the pleiotropic loss of molybdoenzyme activities. Three mutants A1, A4, and M22 were finally selected for further analysis. Even in the presence of 100 microM molybdate in the growth medium, no active nitrate reductase, formate dehydrogenase, and trimethylamine-N-oxide reductase were detected in these mutants, indicating that the intracellular supply of molybdenum was not sufficient. This was also supported by the observation that introduction of plasmid pWK225 carrying the complete nif regulon of Klebsiella pneumoniae did not lead to a functional expression of nitrogenase. Finally, molybdenum determination by induced coupled plasma mass spectroscopy confirmed a significant reduction of cell-bound molybdenum in the mutants compared with that in wild-type E. coli, even at high molybdate concentrations in the medium. A genomic library established with the plasmid mini-F-derived cop(ts) vector pJE258 allowed the isolation of cosmid pBK229 complementing the molybdate uptake deficiency of the chlD mutant and the Tn5lac-induced mutants. Certain subfragments of pBK229 which do not contain the chlD gene are still able to complement the Tn5lac mutants. Mapping experiments showed that the Tn5lac insertions did not occur within the chromosomal region present in pBK229 but did occur very close to that region. We assume that the Tn5lac insertions have a polar effect, thus preventing the expression of transport genes, or that a positively acting regulatory element was inactivated.

  2. Defective glycinergic synaptic transmission in zebrafish motility mutants

    Directory of Open Access Journals (Sweden)

    Hiromi Hirata

    2010-01-01

    Full Text Available Glycine is a major inhibitory neurotransmitter in the spinal cord and brainstem. Recently, in vivo analysis of glycinergic synaptic transmission has been pursued in zebrafish using molecular genetics. An ENU mutagenesis screen identified two behavioral mutants that are defective in glycinergic synaptic transmission. Zebrafish bandoneon (beo mutants have a defect in glrbb, one of the duplicated glycine receptor (GlyR β subunit genes. These mutants exhibit a loss of glycinergic synaptic transmission due to a lack of synaptic aggregation of GlyRs. Due to the consequent loss of reciprocal inhibition of motor circuits between the two sides of the spinal cord, motor neurons activate simultaneously on both sides resulting in bilateral contraction of axial muscles of beo mutants, eliciting the so-called ‘accordion’ phenotype. Similar defects in GlyR subunit genes have been observed in several mammals and are the basis for human hyperekplexia/startle disease. By contrast, zebrafish shocked (sho mutants have a defect in slc6a9, encoding GlyT1, a glycine transporter that is expressed by astroglial cells surrounding the glycinergic synapse in the hindbrain and spinal cord. GlyT1 mediates rapid uptake of glycine from the synaptic cleft, terminating synaptic transmission. In zebrafish sho mutants, there appears to be elevated extracellular glycine resulting in persistent inhibition of postsynaptic neurons and subsequent reduced motility, causing the ‘twitch once’ phenotype. We review current knowledge regarding zebrafish ‘accordion’ and ‘twitch once’ mutants, including beo and sho, and report the identification of a new α2 subunit that revises the phylogeny of zebrafish GlyRs.

  3. Auditory development in progressive motor neuronopathy mouse mutants.

    Science.gov (United States)

    Volkenstein, Stefan; Brors, Dominik; Hansen, Stefan; Berend, Achim; Mlynski, Robert; Aletsee, Christoph; Dazert, Stefan

    2009-11-06

    The present study was performed to elucidate the hearing development in the progressive motor neuronopathy (pmn) mouse mutant. This mouse has been used as a model for human motoneuron disease. A missense mutation in the tubulin-specific chaperon E (Tbce) gene on mouse chromosome 13 was localized as the underlying genetic defect. The protein encoded by the Tbce gene is essential for the formation of primary tubulin complexes. Studies on motoneurons show disorganization in microtubules and disturbed axonal transport, followed by retrograde degeneration of the motoneurons. A similar pathomechanism is also possible for hearing disorders where disrupted microtubules could cause functional deficits in spiral ganglion neurons or in cochlear hair cells. Click auditory brainstem response (ABR) audiometry in homozygous pmn mutants showed a normal onset of hearing, but an increasing hearing threshold from postnatal day 26 (P26) on to death, compared to heterozygous mutants and wild-type mice. Histological sections of the cochlea at different ages showed a regular morphology. Additionally, spiral ganglion explants from mutant and wild-type mice were cultured. The neurite length from pmn mutants was shorter than in wild-type mice, and the neurite number/explant was significantly decreased in pmn mutants. We show that the pmn mouse mutant is a model for a progressive rapid hearing loss from P26 on, after initially normal hearing development. Heterozygous mice are not affected by this defect. With the knowledge of the well-known pathomechanism of this defect in motoneurons, a dysfunction of cellular mechanisms regulating tubulin assembling suggests that tubulin assembling plays an essential role in hearing function and maintenance.

  4. The Trp64Arg amino acid polymorphism of the beta3-adrenergic receptor gene does not contribute to the genetic susceptibility of diabetic microvascular complications in Caucasian type 1 diabetic patients

    DEFF Research Database (Denmark)

    Tarnow, L; Urhammer, S A; Mottlau, B

    1999-01-01

    OBJECTIVE: The beta3-adrenergic receptor is involved in regulation of microvascular blood flow. A missense mutation (Trp64Arg) in the beta3-adrenergic receptor gene has been suggested as a risk factor for proliferative retinopathy in Japanese type 2 diabetic patients. The aim of the present study...

  5. Development of pre-implantation porcine embryos cultured within a three-dimensional alginate hydrogel system either conjugated with Arg-Gly-Asp (RGD) peptide or supplemented with secreted phosphoprotein 1 (SPP1)

    Science.gov (United States)

    Many uterine specific factors have been shown to be increased within the uterine milieu as the porcine embryo initiates elongation. Secreted phosphoprotein 1 (SPP1) is increased during this time and contains an Arg-Gly-Asp (RGD) peptide sequence that has been shown to bind to cell surface integrins ...

  6. Ancestry of the Brazilian TP53 c.1010G>A (p.Arg337His, R337H) Founder Mutation: Clues from Haplotyping of Short Tandem Repeats on Chromosome 17p

    Science.gov (United States)

    Paskulin, Diego Davila; Giacomazzi, Juliana; Achatz, Maria Isabel; Costa, Sandra; Reis, Rui Manoel; Hainaut, Pierre; dos Santos, Sidney Emanuel Batista; Ashton-Prolla, Patricia

    2015-01-01

    Rare germline mutations in TP53 (17p13.1) cause a highly penetrant predisposition to a specific spectrum of early cancers, defining the Li-Fraumeni Syndrome (LFS). A germline mutation at codon 337 (p.Arg337His, c1010G>A) is found in about 0.3% of the population of Southern Brazil. This mutation is associated with partially penetrant LFS traits and is found in the germline of patients with early cancers of the LFS spectrum unselected for familial history. To characterize the extended haplotypes carrying the mutation, we have genotyped 9 short tandem repeats on chromosome 17p in 12 trios of Brazilian p.Arg337His carriers. Results confirm that all share a common ancestor haplotype of Caucasian/Portuguese-Iberic origin, distant in about 72–84 generations (2000 years assuming a 25 years intergenerational distance) and thus pre-dating European migration to Brazil. So far, the founder p.Arg337His haplotype has not been detected outside Brazil, with the exception of two residents of Portugal, one of them of Brazilian origin. On the other hand, increased meiotic recombination in p.Arg337His carriers may account for higher than expected haplotype diversity. Further studies comparing haplotypes in populations of Brazil and of other areas of Portuguese migration are needed to understand the historical context of this mutation in Brazil. PMID:26618902

  7. The common SLC30A8 Arg325Trp variant is associated with reduced first-phase insulin release in 846 non-diabetic offspring of type 2 diabetes patients--the EUGENE2 study

    DEFF Research Database (Denmark)

    Boesgaard, T W; Zilinskaite, J; Vänttinen, M

    2008-01-01

    AIMS/HYPOTHESIS: A recent genome-wide association study identified the SLC30A8 rs13266634 polymorphism encoding an Arg325Trp polymorphism in the zinc transporter protein member 8 (ZnT-8) to be associated with type 2 diabetes. Here, we investigate whether the polymorphism is related to altered ins...

  8. EMMA—mouse mutant resources for the international scientific community

    Science.gov (United States)

    Wilkinson, Phil; Sengerova, Jitka; Matteoni, Raffaele; Chen, Chao-Kung; Soulat, Gaetan; Ureta-Vidal, Abel; Fessele, Sabine; Hagn, Michael; Massimi, Marzia; Pickford, Karen; Butler, Richard H.; Marschall, Susan; Mallon, Ann-Marie; Pickard, Amanda; Raspa, Marcello; Scavizzi, Ferdinando; Fray, Martin; Larrigaldie, Vanessa; Leyritz, Johan; Birney, Ewan; Tocchini-Valentini, Glauco P.; Brown, Steve; Herault, Yann; Montoliu, Lluis; de Angelis, Martin Hrabé; Smedley, Damian

    2010-01-01

    The laboratory mouse is the premier animal model for studying human disease and thousands of mutants have been identified or produced, most recently through gene-specific mutagenesis approaches. High throughput strategies by the International Knockout Mouse Consortium (IKMC) are producing mutants for all protein coding genes. Generating a knock-out line involves huge monetary and time costs so capture of both the data describing each mutant alongside archiving of the line for distribution to future researchers is critical. The European Mouse Mutant Archive (EMMA) is a leading international network infrastructure for archiving and worldwide provision of mouse mutant strains. It operates in collaboration with the other members of the Federation of International Mouse Resources (FIMRe), EMMA being the European component. Additionally EMMA is one of four repositories involved in the IKMC, and therefore the current figure of 1700 archived lines will rise markedly. The EMMA database gathers and curates extensive data on each line and presents it through a user-friendly website. A BioMart interface allows advanced searching including integrated querying with other resources e.g. Ensembl. Other resources are able to display EMMA data by accessing our Distributed Annotation System server. EMMA database access is publicly available at http://www.emmanet.org. PMID:19783817

  9. Effect of different immunosuppressive drugs on calcineurin and its mutants

    Institute of Scientific and Technical Information of China (English)

    阎力君; 于翠娟; 张丽芳; 魏群

    2000-01-01

    Several mutants in Loop7 region and near Loop7 region of calcineurin A (CN A) subunit have been constructed and purified using site-directed mutagenesis. Their phosphatase activity and the corresponding solution conformation were examined. Their phosphatase activities between wild-type CN and mutants were compared to identify the interaction of different immuno-suppressive drugs with CN. The results showed that the phosphatase activities of the mutants at Loop7 were much higher than the one of wild-type CN. Furthermore, circular dichroism spectra of the mutants revealed that their solution conformations gave rise in changes in native structure of the protein. Cyclophilin-CyclosporinA (CyP-CsA) significantly inhibited the phosphatase activity of wild-type CN, and had no effects on the phosphatase activity of mutants in Loop7 region, which indicates that the site-directed mutagenesis at Loop7 region made a significant change in the interaction between CyP-CsA and CN. Examination of the activities of these

  10. Normal aging modulates the neurotoxicity of mutant huntingtin.

    Directory of Open Access Journals (Sweden)

    Elsa Diguet

    Full Text Available Aging likely plays a role in neurodegenerative disorders. In Huntington's disease (HD, a disorder caused by an abnormal expansion of a polyglutamine tract in the protein huntingtin (Htt, the role of aging is unclear. For a given tract length, the probability of disease onset increases with age. There are mainly two hypotheses that could explain adult onset in HD: Either mutant Htt progressively produces cumulative defects over time or "normal" aging renders neurons more vulnerable to mutant Htt toxicity. In the present study, we directly explored whether aging affected the toxicity of mutant Htt in vivo. We studied the impact of aging on the effects produced by overexpression of an N-terminal fragment of mutant Htt, of wild-type Htt or of a beta-Galactosidase (beta-Gal reporter gene in the rat striatum. Stereotaxic injections of lentiviral vectors were performed simultaneously in young (3 week and old (15 month rats. Histological evaluation at different time points after infection demonstrated that the expression of mutant Htt led to pathological changes that were more severe in old rats, including an increase in the number of small Htt-containing aggregates in the neuropil, a greater loss of DARPP-32 immunoreactivity and striatal neurons as assessed by unbiased stereological counts.The present results support the hypothesis that "normal" aging is involved in HD pathogenesis, and suggest that age-related cellular defects might constitute potential therapeutic targets for HD.

  11. Mutant alpha-synuclein and autophagy in PC12 cells

    Institute of Scientific and Technical Information of China (English)

    Kangyong Liu; Chunfeng Liu; Chuancheng Ren; Yaping Yang; Liwei Shen; Xuezhong Li; Fen Wang; Zhenghong Qin

    2011-01-01

    Several studies have demonstrated that overexpression of mutant α-synuclein in PC12 cells is related to occurrence of autophagy.The present study established mutant a-synuclein (A30P)-transfected PC12 cells and treated them with the autophagy inducer rapamycin and autophagy inhibitor wortmannin, respectively.Results demonstrated that mutant o-synuclein resulted in cell death via autophagy and involved α-synuclein accumulation, membrane lipid oxidation, and loss of plasma membrane integrity.Mutant α-synuclein (A30P) also mediated toxicity of1-methyl-4-phenylpyridinium ion.Moreover, rapamycin inhibited a-synuclein aggregation, while wortmannin promoted o-synuclein aggregation and cell death.To further determine the role of autophagy due to mutant a-synuclein, the present study measured expression of microtubule-associated protein light chain 3.Results revealed that wortmannin and 1-methyl-4-phenylpyridinium ion inhibited expression of microtubule-associated protein light chain 3,while rapamycin promoted its expression.These findings suggested that abnormal aggregation of a-synuclein induced autophagic programmed cell death in PC12 cells.

  12. Cytokinin production by plant growth promoting rhizobacteria and selected mutants.

    Science.gov (United States)

    García de Salamone, I E; Hynes, R K; Nelson, L M

    2001-05-01

    One of the proposed mechanisms by which rhizobacteria enhance plant growth is through the production of plant growth regulators. Five plant growth promoting rhizobacterial (PGPR) strains produced the cytokinin dihydrozeatin riboside (DHZR) in pure culture. Cytokinin production by Pseudomonas fluorescens G20-18, a rifampicin-resistant mutant (RIF), and two TnphoA-derived mutants (CNT1, CNT2), with reduced capacity to synthesize cytokinins, was further characterized in pure culture using immunoassay and thin layer chromatography. G20-18 produced higher amounts of three cytokinins, isopentenyl adenosine (IPA), trans-zeatin ribose (ZR), and DHZR than the three mutants during stationary phase. IPA was the major metabolite produced, but the proportion of ZR and DHZR accumulated by CNT1 and CNT2 increased with time. No differences were observed between strain G20-18 and the mutants in the amounts of indole acetic acid synthesized, nor were gibberellins detected in supernatants of any of the strains. Addition of 10(-5) M adenine increased cytokinin production in 96- and 168-h cultures of strain G20-18 by approximately 67%. G20-18 and the mutants CNT1 and CNT2 may be useful for determination of the role of cytokinin production in plant growth promotion by PGPR.

  13. Characterization of a mutant glucose isomerase from Thermoanaerobacterium saccharolyticum.

    Science.gov (United States)

    Xu, Heng; Shen, Dong; Wu, Xue-Qiang; Liu, Zhi-Wei; Yang, Qi-He

    2014-10-01

    A series of site-directed mutant glucose isomerase at tryptophan 139 from Thermoanaerobacterium saccharolyticum strain B6A were purified to gel electrophoretic homogeneity, and the biochemical properties were determined. W139F mutation is the most efficient mutant derivative with a tenfold increase in its catalytic efficiency toward glucose compared with the native GI. With a maximal activity at 80 °C of 59.58 U/mg on glucose, this mutant derivative is the most active type ever reported. The enzyme activity was maximal at 90 °C and like other glucose isomerase, this mutant enzyme required Co(2+) or Mg(2+) for enzyme activity and thermal stability (stable for 20 h at 80 °C in the absence of substrate). Its optimum pH was around 7.0, and it had 86 % of its maximum activity at pH 6.0 incubated for 12 h at 60 °C. This enzyme was determined as thermostable and weak-acid stable. These findings indicated that the mutant GI W139F from T. saccharolyticum strain B6A is appropriate for use as a potential candidate for high-fructose corn syrup producing enzyme.

  14. Mutant number distribution in an exponentially growing population

    Science.gov (United States)

    Keller, Peter; Antal, Tibor

    2015-01-01

    We present an explicit solution to a classic model of cell-population growth introduced by Luria and Delbrück (1943 Genetics 28 491-511) 70 years ago to study the emergence of mutations in bacterial populations. In this model a wild-type population is assumed to grow exponentially in a deterministic fashion. Proportional to the wild-type population size, mutants arrive randomly and initiate new sub-populations of mutants that grow stochastically according to a supercritical birth and death process. We give an exact expression for the generating function of the total number of mutants at a given wild-type population size. We present a simple expression for the probability of finding no mutants, and a recursion formula for the probability of finding a given number of mutants. In the ‘large population-small mutation’ limit we recover recent results of Kessler and Levine (2014 J. Stat. Phys. doi:10.1007/s10955-014-1143-3) for a fully stochastic version of the process.

  15. Hepatitis B escape mutants in Scottish blood donors.

    Science.gov (United States)

    Larralde, Osmany; Dow, Brian; Jarvis, Lisa; Davidson, Fiona; Petrik, Juraj

    2013-06-01

    Hepatitis B virus (HBV) remains as the viral infection with the highest risk of transmission by transfusion. This risk is associated with window period donations, occult HBV infection (OBI) and the emergence of escape mutants, which render blood donations false negative for hepatitis B surface antigen (HBsAg) serological testing. A retrospective study was conducted to gain insights into the molecular epidemiology of HBV escape mutants in Scottish blood donors. The criterion for selection was HBV positivity either by serology or nucleic acid testing (NAT). HBsAg detection was compared across several commercial immunoassays. The full length S gene from plasma samples was PCR amplified, cloned and expressed in HepG2 cells. Eight samples showed HBsAg discordant results, while 5 OBI samples were found. Four escape mutants, containing missense mutations in the S gene, are described here. These mutations impaired HBsAg detection both from HBV infected plasma samples and from recombinant proteins derived from its infected donors. Phylogenetic analysis showed that most of the mutants were clustered in the genotype D and were closely related to strains from Asia and the Middle East. We report here a proline substitution, outside the major hydrophilic region, that impaired HBsAg detection in vivo and in vitro, warning about the risk for the emergence of vaccine escape mutants with mutations outside the major neutralisation site.

  16. Rescue of cystathionine beta-synthase (CBS) mutants with chemical chaperones: purification and characterization of eight CBS mutant enzymes.

    Science.gov (United States)

    Majtan, Tomas; Liu, Lu; Carpenter, John F; Kraus, Jan P

    2010-05-21

    Missense mutations represent the most common cause of many genetic diseases including cystathionine beta-synthase (CBS) deficiency. Many of these mutations result in misfolded proteins, which lack biological function. The presence of chemical chaperones can sometimes alleviate or even restore protein folding and activity of mutant proteins. We present the purification and characterization of eight CBS mutants expressed in the presence of chemical chaperones such as ethanol, dimethyl sulfoxide, or trimethylamine-N-oxide. Preliminary screening in Escherichia coli crude extracts showed that their presence during protein expression had a significant impact on the amount of recovered CBS protein, formation of tetramers, and catalytic activity. Subsequently, we purified eight CBS mutants to homogeneity (P49L, P78R, A114V, R125Q, E176K, P422L, I435T, and S466L). The tetrameric mutant enzymes fully saturated with heme had the same or higher specific activities than wild type CBS. Thermal stability measurements demonstrated that the purified mutants are equally or more thermostable than wild type CBS. The response to S-adenosyl-L-methionine stimulation or thermal activation varied. The lack of response of R125Q and E176K to both stimuli indicated that their specific conformations were unable to reach the activated state. Increased levels of molecular chaperones in crude extracts, particularly DnaJ, indicated a rather indirect effect of the chemical chaperones on folding of CBS mutants. In conclusion, the chemical chaperones present in the expression medium were able to fully restore the activity of eight CBS mutants by improving their protein folding. This finding could have direct implications for the development of a therapeutical approach to pyridoxine unresponsive homocystinuria.

  17. p21-ras effector domain mutants constructed by "cassette" mutagenesis

    DEFF Research Database (Denmark)

    Stone, J C; Vass, W C; Willumsen, B M;

    1988-01-01

    A series of mutations encoding single-amino-acid substitutions within the v-rasH effector domain were constructed, and the ability of the mutants to induce focal transformation of NIH 3T3 cells was studied. The mutations, which spanned codons 32 to 40, were made by a "cassette" mutagenesis...... technique that involved replacing this portion of the v-rasH effector domain with a linker carrying two BspMI sites in opposite orientations. Since BspMI cleaves outside its recognition sequence, BspMI digestion of the plasmid completely removed the linker, creating a double-stranded gap whose missing ras...... sequences were reconstructed as an oligonucleotide cassette. Based upon the ability of the mutants to induce focal transformation of NIH 3T3 cells, a range of phenotypes from virtually full activity to none (null mutants) was seen. Three classes of codons were present in this segment: one which could...

  18. How Life History Can Sway the Fixation Probability of Mutants.

    Science.gov (United States)

    Li, Xiang-Yi; Kurokawa, Shun; Giaimo, Stefano; Traulsen, Arne

    2016-07-01

    In this work, we study the effects of demographic structure on evolutionary dynamics when selection acts on reproduction, survival, or both. In contrast to the previously discovered pattern that the fixation probability of a neutral mutant decreases while the population becomes younger, we show that a mutant with a constant selective advantage may have a maximum or a minimum of the fixation probability in populations with an intermediate fraction of young individuals. This highlights the importance of life history and demographic structure in studying evolutionary dynamics. We also illustrate the fundamental differences between selection on reproduction and selection on survival when age structure is present. In addition, we evaluate the relative importance of size and structure of the population in determining the fixation probability of the mutant. Our work lays the foundation for also studying density- and frequency-dependent effects in populations when demographic structures cannot be neglected.

  19. Antisense downregulation of mutant huntingtin in a cell model

    DEFF Research Database (Denmark)

    Hasholt, L.; Abell, K.; Norremolle, A.

    2003-01-01

    or by addition to the culture medium. Results Expression of the fusion protein containing the mutant huntingtin fragment resulted in diffuse green fluorescence in the cytoplasm and formation of aggregates in some of the NT2 cells and NT2-N neurons. We obtained antisense sequence-specific inhibition of expression...... of specific neurons in the brains of HD patients correlate with the expression of mutant huntingtin. Therefore, we have studied whether mutant huntingtin expression can be downregulated by antisense technique. Methods NT2 precursor cells and differentiated postmitotic NT2-N neurons, respectively, were...... transfected with plasmid constructs containing exon 1 of the HD gene with expanded CAG repeats in frame with the reporter protein EGFP. The transfected cell cultures were treated with a phosphorothioated antisense oligonucleotide (PS-ASHD/20+) or a control oligonucleotide either by cotransfection...

  20. Human mutant huntingtin disrupts vocal learning in transgenic songbirds.

    Science.gov (United States)

    Liu, Wan-Chun; Kohn, Jessica; Szwed, Sarah K; Pariser, Eben; Sepe, Sharon; Haripal, Bhagwattie; Oshimori, Naoki; Marsala, Martin; Miyanohara, Atsushi; Lee, Ramee

    2015-11-01

    Speech and vocal impairments characterize many neurological disorders. However, the neurogenetic mechanisms of these disorders are not well understood, and current animal models do not have the necessary circuitry to recapitulate vocal learning deficits. We developed germline transgenic songbirds, zebra finches (Taneiopygia guttata) expressing human mutant huntingtin (mHTT), a protein responsible for the progressive deterioration of motor and cognitive function in Huntington's disease (HD). Although generally healthy, the mutant songbirds had severe vocal disorders, including poor vocal imitation, stuttering, and progressive syntax and syllable degradation. Their song abnormalities were associated with HD-related neuropathology and dysfunction of the cortical-basal ganglia (CBG) song circuit. These transgenics are, to the best of our knowledge, the first experimentally created, functional mutant songbirds. Their progressive and quantifiable vocal disorder, combined with circuit dysfunction in the CBG song system, offers a model for genetic manipulation and the development of therapeutic strategies for CBG-related vocal and motor disorders.

  1. The use of mutants to probe models of gravitropism.

    Science.gov (United States)

    Firn, R D; Wagstaff, C; Digby, J

    2000-08-01

    It has been widely believed for more than 70 years that auxin plays a central role in the induction of differential growth which causes gravitropic curvature. However, this long-standing consensus about a role for auxin in gravitropism has only been achieved by allowing several mutually exclusive models to coexist. Furthermore, because there is no detailed model which is unchallenged by evidence, consensus is now centred on ill-defined models which have a low predictive value, hence are harder to challenge experimentally. An increasing number of mutants with abnormal gravitropic behaviour are becoming available. Such mutants should be very helpful in challenging existing models of gravitropism and in providing new evidence on which to build improved, more precise models. However, to date, most studies of mutants with abnormal gravitropism have been guided, experimentally and conceptually, by the old inadequate and vague models. Consequently, the full potential of modern molecular analysis in aiding our understanding of gravitropism has yet to be realized.

  2. Candida albicans mutant construction and characterization of selected virulence determinants.

    Science.gov (United States)

    Motaung, T E; Albertyn, J; Pohl, C H; Köhler, Gerwald

    2015-08-01

    Candida albicans is a diploid, polymorphic yeast, associated with humans, where it mostly causes no harm. However, under certain conditions it can cause infections ranging from superficial to life threatening. This ability to become pathogenic is often linked to the immune status of the host as well as the expression of certain virulence factors by the yeast. Due to the importance of C. albicans as a pathogen, determination of the molecular mechanisms that allow this yeast to cause disease is important. These studies rely on the ability of researchers to create deletion mutants of specific genes in order to study their function. This article provides a critical review of the important techniques used to create deletion mutants in C. albicans and highlights how these deletion mutants can be used to determine the role of genes in the expression of virulence factors in vitro.

  3. How does the NPM1 mutant induce leukemia?

    Directory of Open Access Journals (Sweden)

    Paolo Sportoletti

    2011-06-01

    Full Text Available NPM1 is the most frequently mutated gene in AML and the role of the NPM1 mutant in acute myeloid leukemia along with its leukemogenic potential are still under investigation. NPM1 genetic alterations can contribute to leukemogenesis through the direct oncogenic effect of the mutant protein and the concomitant loss of one functional allele. Npm1 loss determines tumor development in the mouse while in human NPM1 maps in a chromosomal region frequently loss in myelodysplastic syndrome (MDS. The NPM1 mutant cytoplasmic delocalization in leukemic blasts alters multiple cellular pathways through either loss or gain of function effects on different protein partners. Here we discuss the most relevant studies on the role of the NPM1 molecule in hematological malignancies and both in vitro and in vivo studies that are trying to elucidate the way by which the NPM1 mutation induces leukemia.

  4. Antisense downregulation of mutant huntingtin in a cell model

    DEFF Research Database (Denmark)

    Hasholt, L.; Abell, K.; Norremolle, A.;

    2003-01-01

    of specific neurons in the brains of HD patients correlate with the expression of mutant huntingtin. Therefore, we have studied whether mutant huntingtin expression can be downregulated by antisense technique. Methods NT2 precursor cells and differentiated postmitotic NT2-N neurons, respectively, were...... transfected with plasmid constructs containing exon 1 of the HD gene with expanded CAG repeats in frame with the reporter protein EGFP. The transfected cell cultures were treated with a phosphorothioated antisense oligonucleotide (PS-ASHD/20+) or a control oligonucleotide either by cotransfection...... or by addition to the culture medium. Results Expression of the fusion protein containing the mutant huntingtin fragment resulted in diffuse green fluorescence in the cytoplasm and formation of aggregates in some of the NT2 cells and NT2-N neurons. We obtained antisense sequence-specific inhibition of expression...

  5. Milder clinical and biochemical phenotypes associated with the c.482G>A (p.Arg161Gln) pathogenic variant in cobalamin C disease: Implications for management and screening.

    Science.gov (United States)

    Almannai, Mohammed; Marom, Ronit; Divin, Kristian; Scaglia, Fernando; Sutton, V Reid; Craigen, William J; Lee, Brendan; Burrage, Lindsay C; Graham, Brett H

    2017-09-01

    Cobalamin C disease is a multisystemic disease with variable manifestations and age of onset. Genotype-phenotype correlations are well-recognized in this disorder. Here, we present a large cohort of individuals with cobalamin C disease, several of whom are heterozygous for the c.482G>A pathogenic variant (p.Arg161Gln). We compared clinical characteristics of individuals with this pathogenic variant to those who do not have this variant. To our knowledge, this study represents the largest single cohort of individuals with the c.482G>A (p.Arg161Gln) pathogenic variant. A retrospective chart review of 27 individuals from 21 families with cobalamin C disease who are followed at our facility was conducted. 13 individuals (48%) are compound heterozygous with the c.482G>A (p.Arg161Gln) on one allele and a second pathogenic variant on the other allele. Individuals with the c.482G>A (p.Arg161Gln) pathogenic variant had later onset of symptoms and easier metabolic control. Moreover, they had milder biochemical abnormalities at presentation which likely contributed to the observation that 4 individuals (31%) in this group were missed by newborn screening. The c.482G>A (p.Arg161Gln) pathogenic variant is associated with milder disease. These individuals may not receive a timely diagnosis as they may not be identified on newborn screening or because of unrecognized, late onset symptoms. Despite the milder presentation, significant complications can occur, especially if treatment is delayed. Copyright © 2017 Elsevier Inc. All rights reserved.

  6. Probing the interaction of human serum albumin with vitamin B2 (riboflavin) and L-Arginine (L-Arg) using multi-spectroscopic, molecular modeling and zeta potential techniques

    Energy Technology Data Exchange (ETDEWEB)

    Memarpoor-Yazdi, Mina [Department of Biology, Faculty of Sciences, Mashhad Branch, Islamic Azad University, Mashhad (Iran, Islamic Republic of); Mahaki, Hanie, E-mail: hanieh.mahaki@gmail.com [Department of Biology, Faculty of Sciences, Mashhad Branch, Islamic Azad University, Mashhad (Iran, Islamic Republic of)

    2013-04-15

    This study was designed to examine the interaction of Riboflavin (RB) and L-Arginine (L-Arg) with human serum albumin (HSA) using different spectroscopic, zeta potential and molecular modeling techniques under imitated physiological conditions. The resonance light scattering (RLS) method determined the critical aggregation concentration of RB on HSA in the presence and absence of L-Arg which confirmed the zeta potential results. The binding constants (K{sub a}) of HSA–RB were 2.5×10{sup 4} and 9.7×10{sup 3} M{sup −1}, respectively in binary and ternary system at the excitation wavelength of 280 nm, also were 7.5×10{sup 3} and 7.3×10{sup 3}, respectively in binary and ternary system at the excitation wavelength of 295 nm. Fluorescence spectroscopy demonstrated that in the presence of L-Arg, the binding constant of HSA–RB was increased. Static quenching was confirmed to results in the fluorescence quenching and FRET. The binding distances between HSA and RB in two- and three-component systems were estimated by the Forster theory which revealed that nonradiative energy transfer from HSA to RB occurred with a high probability. The effect of RB on the conformation of HSA was analyzed using synchronous fluorescence spectroscopy and circular dichroism (CD) in both systems. Docking studies demonstrated a reduction in the binding affinity between RB and HSA in the presence of L-Arg. -- Highlights: ► We studied the interaction of riboflavin with HSA in presence and absence of L-Arg. ► Molecular modeling and zeta-potential used to describe competitive interaction. ► We compared the binding mechanism of riboflavin (RB) to HSA in both systems. ► We determined critical aggregation concentration of RB on HSA in both systems. ► The binding site of RB on HSA in both systems has been determined.

  7. 广西壮族人群CYP1B1基因Arg48Gly和Ala119Ser多态性研究%Study on gene polymorphisms of CYP1B1 Arg48Gly and Ala119Ser among Guangxi Zhuang Nationality population

    Institute of Scientific and Technical Information of China (English)

    李曙波; 廖长秀; 张梁; 李珊; 许小林; 罗莹

    2013-01-01

    Objective To investigate hereditary character of cytochrome P450 1B1 (CYP1B1) codon 48 and codon 119 among Guangxi Zhuang Nationality population (GZNP). Methods Gene polymorphisms of CYP1B1 in codon 48 and codon 119 were detected with polymerase chain reaction -restriction fragment length polymorphism techniques in 278 healthy GZNP. Linkage disequilibrium (LD) between the two loci was analyzed. The genotype frequencies of CYP1B1 codon 48 and codon 119 in GZNP were compared with other groups in China, other countries and regions. Results There was no difference between female and male population in allele and genotype frequencies of CYP1B1 Arg48Gly and Alall9Ser among GZNP. In the other hand, there was LD between the two loci (D' =0. 81, r2= 0.59). The frequency distribution of genotype in GZNP had no great differences with Shanghai and Sichuan, but those had significant differences with other contries such as Japan, India, Poland, and America (P <0. 01). Conclusion Gene polymorphisms of CYP1B1 in Arg48Gly and Alal 19Ser had LD, no gender dif