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Sample records for biuret trimer hdi-bt

  1. Exposure, lung function, and symptoms in car painters exposed to hexamethylendiisocyanate and biuret modified hexamethylendiisocyanate

    Energy Technology Data Exchange (ETDEWEB)

    Alexandersson, R.; Hedenstierna, G.; Plato, N.; Kolmodin-Hedman, B.

    1987-11-01

    Individuals who paint cars often complain to doctors about respiratory problems. Car painters are exposed to isocyanates, especially hexamethylendiisocyanate (HDI), and biuret modified HDI (HDI-BT). The mean exposure to HDI-BT was 115 micrograms/m3 in the air (range 10-385 micrograms/m3), which exceeds the time-weighted Swedish threshold level of 90 micrograms/m3. Exposure to HDI was about 1.0 microgram/m3 with brief peaks. This study investigated the effect of HDI and HDI-BT on lung function and included two control groups: (1) car platers, exposed to the same solvents and grinding dust as car painters, but not to isocyanates, and (2) car mechanics (controls), not exposed to the mentioned agents. Car painters and car platers were compared to car mechanics on Monday before work. Acute effects of car painting were tested by comparing the lung function values on Monday morning with those on Friday afternoon. Pulmonary function was evaluated by means of spirometry and a single breath nitrogen washout. Spirometry in painters and platers did not differ from that in controls, i.e., car mechanics. Closing volume in relation to vital capacity (CV%) was increased in car painters, suggestive of a small airways disease on Monday before work and tended to increase during a work week. Car platers did not differ from controls.

  2. Neuronal modulation of lung injury induced by polymeric hexamethylene diisocyanate in mice

    International Nuclear Information System (INIS)

    Lee, C.-T.; Poovey, Halet G.; Rando, Roy J.; Hoyle, Gary W.

    2007-01-01

    1,6-Hexamethylene diisocyanate biuret trimer (HDI-BT) is a nonvolatile isocyanate that is a component of polyurethane spray paints. HDI-BT is a potent irritant that when inhaled stimulates sensory nerves of the respiratory tract. The role of sensory nerves in modulating lung injury following inhalation of HDI-BT was assessed in genetically manipulated mice with altered innervation of the lung. Knockout mice with a mutation in the low-affinity nerve growth factor receptor (NGFR), which have decreased innervation by nociceptive nerve fibers, and transgenic mice expressing nerve growth factor (NGF) from the lung-specific Clara cell secretory protein (CCSP) promoter, which have increased innervation of the airways, were exposed to HDI-BT aerosol and evaluated at various times after exposure. NGFR knockout mice exhibited significantly more, and CCSP-NGF transgenic mice exhibited significantly less injury and inflammation compared with wild-type mice, indicative of a protective effect of nociceptive nerves on the lung following HDI-BT inhalation. Transgenic mice overexpressing the tachykinin 1 receptor (Tacr1) in lung epithelial cells also showed less severe injury and inflammation compared with wild-type mice after HDI-BT exposure, establishing a role for released tachykinins acting through Tacr1 in mediating at least part of the protective effect. Treatment of lung fragments from Tacr1 transgenic mice with the Tacr1 ligand substance P resulted in increased cAMP accumulation, suggesting this compound as a possible signaling mediator of protective effects on the lung following nociceptive nerve stimulation. The results indicate that sensory nerves acting through Tacr1 can exert protective or anti-inflammatory effects in the lung following isocyanate exposure

  3. Gene expression profiling in mouse lung following polymeric hexamethylene diisocyanate exposure

    International Nuclear Information System (INIS)

    Lee, C.-T.; Ylostalo, Joni; Friedman, Mitchell; Hoyle, Gary W.

    2005-01-01

    Isocyanates are a common cause of occupational lung disease. Hexamethylene diisocyanate (HDI), a component of polyurethane spray paints, can induce respiratory symptoms, inflammation, lung function impairment, and isocyanate asthma. The predominant form of HDI in polyurethane paints is a nonvolatile polyisocyanate known as HDI biuret trimer (HDI-BT). Exposure of mice to aerosolized HDI-BT results in pathological effects, including pulmonary edema, lung inflammation, cellular proliferation, and fibrotic lesions, which occur with distinct time courses following exposure. To identify genes that mediate lung pathology in the distinct temporal phases after exposure, gene expression profiles in HDI-BT-exposed C57BL/6J mouse lungs were analyzed. RNase protection assay (RPA) of genes involved in apoptosis, cell survival, and inflammation revealed increased expression of IκBα, Fas, Bcl-X L , TNFα, KC, MIP-2, IL-6, and GM-CSF following HDI-BT exposure. Microarray analysis of approximately 10 000 genes was performed on lung RNA collected from mice 6, 18, and 90 h after HDI-BT exposure and from unexposed mice. Classes of genes whose expression was increased 6 h after exposure included those involved in stress responses (particularly oxidative stress and thiol redox balance), growth arrest, apoptosis, signal transduction, and inflammation. Types of genes whose expression was increased at 18 h included proteinases, anti-proteinases, cytoskeletal molecules, and inflammatory mediators. Transcripts increased at 90 h included extracellular matrix components, transcription factors, inflammatory mediators, and cell cycle regulators. This characterization of the gene expression profile in lungs exposed to HDI-BT will provide a basis for investigating injury and repair pathways that are operative during isocyanate-induced lung disease

  4. 21 CFR 573.220 - Feed-grade biuret.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Feed-grade biuret. 573.220 Section 573.220 Food... Additive Listing § 573.220 Feed-grade biuret. The food additive feed grade biuret may be safely used in... resulting from the controlled pyrolysis of urea conforming to the following specifications: Percent Biuret...

  5. Microbial biodegradation of biuret: defining biuret hydrolases within the isochorismatase superfamily.

    Science.gov (United States)

    Robinson, Serina L; Badalamenti, Jonathan P; Dodge, Anthony G; Tassoulas, Lambros J; Wackett, Lawrence P

    2018-03-12

    Biuret is a minor component of urea fertilizer and an intermediate in s-triazine herbicide biodegradation. The microbial metabolism of biuret has never been comprehensively studied. Here, we enriched and isolated bacteria from a potato field that grew on biuret as a sole nitrogen source. We sequenced the genome of the fastest-growing isolate, Herbaspirillum sp. BH-1 and identified genes encoding putative biuret hydrolases (BHs). We purified and characterized a functional BH enzyme from Herbaspirillum sp. BH-1 and two other bacteria from divergent phyla. The BH enzymes reacted exclusively with biuret in the range of 2-11 µmol min -1 mg -1 protein. We then constructed a global protein superfamily network to map structure-function relationships in the BH subfamily and used this to mine > 7000 genomes. High-confidence BH sequences were detected in Actinobacteria, Alpha- and Beta-proteobacteria, and some fungi, archaea and green algae, but not animals or land plants. Unexpectedly, no cyanuric acid hydrolase homologs were detected in > 90% of genomes with BH homologs, suggesting BHs may have arisen independently of s-triazine ring metabolism. This work links genotype to phenotype by enabling accurate genome-mining to predict microbial utilization of biuret. Importantly, it advances understanding of the microbial capacity for biuret biodegradation in agricultural systems. © 2018 Society for Applied Microbiology and John Wiley & Sons Ltd.

  6. Biuret, a NPN source for ruminants -- a review

    International Nuclear Information System (INIS)

    Singhal, K.K.; Mudgal, V.D.

    1980-01-01

    Biuret is a non toxic and palatable source of non protein nitrogen which is hydrolyzed slowly and has a slower solubility as compared to urea. It is hydrolyzed in rumen with the help of biuretase which is an induced enzyme, therefore animals require some time of adaptation before its proper utilization. Biuret is degraded into ammonia and urea which later on gets hydrolysed further into ammonia with the help of urease. The activity of urease might have been supressed in the rumen when the animals were fed on biuret supplemented diet. Biuret degradation into ammonia increases as the adaptation reaches. Biuret hydrolyzed slowly upto 24 hr of its feeding in adapted animals. For the best utilization of this NPN compound, the diet should contain low levels of natural protein and sufficient quantity of readily available source of energy and provide adequate essential minerals. Biuret can be successfully used for the maintenance and production ratio of the ruminants. (author)

  7. The measurement of total serum proteins by the Biuret method.

    Science.gov (United States)

    Lubran, M M

    1978-01-01

    The biuret reaction for proteins provides a simple and precise method for measuring serum proteins; Beer's law is obeyed to at least 10 g per dl. Several stable biuret reagents are available. Hemoglobin is the only important cause of interference which cannot be minimized by use of a sample blank. The mechanism of the biuret reaction is described and attention is drawn to the heterogeneity of the serum proteins and to the use of a certified albumin standard.

  8. Metformin reactivity with biuret: application to tablets

    OpenAIRE

    Chahin, Cecilia L.; Delfino, Mario R.; Sarno, María del C.

    2011-01-01

    A spectrophotometric method was developed for the analysis of metformin, based on its reaction with the biuret reagent. A complex with λmax = 495 nm and stoichiometry 2:1 (metformin-Cu) is generated wich allows the quantification of metformin in tablets. Physicochemical parameters of the product were determined. Sample conditioning by centrifugation was necessary to separate insoluble excipients. Polyvinylpyrrolidone (PVP) was isolated by a solid-liquid extraction with SPE-SCX resin in o...

  9. 1-Benzoyl-4-thio-biuret.

    Science.gov (United States)

    Kang, Sung Kwon

    2013-01-01

    {[(phenyl-formamido)-carbon-yl]amino}-methane-thio-amide), C9H9N3O2S, both benzoyl and terminal thio-urea fragments adopt transoid conformations with respect to the central carbonyl O atom. The benzoyl and thio-biuret groups are almost coplanar, making a dihedral angle of 4.40 (8)°. The mol-ecular structure is stabilized by two intra-molecular N-H⋯O hydrogen bonds. In the crystal, N-H⋯O and N-H⋯S hydrogen bonds link the mol-ecules into a tape running along [101].

  10. 1-Benzoyl-2-thio-biuret.

    Science.gov (United States)

    Kang, Sung Kwon; Cho, Nam Sook; Jeon, Min Kyeong

    2012-02-01

    IN THE TITLE COMPOUND [SYSTEMATIC NAME: N-(carbamoyl-carb-a-mo-thio-yl)benzamide], C(9)H(9)N(3)O(2)S, the benzoyl and terminal urea fragments adopt cisoid and transoid conformations, respectively, with respect to the S atom. The benzoyl and thio-biuret groups are almost coplanar, making a dihedral angle of 8.48 (5)°. The mol-ecular structure is stabilized by an intra-molecular N-H⋯O hydrogen bond. In the crystal, N-H⋯O and N-H⋯S hydrogen bonds link the mol-ecules into a sheet parallel to the bc plane.

  11. [Application research of protein test by using biuret reagent].

    Science.gov (United States)

    Chen, Ningqing; Zheng, Siyu

    2014-11-01

    To investigate the biuret reagent to detect proteins in the application, the impact of different test conditions for test results. The biuret method to select three different instruments, reagents, calibrators are arranged in combination to form 27 sets of detection systems, each detection system is a combination of 5 serum samples for testing, 5 measured values obtained, the selection process normality good a serum for the study to determine the mean value of all AST after culling outliers obtained in order to calculate the various detection systems use a combination of biuret reagent to detect proteins bias. The use of different detection equipment to detect proteins biuret reagent bias, homogeneity of variance (P = 0.467), the difference was not statistically significant (F = 1.688, P = 0.421). different detection reagents using biuret reagent to detect proteins bias, homogeneity of variance (P = 0.574), a statistically significant difference (F = 5.784, P = 0.011). different calibrators use biuret reagent to detect proteins bias, homogeneity of variance (P = 0.467), the difference was statistically significant (F = 5.289, P = 0.000). Biuret reagent in the detection of protein applications, impact detection reagents and calibrators will test result, during the test than when it is necessary to detect deviation detection reagents and calibrators due to be considered.

  12. 1-Benzoyl-4-thio­biuret

    Science.gov (United States)

    Kang, Sung Kwon

    2013-01-01

    In the title compound (systematic name: {[(phenyl­formamido)­carbon­yl]amino}­methane­thio­amide), C9H9N3O2S, both benzoyl and terminal thio­urea fragments adopt transoid conformations with respect to the central carbonyl O atom. The benzoyl and thio­biuret groups are almost coplanar, making a dihedral angle of 4.40 (8)°. The mol­ecular structure is stabilized by two intra­molecular N—H⋯O hydrogen bonds. In the crystal, N—H⋯O and N—H⋯S hydrogen bonds link the mol­ecules into a tape running along [101]. PMID:24109399

  13. 1-Benzoyl-2-thio­biuret

    Science.gov (United States)

    Kang, Sung Kwon; Cho, Nam Sook; Jeon, Min Kyeong

    2012-01-01

    In the title compound [systematic name: N-(carbamoyl­carb­a­mo­thio­yl)benzamide], C9H9N3O2S, the benzoyl and terminal urea fragments adopt cisoid and transoid conformations, respectively, with respect to the S atom. The benzoyl and thio­biuret groups are almost coplanar, making a dihedral angle of 8.48 (5)°. The mol­ecular structure is stabilized by an intra­molecular N—H⋯O hydrogen bond. In the crystal, N—H⋯O and N—H⋯S hydrogen bonds link the mol­ecules into a sheet parallel to the bc plane. PMID:22347017

  14. Synthesis and Characterization of Polyurethanes With High Content of Biuret Groups

    OpenAIRE

    Šebenik, A.

    1987-01-01

    An 'H-NMR and 13C NMR study of the reaction between urea and 4,4'-diphenylmethanedisocyanate is reported. Assignments for individual signals of biuret groups were determined and their concentration dependence on the reaction time was established. The rate constants for different biuret oligomers were calculated. From biuret oligomers and urethane oligomers block cooligomers and later block copolymers were formed.

  15. Preparation of guanine and diaminopurine from biuret. Part III.

    Science.gov (United States)

    Lagoja, Irene M; Herdewijn, Piet

    2007-04-01

    Because of their potential prebiotic origin and relative chemical stability, urea, biuret, formic acid, and glycine amide might have played a role in the assembly process of purine bases. In this paper, we describe a short reaction path to purine nucleobases from these acyclic precursors. The formation of different purines was verified by UV and NMR spectroscopy, as well as by mass spectrometry.

  16. A New Family of Biuret Hydrolases Involved in S-Triazine Ring Metabolism.

    Science.gov (United States)

    Cameron, Stephan M; Durchschein, Katharina; Richman, Jack E; Sadowsky, Michael J; Wackett, Lawrence P

    2011-08-01

    Biuret is an intermediate in the bacterial metabolism of s-triazine ring compounds and is occasionally used as a ruminant feed supplement. We used bioinformatics to identify a biuret hydrolase, an enzyme that has previously resisted efforts to stabilize, purify and characterize. This newly discovered enzyme is a member of the cysteine hydrolase superfamily, a family of enzymes previously not found to be involved in s-triazine metabolism. The gene from Rhizobium leguminosarum bv. viciae strain 3841 encoding biuret hydrolase was synthesized, transformed into Escherichia coli, and expressed. The enzyme was purified and found to be stable. Biuret hydrolase catalyzed the hydrolysis of biuret to allophanate and ammonia. The k(cat)/K(M) of 1.7 × 10(5) M(-1)s(-1) and the relatively low K(M) of 23 ± 4 μM together suggested that this enzyme acts uniquely on biuret physiologically. This is supported by the fact that of the 34 substrate analogs of biuret tested, only two demonstrated reactivity, both at less than 5% of the rate determined for biuret. Biuret hydrolase does not react with carboxybiuret, the product of the enzyme immediately preceding biuret hydrolase in the metabolic pathway for cyanuric acid. This suggests an unusual metabolic strategy of an enzymatically-produced intermediate undergoing non-enzymatic decarboxylation to produce the substrate for the next enzyme in the pathway.

  17. Purification and characterization of TrzF: biuret hydrolysis by allophanate hydrolase supports growth.

    Science.gov (United States)

    Shapir, Nir; Cheng, Gang; Sadowsky, Michael J; Wackett, Lawrence P

    2006-04-01

    TrzF, the allophanate hydrolase from Enterobacter cloacae strain 99, was cloned, overexpressed in the presence of a chaperone protein, and purified to homogeneity. Native TrzF had a subunit molecular weight of 65,401 and a subunit stoichiometry of alpha(2) and did not contain significant levels of metals. TrzF showed time-dependent inhibition by phenyl phosphorodiamidate and is a member of the amidase signature protein family. TrzF was highly active in the hydrolysis of allophanate but was not active with urea, despite having been previously considered a urea amidolyase. TrzF showed lower activity with malonamate, malonamide, and biuret. The allophanate hydrolase from Pseudomonas sp. strain ADP, AtzF, was also shown to hydrolyze biuret slowly. Since biuret and allophanate are consecutive metabolites in cyanuric acid metabolism, the low level of biuret hydrolase activity can have physiological significance. A recombinant Escherichia coli strain containing atzD, encoding cyanuric acid hydrolase that produces biuret, and atzF grew slowly on cyanuric acid as a source of nitrogen. The amount of growth produced was consistent with the liberation of 3 mol of ammonia from cyanuric acid. In vitro, TrzF was shown to hydrolyze biuret to liberate 3 mol of ammonia. The biuret hydrolyzing activity of TrzF might also be physiologically relevant in native strains. E. cloacae strain 99 grows on cyanuric acid with a significant accumulation of biuret.

  18. The effect of applying biuret to certain grasses | RE | African Journal ...

    African Journals Online (AJOL)

    The effects of applications of biuret to Digitaria swazilandenis, Cynodon dactylon, Eragrostis curvula and sugarcane are described. Slight leaf tip-burn was observed in the cases of the Digitaria, Cynodon and Eragrostis a few days after application of the biuret but sugarcane was not affected. The results show that crude ...

  19. Synthesis and cytotoxicity of some biurets against human breast cancer T47D cell line.

    Science.gov (United States)

    Fouladdel, Shamileh; Khalaj, Ali; Adibpour, Neda; Azizi, Ebrahim

    2010-10-01

    Design, synthesis and cytotoxicity of several known and novel biurets against human breast cancer T47D cell line in comparison to doxorubicin are described. Biurets incorporating 2-methyl quinoline-4-yl and benzo[d]thiazol-2-ylthio moieties showed higher cytotoxicity and decreased cell viability in a concentration- and time-dependent manner. Copyright © 2010 Elsevier Ltd. All rights reserved.

  20. Enthalpy of solution of biuret in various aqueous electrolyte solutions and in an urea solution

    Energy Technology Data Exchange (ETDEWEB)

    Nishimura, Hisashi; Murakami, Sachio (Osaka City Univ. (Japan))

    1989-06-30

    Enthalpies of biuret which is simplest amido acid and does not have hydrophobic group, in various aqueous solutions and in urea solution were measured at 298.15K to clarify the influence of salt on polymer conformal changes of organism or structure and functions of membrane. The isoperibol calorimeter where the quartz thermometer was used as the thermal sensor, was used in the measurement. The performance of this calorimeter was tested by comparing measurements with values in references. From the analysis of enthalpies of biuret solutions, following results of solute-solute-solvent interactions could be obtained: the rate where biuret destroys water structure is smaller in salt solutions than in aqueous solutions and the tendency depends on the ion size; ion-water interaction weakens owing to the ion-biuret interaction in salt solution of biuret; and water molecules are made free. 25 refs., 4 figs., 5 tabs.

  1. [The determination of protein content of milk, cheese, and meat with the use of the biuret reaction].

    Science.gov (United States)

    Reichardt, W; Eckert, W

    1991-01-01

    A review is given of the different composition of biuret test solutions and experiments with the estimation of protein content of milk, cheese and meat by means of the biuret method are reported. The pure protein and the casein content of skimmed milk and the protein content of whole milk could be determined with biuret test solutions containing potassium hydroxide and a detergent. The use of the biuret method is advantageous in the case of cheese and meat, which are dissolved in potash lye or alkaline detergent solutions. Disturbances of the biuret method caused by lactose, fat or turbidity were eliminated after addition of hydrogen peroxide, by means of extraction or of additional measurements with a copper-free, zinc-containing biuret reagent. Deviations of the color intensity of biuret complexes were compensated by inclusion of standards in the measurement series.

  2. Assay of cerebrospinal fluid protein: a rate biuret method evaluated.

    Science.gov (United States)

    Finley, P R; Williams, R J

    1983-01-01

    We evaluated a rate colorimetric method (Beckman) for measuring total protein in cerebrospinal fluid. The automated instrument we used was Beckman's ASTRA TM. A 100-microL sample of spinal fluid is introduced into the biuret reagent in the reaction cell and the increase in absorbance at 545 nm is monitored for 20.5 s. Solid-state circuits determine the rate of alkaline biuret-protein chelate formation, which is directly proportional to the total protein concentration in the sample. The linear range of measurement is 120 to 7500 mg/L. Day-to-day precision (CV) over the range of 150 to 1200 mg/L ranged from 15.2 to 2.3%. The method was unaffected by radical alteration of the albumin/globulin ratio, but there is a positive interference in the presence of hemoglobin, a suppression in the presence of bilirubin, and no effect by xanthochromia. The method is precise, accurate, rapid, and convenient. The method was compared with the trichloroacetic acid method as performed on the Du Pont aca III, giving a correlation coefficient (r2) of 0.9693. The method is precise, accurate, rapid, and convenient.

  3. The influence of biuret on the growth kinetics of urea crystals from aqueous solutions

    Science.gov (United States)

    Davey, R.; Fila, W.; Garside, J.

    1986-12-01

    The growth kinetics of urea in the [001] and [110] directions are presented for both pure solutions and solutions containing up to 6% biuret. This impurity is formed during the synthesis of urea and acts as a powerful habit modifier. In pure solutions urea grows very quickly in the [001] but very slowly in the [110] direction. As a result long needle-like crystals are produced. In the presence of biuret [001] growth is dramatically reduced while [110] growth is virtually unaffected, so producing a crystal with a much lower length: breadth ratio. The influence of biuret is explained by the replacement of two urea molecules in the crystal lattice by a biuret molecule. The resulting disruption of the (001) face leaves insufficient NH 2-groups to satisfy the hydrogen bonding linkages.

  4. A New Family of Biuret Hydrolases Involved in S-Triazine Ring Metabolism

    OpenAIRE

    Cameron, Stephan M.; Durchschein, Katharina; Richman, Jack E.; Sadowsky, Michael J.; Wackett, Lawrence P.

    2011-01-01

    Biuret is an intermediate in the bacterial metabolism of s-triazine ring compounds and is occasionally used as a ruminant feed supplement. We used bioinformatics to identify a biuret hydrolase, an enzyme that has previously resisted efforts to stabilize, purify and characterize. This newly discovered enzyme is a member of the cysteine hydrolase superfamily, a family of enzymes previously not found to be involved in s-triazine metabolism. The gene from Rhizobium leguminosarum bv. viciae strain...

  5. Comparison of biuret and refractometry methods for the serum total proteins measurement in ruminants.

    Science.gov (United States)

    Katsoulos, Panagiotis D; Athanasiou, Labrini V; Karatzia, Maria A; Giadinis, Nektarios; Karatzias, Harilaos; Boscos, Constantin; Polizopoulou, Zoe S

    2017-12-01

    Determination of serum total protein concentration is commonly performed by the biuret method. Refractometric measurement is a faster and less expensive alternative but its accuracy has not been determined in ruminants. The purpose of the study was to compare the serum total protein concentrations in cattle, sheep, and goats measured by the biuret method with those obtained by refractometry. Serum total protein concentration was determined in 120 cattle, 67 sheep, and 58 goat blood samples refractometrically and with the biuret method. The data were analyzed with a paired samples t-test, and Passing and Bablok regression equations and Bland and Altman plots were generated. There was a strong linear relationship between the total protein values determined with the refractometer and the biuret method in cattle, sheep, and goats. The statistical accuracy, which represents a bias correction factor that measures the deviation of the best-fit line from the 45° line through the origin, was 90.63% for cattle, 93.05% for sheep, and 91.76% for goats. The mean protein values determined with the refractometer were significantly lower than those measured with the biuret method in cattle and goats (P  .05). The evaluated refractometer was sufficiently accurate for the determination of serum total proteins in cattle, sheep, and goats, although it cannot be used interchangeably with the biuret method. The RIs should be corrected for negative bias based on the created equations. © 2017 American Society for Veterinary Clinical Pathology.

  6. Hydrogen-Bonding Modification in Biuret Under Pressure.

    Science.gov (United States)

    Borstad, Gustav M; Ciezak-Jenkins, Jennifer A

    2017-02-02

    Biuret (C 2 H 5 N 3 O 2 ) has been studied to 30 GPa by Raman spectroscopy and 50 GPa by X-ray diffraction. Raman peaks exhibit shoulders and splitting that suggests that the molecules undergo reorientation in response to compression. These are observed in three pressure ranges: the first from 3-5 GPa, the second from 8-12 GPa, and finally from 16-20 GPa. The particular modes in the sample that are observed to change in the Raman are strongly linked to the molecular vibrations involving the N-H and the C═O bond, which are most strongly coupled to the hydrogen-bonded lattice structure. The X-ray diffraction suggests that the crystal maintains a monoclinic structure to the highest pressures studied. Although there was a considerable degree of hysteresis observed in some X-ray runs, all the changes observed under pressure are reversible.

  7. The effects of isomerism on the vibrational spectra and thermodynamic characteristics of biuret in the gas phase

    Science.gov (United States)

    Korolevich, M. V.; Lastochkina, V. A.; Frenkel, M. L.; Kabo, G. Ya.; Zhbankov, R. G.

    1991-03-01

    A theoretical study of the IR spectra of cis- and trans-isomers of biuret, based on coupled calculations of vibrational frequencies as a valence force field approximation and of absorption band intensities by the CNDO/2 method, is reported. Calculated thermodynamic functions for biuret agreed with experimental thermochemical data. For calculating the vibrations of an isolated biuret molecule, the experimental vapour-phase IR absorption spectra were used.

  8. Cross-reactivity of amino acids and other compounds in the biuret reaction: interference with urinary peptide measurements.

    Science.gov (United States)

    Hortin, Glen L; Meilinger, Bonnie

    2005-08-01

    Biuret assays for total protein measurement are considered to react with all peptides longer than 2 residues. Some studies using biuret assays of urine suggest that small peptides generally are more abundant than proteins in urine, but it is not clear whether this is a problem of assay specificity. We analyzed the specificity and kinetics of a biuret reaction for solutions of amino acids, organic compounds, peptides, proteins, and ultrafiltered urine specimens and compared the results with standard clinical assays for protein measurement. The biuret assay cross-reacted with several amino acids, dipeptides, and other organic compounds able to form 5- or 6-member ring chelation complexes with copper. Reactions with amino acids and dipeptides had higher absorbance maxima (blue color) than with larger peptides and proteins (purple). Compounds forming potential 4-, 7-, 8-, or 9-member ring complexes with copper had low reactivity. Amino acid amides, dipeptides, and longer peptides had substantial reactivity, except those containing proline. Proteins and polypeptides had similar biuret reactivities per peptide bond, but reaction kinetics were slower for proteins than peptides. Urine specimens ultrafiltered through 3-kDa-cutoff membranes had substantial biuret reactivity, but absorbance maxima were consistent with cross-reactive amino acids rather than peptides. Many compounds, including amino acids, amino acid derivatives, and dipeptides, cross-react in biuret assays. Our studies improve understanding of the specificity of endpoint and kinetic biuret assays widely used in clinical laboratories. Amino acids, urea, and creatinine contribute to overestimation of urinary peptide content by biuret assays.

  9. Penentuan Kadar Biuret, H2O, Dan Total Nitrogen Pada Urea Prill Dari Bulk Storage Di PT. Pupuk Iskandar Muda Lhokseumawe

    OpenAIRE

    Suriani, Evie A.S.

    2011-01-01

    Urea merupakan salah satu bentuk penggunaan nitrogen sebagai pupuk. Urea dibentuk dari reaksi antara ammonia dengan karbondioksida pada suhu dan tekanan tertentu. Namun pada suhu yang lebih tinggi saat proses pembuatan urea akan terbentuk senyawa biuret. Jika pupuk urea mengandung kadar senyawa biuret yang cukup tinggi, maka senyawa biuret ini akan menjadi racun bagi tanaman. Urea juga bersifat hidroskopis yaitu sangat mudah menyerap air. Dari hasil penelitian ini akan diperoleh kadar biuret,...

  10. Chiral all-dielectric trimer nanoantenna

    Science.gov (United States)

    Ullah, Kaleem; Garcia-Camara, Braulio; Habib, Muhammad; Liu, Xuefeng; Krasnok, Alex; Lepeshov, Sergey; Hao, Jingjing; Liu, Juan; Yadav, N. P.

    2018-03-01

    Chirality is a property of certain molecules, materials or artificial nanostructures, which enables them to interact with the spin angular momentum of an incident light. This provides a different optical response, depending on the incident polarization, which gives rise to chiral optical spectroscopies. However, low-detection limits require an enhanced chiral light-matter interaction. Here, we propose a novel type of resonant chiral optical nanoantenna based on high-index dielectric (Cu2O) spherical nanoparticles arranged in a trimer geometry. We demonstrate both numerically and experimentally that this trimer nanoantenna exhibits resonantly enhanced optical dichroism and light hotspots, which are associated with this geometry. Moreover, we apply near-field optical microscopy to measure the near-field peculiarities of the proposed nanoantenna. The ability of changing the nanoantenna near field scattering by varying the light polarization is shown.

  11. Antinociceptive Effect of Some Biuret Derivatives on Formalin Test in Mice

    Directory of Open Access Journals (Sweden)

    Neda Adibpour

    2014-03-01

    Full Text Available Purpose: The current study was designed to investigate the antinociceptive effects of several biuret derivatives with N, N`-diphenyl, N-phenyl-N`-alkylphenyl, N,N`-bis alkylphenyl, 2-methylquinoline-4-yl, benzo[d]thiazol-2-ylthio and (1-phenyl-1H-tetrazol-5-ylthio substituents on the formalin-evoked pain in mice. Methods: Antinociceptive activity of the nine biurets derivatives were assessed at different doses in mice using formalin test and the results were compared with those of indomethacin(20 mg/kg and vehicle of the compounds. Area under the pain score curve against time (AUEC up to 60 minutes was used as the measure of pain behavior. Results: A rather good analgesic effect was seen for most of the tested biuret derivatives. Significant reduction in median AUEC0-5 minutes was observed at the doses of 50 and 25 mg/kg for biurets with either benzyl and 2-methylquinoline-4-yl (C8 or phenylethyl and benzo[d]thiazol-2-ylthio(C9 moieties, respectively(p-value<0.0044. Antinociceptive activities of compound C7 (with bis phenylropyl substituent, C8 and C9 during the late phase of formaldehyde-induced pain were comparable to that of indomethacin. Conclusion: Unlike indomethacin, the tested biuret compounds are able to induce antinociception in both phases of formalin test and could be considered comparable to indomethacin at the selected doses.

  12. Determination of chicken and turkey plasma and serum protein concentrations by refractometry and the biuret method.

    Science.gov (United States)

    Andreasen, C B; Latimer, K S; Kircher, I M; Brown, J

    1989-01-01

    Plasma and serum protein concentrations were determined in chickens and turkeys by refractometry (with human and veterinary refractometers) and by the biuret method. Chicken and turkey serum protein values were significantly lower than respective plasma protein values according to both methods. Refractometer readings for both plasma and serum correlated closely with the results of the biuret test (r2 = 0.72 to 0.97). These findings indicate that plasma and serum protein values may be determined accurately in chickens and turkeys with a handheld refractometer.

  13. Interference of dextran in biuret-type assays of serum proteins.

    Science.gov (United States)

    Delanghe, Joris R; Hamers, Nicole; Taes, Youri E; Libeer, Jean-Claude

    2005-01-01

    Dextran interference in biuret-type assays of total serum proteins was investigated in a Belgian National External Quality Assurance Survey with 256 participants. In vitro supplementation of therapeutic (10% Gentran 70) dextran concentrations showed a broadly varying (from 0 to 20%) negative interference. The analytical interference was found to depend on both the sodium hydroxide and tartrate concentrations in the reagent formulation. The dry chemistry biuret method was not affected by the dextran interference. In a number of cases, the effects observed may be of clinical importance. Both clinicians and laboratory staff should be aware of the persistence of this analytical problem.

  14. Improved biuret procedure for routine determination of urinary total proteins in clinical proteinuria.

    Science.gov (United States)

    Rice, E W

    1975-03-01

    This communication describes and evaluates an improved routine methodology for quantitating clinical proteinuria. Based on investigations of Piscator and of Savory et al., a modified Tsuchiya's reagent (ethanolic HCI-phosphotungstic acid) is used to precipitate proteins at 56 degrees C, followed by biuret spectrophotometry at 540 nm. The accuracy of the proposed procedure was assessed by comparisons with results obtained by using an ultrafiltration membrane that retains solutes with an average molecular weight in excess of 10 000 for separating of urinary proteins before they are measured with the biuret reaction. Precision of the method (coefficient of variation) is typically 2-3%.

  15. Tetranectin, a trimeric plasminogen-binding C-type lectin

    DEFF Research Database (Denmark)

    Holtet, T L; Graversen, Jonas Heilskov; Clemmensen, I

    1997-01-01

    -linking analysis and SDS-PAGE to be a homo-trimer in solution as are other known members of the collectin family of C-type lectins. Biochemical evidence is presented showing that an N-terminal domain encoded within exons 1 and 2 of the tetranectin gene is necessary and sufficient to govern subunit trimerization....

  16. Thermodynamics of alcohols and monosaccharides in aqueous solutions of biuret at 25/sup 0/C

    Energy Technology Data Exchange (ETDEWEB)

    Barone, G.; Castronuovo, G.; Del Vecchio, P.; Elia, V.; Tosto, M.T.

    1988-10-01

    The excess enthalpies of ternary aqueous solutions of biuret with four aliphatic alcohols or four isomeric pentoses have been determined by flow microcalorimetry at 25/sup 0/C. The coefficients of the virial expansion of the excess enthalpies have been evaluated and compared with those already reported for urea and thiourea with the same alcohols and pentoses used here. For biuret-alcohol systems, the cross coefficients are positive and depend on the length and branching of the alkyl chain of the alcohols. On the contrary, they are negative for biuret-pentose systems. Hence, biuret, as urea and thiourea, is able to differentiate the behavior of families of solutes characterized by the same functional groups. The results are interpreted in terms of the relative stabilities of the hydration cospheres of predominantly hydrophobic or predominantly hydrophilic cosolutes. However, hypotheses about the hydration cospheres of the saccharides, which are more complex than those used previously, seem to be necessary to rationalize their complete behavior in water.

  17. THE INTERACTION OF SULPHUR DIOXIDE WITH 2-IMIDAZOLIDINONE AND BIURET IN AQUEOUS SOLUTIONS

    Directory of Open Access Journals (Sweden)

    R. E. Khoma

    2016-04-01

    Full Text Available The interaction in «sulphur dioxide — 2-imidazolidinone (biuret — water» systems has been investigated by pH-metric titration method. The formation of molecular and anion complexes has been stated, their structure and stability depend on the nature and concentration of components in a solution as well as on temperature and electronic characteristics of complexes.

  18. Fe(III) complex of biuret-amide based macrocyclic ligand as peroxidase enzyme mimic.

    Science.gov (United States)

    Panda, Chakadola; Ghosh, Munmun; Panda, Tamas; Banerjee, Rahul; Sen Gupta, Sayam

    2011-07-28

    An Fe(III) complex of a biuret-amide based macrocyclic ligand that exhibits both excellent reactivity for the activation of H(2)O(2) and high stability, especially at low pH and high ionic strength, is reported.

  19. Antinociceptive effect of some biuret derivatives on formalin test in mice.

    Science.gov (United States)

    Adibpour, Neda; Poornajjari, Ali; Khodayar, Mohammad Javad; Rezaee, Saeed

    2014-01-01

    The current study was designed to investigate the antinociceptive effects of several biuret derivatives with N, N`-diphenyl, N-phenyl-N`-alkylphenyl, N,N`-bis alkylphenyl, 2-methylquinoline-4-yl, benzo[d]thiazol-2-ylthio and (1-phenyl-1H-tetrazol-5-yl)thio substituents on the formalin-evoked pain in mice. Antinociceptive activity of the nine biurets derivatives were assessed at different doses in mice using formalin test and the results were compared with those of indomethacin(20 mg/kg) and vehicle of the compounds. Area under the pain score curve against time (AUEC) up to 60 minutes was used as the measure of pain behavior. A rather good analgesic effect was seen for most of the tested biuret derivatives. Significant reduction in median AUEC0-5 minutes was observed at the doses of 50 and 25 mg/kg for biurets with either benzyl and 2-methylquinoline-4-yl (C8) or phenylethyl and benzo[d]thiazol-2-ylthio(C9) moieties, respectively(p-valuebiuret compounds are able to induce antinociception in both phases of formalin test and could be considered comparable to indomethacin at the selected doses.

  20. Determination of serum proteins in the presence of dextran by means of the biuret reaction.

    Science.gov (United States)

    Gregor, A; Kostrzewska, E; Godorowska, W

    1977-02-01

    264 biuret reagents for total protein determination in serum containing dextran were investigated. The usefulness of the method is dependent on proper ratio of the components of the reagent: concentrations of NaOH and copper sulphate, and the ratio of sodium-potassium tartrate to copper sulphate.

  1. Vibrational infrared spectra of biuret and its thioanalogs. An ab initio SCF/3-21G study

    Science.gov (United States)

    Sullivan, Richard H.; Kwiatkowski, Józef S.; Leszczyński, Jerzy; Jabalameli, Ali

    1993-05-01

    The results of ab initio SCF/3-21G * calculations are reported for vibrational IR spectra (harmonic wavenumbers, absolute intensities) of biuret, thiobiuret, dithiobiuret and their model systems, formamide and thioformamide. The predicted vibrational spectra of both biurets and model systems compare well with the available experimental data, and the observed shifts of the IR wavenumbers upon deuteration of the molecules are correctly predicted by the calculations. The positions and relative intensities of characteristic IR bands of formamide and thioformamide correlate well with the corresponding data for the biurets.

  2. Negative interference of icteric serum on a bichromatic biuret total protein assay.

    Science.gov (United States)

    Gupta, Aradhana; Stockham, Steven L

    2014-09-01

    Bilirubin is stated to be a negative interferent in some biuret assays and thus could contribute to pseudohypoproteinemia in icteric samples. The purpose of the study was to evaluate the magnitude of and reason for a falsely low total protein concentration in icteric serum when the protein concentration is measured with a bichromatic spectrophotometric biuret assay. Commercially available bilirubin was dissolved in 0.1 M NaOH and mixed with sera from 2 dogs to achieve various bilirubin concentrations of up to 40 mg/dL (first set of samples) and 35 mg/dL (second set of samples, for confirmation of first set of results and to explore the interference). Biuret total protein and bilirubin concentrations were determined with a chemistry analyzer (Cobas 6000 with c501 module). Line graphs were drawn to illustrate the effects of increasing bilirubin concentrations on the total protein concentrations. Specific spectrophotometric absorbance readings were examined to identify the reason for the negative interference. High bilirubin concentrations created a negative interference in the Cobas biuret assay. The detectable interference occurred with a spiked bilirubin concentration of 10.7 mg/dL in one set of samples, 20.8 mg/dL in a second set. The interference was due to a greater secondary-absorbance reading at the second measuring point in the samples spiked with bilirubin, which possibly had converted to biliverdin. Marked hyperbilirubinemia is associated with a falsely low serum total protein concentration when measured with a bichromatic spectrophotometric biuret assay. This can result in pseudohypoproteinemia and pseudohypoglobulinemia in icteric serum. © 2014 American Society for Veterinary Clinical Pathology and European Society for Veterinary Clinical Pathology.

  3. Detection and quantification of additives (urea, biuret and poultry litter) in alfalfas by NIR spectroscopy with fibre-optic probe.

    Science.gov (United States)

    González-Martín, Inmaculada; Hernández-Hierro, José Miguel

    2008-09-15

    The additives (urea, biuret and poultry litter) present in alfalfa, which contribute non-proteic nitrogen, were analysed using near infrared spectroscopy (NIRS) technology together with a remote reflectance fibre-optic probe. We used 75 samples of known alfalfa without additives and 75 samples with each of the additives, urea (0.01-10%), biuret (0.01-10%) and poultry litter (1-25%). Using the discriminant partial least squares (DPLS) algorithm, the presence or absence of the additives urea, biuret and poultry litter is classified and predicted with a high prediction rate of 96.9%, 100% and 100%, obtaining the equations of discrimination for each additive. The regression method employed for the quantification was modified partial least squares (MPLS). The equations were developed using the fibre-optic probe to determine the content of urea, biuret and poultry litter with multiple correlation coefficients (RSQ) and prediction corrected standard errors (SEP (C)) of 0.990, 0.28% for urea, 0.991, 0.29% for biuret and 0.925, 2.08% for poultry litter. The work permits the instantaneous and simultaneous prediction and determination of urea, biuret and poultry litter in alfalfas, applying the fibre-optic directly on the ground samples of alfalfa.

  4. Insights into the trimeric HIV-1 envelope glycoprotein structure.

    Science.gov (United States)

    Ward, Andrew B; Wilson, Ian A

    2015-02-01

    The HIV-1 envelope glycoprotein (Env) trimer is responsible for receptor recognition and viral fusion with CD4(+) T cells, and is the sole target for neutralizing antibodies. Thus, understanding its molecular architecture is of significant interest. However, the Env trimer has proved to be a challenging target for 3D structure determination. Recent electron microscopy (EM) and X-ray structures have at last enabled us to decipher the structural complexity and unique features of the Env trimer, and how it is recognized by an ever-expanding arsenal of potent broadly neutralizing antibodies. We describe our current knowledge of the Env trimer structure in the context of exciting recent developments in the identification and characterization of HIV broadly neutralizing antibodies. Copyright © 2015 Elsevier Ltd. All rights reserved.

  5. Electrochemical detection of oligopeptides through the precolumn formation of biuret complexes.

    Science.gov (United States)

    Tsai, H Y; Weber, S G

    1991-04-12

    The relatively slow kinetics of formation of the electroactive Cu(II)-peptide complexes from larger (greater than 6 amino acids) peptides requires relatively high temperature and long reaction times for a postcolumn reactor. The precolumn incubation of bradykinin, Tyr8-bradykinin and insulin A chain with biuret reagent for 20 min at 60 degrees C leads to the formation of biuret complexes which can be subjected to chromatography in acidic or basic eluents. These complexes are detected electrochemically with a sensitivity similar to the Cu(II)-(ala)3 complex (1 nC/pmol at 1.0 ml/min). The influence of the column-packing material on the electrochemical detector response of the Cu-peptide complexes has also been studied.

  6. Influence of biuret and cyanuric acid on dewaxing petroleum stocks with alcoholic urea solution

    Energy Technology Data Exchange (ETDEWEB)

    Abdullaev, E.Sh.; Ismailov, A.G.; Gadzhiev, A.Sh.; Balayan, R.D.

    1987-11-01

    The influence of biuret and cyanuric acid contents on the formation and separation of the adduct in commercial dewaxing of petroleum stocks by a urea solution in a water and isopropyl alcohol mixture was studied. The stock was a diesel fuel distillate with a solid point of -12/sup 0/C. Experiments were performed with a 3.5:1 volume ratio of urea solution to feed, urea content 38% by weight, isopropyl alcohol concentration 70% by weight, adduct formation temperature 55-30/sup 0/C, and adduct formation duration 30 min. Test results show the adverse effects at different quantities of cyanuric and biuret acids on adduct formation. Solutions for overcoming these effects are proposed.

  7. 5'-Uridyl derivatives of N-glycosyl allophanic acid and biuret.

    Science.gov (United States)

    Tóth, Marietta; Somsák, László

    2010-01-11

    (2',3'-O-Isopropylidene-5'-uridyl) 4-(2,3,4,6-tetra-O-acetyl-beta-d-glycopyranosyl)allophanates were obtained in the reactions of 2',3'-O-isopropylidene-uridine and O-peracetylated beta-d-gluco-, galacto- and xylopyranosylamines, and OCNCOCl. 2,3,4,6-Tetra-O-acetyl-beta-d-glucopyranosyl isocyanate and N-(2',3'-O-isopropylidene-5'-uridyl)urea gave 1-(2,3,4,6-tetra-O-acetyl-beta-d-glucopyranosyl)-5-(2',3'-O-isopropylidene-5'-uridyl)biuret. Deprotection of the beta-d-gluco configured allophanate and biuret was carried out by standard methods. Copyright 2009 Elsevier Ltd. All rights reserved.

  8. Prevention of interference by dextran with biuret-type assay of serum proteins.

    Science.gov (United States)

    Flack, C P; Woollen, J W

    1984-04-01

    In assay of serum proteins by use of the biuret reaction, dextran can cause turbidity by formation of an insoluble complex of dextran with copper and tartrate (or EDTA) in strongly alkaline solution. Whether or not the turbidity occurs depends on the tartrate concentration: turbidity is maximal at about 10 g/L, absent at 20 g/L or more, and only slight and delayed at 4 g/L. Two biuret reagents, containing respectively 5.6 and 22.5 g of tartrate per liter, obviate the interference, but the former is suitable only when a short (5 min) incubation is used. Both reagents show linear calibration curves and yield virtually identical results.

  9. Postcolumn detection of serum proteins with the biuret and Lowry reactions.

    Science.gov (United States)

    Schlabach, T D

    1984-06-01

    The Lowry and biuret reactions have been adapted for the selective detection of chromatographically resolved proteins, specifically proteins separated by high-performance liquid chromatography. The protein reagents are continuously added to the column effluent and produce the characteristic chromophores with both proteins and peptides. The reaction chemistries are compatible with ion-exchange, steric exclusion, and reverse-phase chromatography. Detection limits for proteins resolved by ion-exchange are about 5 to 10 micrograms with the Lowry reaction. Peptides containing tyrosine can be detected at the 100-ng level when chromatographed on reverse-phase columns. The biuret reaction is about 8 times less sensitive for proteins and not very effective for peptides. Reaction detection can be combined with direct absorbance detection in the uv to distinguish proteinaceous peaks from other peaks containing uv-absorbing compounds.

  10. Interaction of erbium iodide with biuret in aqueous media at 25 deg C

    International Nuclear Information System (INIS)

    Rukk, N.S.; Zajtseva, M.G.; Alikberova, L.Yu.; Stepin, B.D.

    1989-01-01

    Solubility in ErI 3 -biuret (L)-H 2 O system is studied by isothermal method at 25 deg C. The results of determining crystallization fields by solubility method are confirmed by determining the liquid phase refractive indices. The formation of incongruently soluble complex compound of (ErL 4 )I 3 composition is ascertained, formation conditions are determined and some of its properties are investigated

  11. Preparation and Analytical Properties of 4-Hydroxybenzaldehyde, Biuret and Formaldehyde Terpolymer Resin

    OpenAIRE

    Bunian A. Shareef; Ibrahim F. Waheed; Kariem K. Jalaot

    2014-01-01

    The chelating resin was synthesized by polycondensation reaction of 4-hydroxy benzaldehyde, formaldehyde and biuret under alkaline condition. The prepared resin was characterized by IR spectroscopy and 1HNMR spectroscopy. The resin sample was cured isothermally at 1200C.The DSC scans was measured for the prepared resin. The analytical evaluation of the prepared resin toward the studied ions (Hg2+, Ni2+ and Pb2+) using the batch method. It was employed to study selectivity of metal ion...

  12. Tautomeric preferences and electron delocalization in biurets, thiobiurets, and dithiobiurets: An ab initio study

    Science.gov (United States)

    Adane, Legesse; Bharatam, Prasad V.

    In several literature reports biuret and its sulfur analogs are reported to exist in their diketo form with general formula H2N bond CX bond NH bond CY bond NH2 (X = O, Y = O, biuret; X = Y = S, dithiobiuret; and X = O, Y = S, thiobiuret). On the other hand, recently reported results on the electronic structure of biguanide analogs (X = Y = NH)demonstrated that a form equivalent to diketo is not the preferred structure. Thus, a systematic ab initio study on the tautomeric preferences of biuret and its sulfur analogs (dithiobiuret and thiobiuret) has been carried out. The results indicate that an interplay of conjugative stabilization and intramolecular hydrogen bonding to play a role in tautomeric preferences. Energy and geometric parameters, natural bond orbital analyses have been employed to understand the chemistry of the title compounds. The results indicate that unlike biguanides, these compounds prefer diketo forms containing hydrogen on the bridging nitrogen (N4) and in a trans-arrangement (1a-4a). However, tautomerization of these keto forms to the corresponding enol isomers was also found to be highly probable.

  13. Comparison of refractometer and biuret methods for total protein measurement in body cavity fluids.

    Science.gov (United States)

    George, Jeanne W.; O'Neill, Sharron L.

    2001-01-01

    Most hand-held medical refractometers have internal scales that limit protein measurement to results >/=2.5 g/dL. Tables for conversion of refraction (r) to protein concentration for values as low as 0.1 g/dL were published in the 1960s, but their accuracy for use on body fluids has not been established. The purpose of this study was to assess the reliability of body cavity fluid protein determination by refractometry. We compared the protein concentration of 25 body cavity fluids as determined by 2 Goldberg type hand-held refractometers with results obtained by the biuret method. Published charts converting refraction (r) to protein concentration were used to determine protein concentration in samples with protein /=0.6 g/dL, the lowest concentration of the biuret method's standard curve. Twenty-one peritoneal fluid, 2 pleural fluid and 2 pericardial fluid samples from 16 horses, 5 cattle, 3 dogs, 2 llamas and 1 cat were tested. The results obtained by the two refractometers were closely and linearly related to biuret results (P0.977. Based on this study, the range for quantification of body cavity fluid protein concentration by refractometry can be extended below 2.5 g/dL, allowing for quantitative assessment of most clinical samples.

  14. Determination of proteins by a reverse biuret method combined with the copper-bathocuproine chelate reaction.

    Science.gov (United States)

    Matsushita, M; Irino, T; Komoda, T; Sakagishi, Y

    1993-07-16

    A method of protein determination has been developed which combines the biuret reaction and the copper(I)-bathocuproine chelate reaction. Protein in the specimen forms a Cu(2+)-protein chelate complex (biuret reaction) during the first step. Excess Cu2+ is reduced to Cu+ by ascrobic acid, allowing the Cu+ to form a Cu(+)-bathocuproine chelate complex during the second step. The amount of Cu(+)-bathocuproine chelate complex formed is inversely proportional to the protein concentration. The sensitivity (epsilon = 1.4 x 10(6) 1.mol-1.cm-1 against human albumin) of this method was higher than that of the original Lowry (9.8 x 10(5)), pyrogallol red (1.0 x 10(6)) and commercially available Coomassie Brilliant Blue G.250 methods (6.7 x 10(5)). The color intensities of human gamma-globulin, human globulin (fractions IV-1 and IV-4), bovine albumin, egg albumin and horse gamma-globulin against human albumin (100%) ranged from 92 to 101%. The results obtained with the present method (y) correlated well with those determined by the biuret method (r = 0.998, y = 0.98 chi - 0.002, x = 1.31, y = 1.29 g/l) in 30 diluted sera. These results confirm that this assay is similar in sensitivity to the original Lowry method, is rapid and has similar reactivity to each of the various proteins in biological fluids.

  15. Automated multiple flow-injection analysis in clinical chemistry: determination of total protein with Biuret reagent.

    Science.gov (United States)

    Shideler, C E; Stewart, K K; Crump, J; Wills, M R; Savory, J; Renoe, B W

    1980-09-01

    We have examined the feasibility of the automated multiple flow-injection technique for application to clinical chemistry by adapting to this system the biuret method for the determination of total protein. Samples were discretely and rapidly introduced into a continuously flowing, nonsegmented reagent stream by means of an automatic sampler and high-pressure injection valve. Pumps operating at 1380-2070 kPa (200-300 psi) were utilized to introduce the biuret reagent and saline diluent into the system separately at flow rates of 72 and 47 microL/s, respectively. Use of 20-microL sample and a 3.0-s reaction-delay coil was adequately sensitive for analysis for total protein by this method. Samples were analyzed at a rate of 150/h with no detectable between-sample carryover. Within-run precision studies yielded relative standard deviations of 2.5% and less. Total protein values obtained by this method correlated well with those obtained by centrifugal analyzer and bubble-segmented continuous-flow biuret methods.

  16. Comparison of refractometry and biuret assay for measurement of total protein concentration in canine abdominal and pleural fluid specimens.

    Science.gov (United States)

    Rose, Alexandra; Funk, Deborah; Neiger, Reto

    2016-04-01

    To compare total protein (TP) concentrations in canine pleural and abdominal fluid specimens as measured by refractometry and biuret assay. Diagnostic test evaluation. Data regarding 92 pleural and 148 abdominal fluid specimens from dogs with various diseases. TP concentrations in fluid specimens as measured by refractometry and biuret assay were recorded. Strength of association between sets of measurements was assessed by Spearman rank correlations and Bland-Altman plots. Optimal concentration cutoff for diagnostic discrimination between exudate and nonexudate was identified by construction of receiver operating characteristic curves. Median TP concentration in pleural fluid specimens was 2.7 g/dL (range, 0.3 to 4.8 g/dL) for refractometry and 2.9 g/dL (range, 0.7 to 5.8 g/dL) for biuret assay. Median TP concentration in abdominal fluid specimens was 3.5 g/dL (range, 0.1 to 6.0 g/dL) for refractometry and 3.5 g/dL (range, 0.6 to 5.7 g/dL) for biuret assay. Correlation was significant between refractometric and biuret results for pleural (ρ = 0.921) and abdominal (ρ = 0.908) fluid. Bland-Altman plots revealed bias of -0.18 g/dL for pleural fluid and -0.03 g/dL for abdominal fluid for refractometry versus biuret assay. With a TP concentration of ≥ 3 g/dL used to distinguish exudate from nonexudate, sensitivity of refractometry was 77% for pleural fluid and 80% for abdominal fluid. Specificity was 100% and 94%, respectively. Refractometry yielded acceptable results for measurement of TP concentration in canine pleural and abdominal fluid specimens, providing a more rapid and convenient method than biuret assay.

  17. Predicted 3D Model of the Rabies Virus Glycoprotein Trimer

    Directory of Open Access Journals (Sweden)

    Bastida-González Fernando

    2016-01-01

    Full Text Available The RABVG ectodomain is a homotrimer, and trimers are often called spikes. They are responsible for the attachment of the virus through the interaction with nicotinic acetylcholine receptors, neural cell adhesion molecule (NCAM, and the p75 neurotrophin receptor (p75NTR. This makes them relevant in viral pathogenesis. The antigenic structure differs significantly between the trimers and monomers. Surfaces rich in hydrophobic amino acids are important for trimer stabilization in which the C-terminal of the ectodomain plays an important role; to understand these interactions between the G proteins, a mechanistic study of their functions was performed with a molecular model of G protein in its trimeric form. This verified its 3D conformation. The molecular modeling of G protein was performed by a I-TASSER server and was evaluated via a Rachamandran plot and ERRAT program obtained 84.64% and 89.9% of the residues in the favorable regions and overall quality factor, respectively. The molecular dynamics simulations were carried out on RABVG trimer at 310 K. From these theoretical studies, we retrieved the RMSD values from Cα atoms to assess stability. Preliminary model of G protein of rabies virus stable at 12 ns with molecular dynamics was obtained.

  18. Pulmonary effects of a polyisocyanate aerosol: hexamethylene diisocyanate trimer (HDIt) or Desmodur-N (DES-N)

    Energy Technology Data Exchange (ETDEWEB)

    Ferguson, J.S.; Schaper, M.; Alarie, Y.

    1987-07-01

    Desmodur-N (DES-N) or hexamethylene diisocyanate trimer (HDIt), a biuret structure of hexamethylene diisocyanate, is a viscous liquid used for durable coatings and is applied by brushing or spraying. DES-N aerosol has been shown to be primarily a pulmonary irritant following a single exposure in mice. To explore the pulmonary effects of this agent further, groups of guinea pigs were exposed to concentrations ranging from 8 to 121 mg/m3 of DES-N for 3 hr. Prior to and following exposure, each animal was challenged with 10% CO/sub 2/ in 20% O/sub 2/ and 70% N/sub 2/ to evaluate their pulmonary performance. Following a single exposure, these animals displayed a concentration-dependent increase in respiratory rate and decrease in tidal volume, as well as coughing and apnea. Their ventilatory response to 10% CO/sub 2/ was abnormal and characteristic of a lung restriction response. Some airflow limitation was seen during expiration but this occurred more often during air breathing than during CO/sub 2/ challenge. With daily exposures repeated for 11 consecutive days, guinea pigs began to adapt to the exposures as indicated by a return to a normal ventilatory response to CO/sub 2/. This adaptation occurred within the first 5 days of exposures. From Days 6 to 11, there was a demonstrable effect, but the level of response was much less than that following the first exposure. No cumulative effect could be demonstrated with this polyisocyanate and the effect was found to be different than that for mono- or diisocyanates. Acceptable levels of exposure to this polyisocyanate for industrial workers are suggested.

  19. Photoelectron Spectroscopy of 4-Bromochlorobenzene Dimer and Trimer Anions

    International Nuclear Information System (INIS)

    Kim, Namdoo

    2013-01-01

    I investigated the electron attachment to 4-BCB dimer and trimer anions using anion photoelectron spectroscopy and theoretical calculations. I found that an excess electron can be delocalized to these clusters through extended π-network. However, the nature of the ion core is different for the dimer and trimer. For the heterodimer anions, the geometry is determined by the differences in electronegativity of the molecules. Dimer anions with a large electronegativity difference have T-shaped geometries and those with a small electronegativity difference have PD geometries

  20. Albumin-based or albumin-linked calibrators cause a positive bias in serum proteins assayed by the biuret method.

    Science.gov (United States)

    Chromý, Vratislav; Sváchová, Lenka; Novosád, Lukás; Jarkovský, Jirí; Sedlák, Pavel; Horák, Petr; Dobrovolná, Hana; Hlavácová, Barbora

    2009-01-01

    Assay of total serum protein by the biuret method calibrated with albumin standards according to the reference method provides results with a positive bias approximately 3%-5% exceeding the total error of 3.4% allowable for total protein in serum analysis made by analysers using two-part reagents and short-term procedures. We used two types of two-part biuret reagents utilised in a short-term measurement in analysers with albumin or serum calibrators, in which protein was attested by the Kjeldahl method. Tests with potentially interfering substances proved that serum blanking used in a short-term biuret procedure is not capable of sufficiently eliminating effects of serum interferents. A short-term blanking is evidently capable of suppressing only an absorbance caused by serum-present coloured and turbid interferents, but its capacity to transform them (oxidise, hydrolyse, saponify, etc.) to some other not-interfering substances is very low compared with a long-term blanking. Lipids and bilirubin are responsible for significant positive bias of total protein in normal serum samples (approximately 3%) and even a greater positive offset in lipaemic and icteric sera (approximately 5%). We verified that interference tests based on a normal serum spiked with endogenous lipids and bilirubin give quite false and misleading results in the biuret reaction. A pure albumin, not depending on its bovine/human origin, gives absorbance responding only to its copper complexes with protein with a biuret regent, while its absorbance with a serum also includes the absorbance of interferents present in serum. The simplest way to improve current short-term biuret procedures is the use of a human serum calibrator with total protein attested by the Kjeldahl method. A serum calibrator, behaving analogously to serum samples, compensates for a positive bias in most normal sera. Reagents with a greater concentration of active biuret components (copper and alkali, reference method included

  1. Epitope-Independent Purification of Native-Like Envelope Trimers from Diverse HIV-1 Isolates

    OpenAIRE

    Verkerke, Hans P.; Williams, James A.; Guttman, Miklos; Simonich, Cassandra A.; Liang, Yu; Filipavicius, Modestas; Hu, Shiu-Lok; Overbaugh, Julie; Lee, Kelly K.

    2016-01-01

    Soluble forms of trimeric HIV-1 envelope glycoprotein (Env) have long been sought as immunogens and as reagents for analysis of Env structure and function. Isolation of trimers that mimic native Env, derived from diverse viruses, however, represents a major challenge. Thus far, the most promising native-like (NL) structures have been obtained by engineering trimer-stabilizing mutations, termed SOSIP, into truncated Env sequences. However, the abundances of NL trimeric conformers vary among En...

  2. Tetra-aqua-bis(biuret-κO,O')yttrium(III) trichloride.

    Science.gov (United States)

    Harrison, William T A

    2008-04-02

    In the title compound, [Y(C(2)H(5)N(3)O(2))(2)(H(2)O)(4)]Cl(3), the Y(3+) ion (site symmetry 2) is bonded to eight O atoms (arising from two O,O'-bidentate biuret mol-ecules and four water mol-ecules) in a distorted square-anti-prismatic arrangement. A network of N-H⋯O, N-H⋯Cl and O-H⋯Cl hydrogen bonds help to establish the packing, leading to a three-dimensional network. One of the chloride ions has site symmetry 2.

  3. Tetra­aqua­bis(biuret-κ2 O,O′)yttrium(III) trichloride

    Science.gov (United States)

    Harrison, William T. A.

    2008-01-01

    In the title compound, [Y(C2H5N3O2)2(H2O)4]Cl3, the Y3+ ion (site symmetry 2) is bonded to eight O atoms (arising from two O,O′-bidentate biuret mol­ecules and four water mol­ecules) in a distorted square-anti­prismatic arrangement. A network of N—H⋯O, N—H⋯Cl and O—H⋯Cl hydrogen bonds help to establish the packing, leading to a three-dimensional network. One of the chloride ions has site symmetry 2. PMID:21202174

  4. Tetra-aqua-bis(biuret-κO,O')gadolinium(III) trichloride.

    Science.gov (United States)

    Harrison, William T A

    2008-04-02

    In the title compound, [Gd(C(2)H(5)N(3)O(2))(2)(H(2)O)(4)]Cl(3), which is isostrucutural with its yttrium analogue, the Gd(3+) ion (site symmetry 2) is bonded to eight O atoms (arising from two O,O'-bidentate biuret mol-ecules and four water mol-ecules) in a distorted square-anti-prismatic arrangement. A network of N-H⋯O, N-H⋯Cl and O-H⋯Cl hydrogen bonds helps to establish the packing, leading to a three-dimensional network. One of the chloride ions has site symmetry 2.

  5. Tetra­aqua­bis(biuret-κ2 O,O′)gadolinium(III) trichloride

    Science.gov (United States)

    Harrison, William T. A.

    2008-01-01

    In the title compound, [Gd(C2H5N3O2)2(H2O)4]Cl3, which is isostrucutural with its yttrium analogue, the Gd3+ ion (site symmetry 2) is bonded to eight O atoms (arising from two O,O′-bidentate biuret mol­ecules and four water mol­ecules) in a distorted square-anti­prismatic arrangement. A network of N—H⋯O, N—H⋯Cl and O—H⋯Cl hydrogen bonds helps to establish the packing, leading to a three-dimensional network. One of the chloride ions has site symmetry 2. PMID:21202175

  6. Hydroxonium triaquabis(biuret-κ2O,O′dichloridolanthanum(III dichloride dihydrate

    Directory of Open Access Journals (Sweden)

    William T. A. Harrison

    2008-05-01

    Full Text Available In the title compound, (H3O[LaCl2(C2H5N3O22(H2O3]Cl2·2H2O, the La atom is bonded to seven O atoms (arising from two O,O′-bidentate biuret molecules and three water molecules and two chloride ions in an irregular arrangement. A network of N—H...O, N—H...Cl, O—H...O and O—H...Cl hydrogen bonds helps to establish the packing, leading to a three-dimensional network. The La atom, one Cl atom and four O atoms lie on a crystallographic mirror plane.

  7. Tetraaquabis(biuret-κ2O,O′yttrium(III trichloride

    Directory of Open Access Journals (Sweden)

    William T. A. Harrison

    2008-05-01

    Full Text Available In the title compound, [Y(C2H5N3O22(H2O4]Cl3, the Y3+ ion (site symmetry 2 is bonded to eight O atoms (arising from two O,O′-bidentate biuret molecules and four water molecules in a distorted square-antiprismatic arrangement. A network of N—H...O, N—H...Cl and O—H...Cl hydrogen bonds help to establish the packing, leading to a three-dimensional network. One of the chloride ions has site symmetry 2.

  8. Tetraaquabis(biuret-κ2O,O′gadolinium(III trichloride

    Directory of Open Access Journals (Sweden)

    William T. A. Harrison

    2008-05-01

    Full Text Available In the title compound, [Gd(C2H5N3O22(H2O4]Cl3, which is isostrucutural with its yttrium analogue, the Gd3+ ion (site symmetry 2 is bonded to eight O atoms (arising from two O,O′-bidentate biuret molecules and four water molecules in a distorted square-antiprismatic arrangement. A network of N—H...O, N—H...Cl and O—H...Cl hydrogen bonds helps to establish the packing, leading to a three-dimensional network. One of the chloride ions has site symmetry 2.

  9. Combined gel filtration, biuret/copper method compared with an immunochemical method for urinary protein measurement.

    Science.gov (United States)

    Peele, J D; Gadsden, R H; Loadholt, C B

    1977-01-01

    We compared an immunochemical method specific for plasma proteins with a chemical method, in which interfering substances are separated by gel filtration and "total" urinary protein is determined by the biuret reaction followed by reaction of protein-bound copper with diethyldithiocarbamate after a second gel filtration to remove nonprotein-bound copper. More than 250 24-h urine samples were analyzed by each method. There was linear agreement and a correlation of 0.96 between the two methods, but urinary protein values determined by the immunochemical method, especially for patients with multiple myeloma, were lower than by the chemical method.

  10. Cleavage strongly influences whether soluble HIV-1 envelope glycoprotein trimers adopt a native-like conformation

    NARCIS (Netherlands)

    Ringe, Rajesh P.; Sanders, Rogier W.; Yasmeen, Anila; Kim, Helen J.; Lee, Jeong Hyun; Cupo, Albert; Korzun, Jacob; Derking, Ronald; van Montfort, Thijs; Julien, Jean-Philippe; Wilson, Ian A.; Klasse, Per Johan; Ward, Andrew B.; Moore, John P.

    2013-01-01

    We compare the antigenicity and conformation of soluble, cleaved vs. uncleaved envelope glycoprotein (Env gp) 140 trimers from the subtype A HIV type 1 (HIV-1) strain BG505. The impact of gp120-gp41 cleavage on trimer structure, in the presence or absence of trimer-stabilizing modifications (i.e., a

  11. DNAzyme-Controlled Cleavage of Dimer and Trimer Origami Tiles.

    Science.gov (United States)

    Wu, Na; Willner, Itamar

    2016-04-13

    Dimers of origami tiles are bridged by the Pb(2+)-dependent DNAzyme sequence and its substrate or by the histidine-dependent DNAzyme sequence and its substrate to yield the dimers T1-T2 and T3-T4, respectively. The dimers are cleaved to monomer tiles in the presence of Pb(2+)-ions or histidine as triggers. Similarly, trimers of origami tiles are constructed by bridging the tiles with the Pb(2+)-ion-dependent DNAzyme sequence and the histidine-dependent DNAzyme sequence and their substrates yielding the trimer T1-T5-T4. In the presence of Pb(2+)-ions and/or histidine as triggers, the programmed cleavage of trimer proceeds. Using Pb(2+) or histidine as trigger cleaves the trimer to yield T5-T4 and T1 or the dimer T1-T5 and T4, respectively. In the presence of Pb(2+)-ions and histidine as triggers, the cleavage products are the monomer tiles T1, T5, and T4. The different cleavage products are identified by labeling the tiles with 0, 1, or 2 streptavidin labels and AFM imaging.

  12. Identification of Trimer and Dimer of 4-hydroxy-3-methoxy ...

    African Journals Online (AJOL)

    The effect of intermolecular hydrogen bonding on the molecular structure of vanillin has been studied using negative ion chemical ionization (NICl) mass spectrometry methods. The [Trimer-H2O] and [Dimer-H2O] were observed at m/z 438 and 286 respectively in NICl (CH4) mass spectrum of vanillin. The NICl (Cl) mass ...

  13. Photosystem I : a search for green plant trimers

    NARCIS (Netherlands)

    Oosterwijk, Niels van; Yakushevska, Alevtyna E.; Boekema, Egbert J.; Kouril, R.

    2005-01-01

    Recent blue-native gel electrophoresis studies gave evidence for the existence of dimeric and trimeric PSI complexes in green plants. We used single particle electron microscopy to investigate all the larger particles from the thylakoid membrane of pea (Pisum sativum var. Charmette). Peak fractions

  14. Synthesis of new glycosyl biuret and urea derivatives as potential glycoenzyme inhibitors.

    Science.gov (United States)

    Felföldi, Nóra; Tóth, Marietta; Chrysina, Evangelia D; Charavgi, Maria-Despoina; Alexacou, Kyra-Melinda; Somsák, László

    2010-01-26

    O-peracetylated 1-(beta-D-glucopyranosyl)-5-phenylbiuret was prepared in the reaction of O-peracetylated beta-D-glucopyranosylisocyanate and phenylurea. The reaction of O-peracetylated N-beta-D-glucopyranosylurea with phenylisocyanate furnished the corresponding 1-(beta-D-glucopyranosyl)-3,5-diphenyl- as well as 3-(beta-D-glucopyranosyl)-1,5-diphenyl biurets besides 1-(beta-D-glucopyranosyl)-3-phenylurea. O-Peracetylated 1-(beta-D-glucopyranosyl)-5-(beta-D-glycopyranosyl)biurets were obtained in one-pot reactions of O-peracetylated beta-D-glucopyranosylamine with OCNCOCl followed by a second glycopyranosylamine of beta-D-gluco, beta-D-galacto and beta-D-xylo configurations. O-Acyl protected 1-(beta-D-glucopyranosyl)-3-(beta-D-glycopyranosylcarbonyl)ureas were obtained from the reaction of beta-D-glucopyranosylisocyanate with C-(glycopyranosyl)formamides of beta-D-gluco and beta-D-galacto configurations. The O-acyl protecting groups were removed under acid- or base-catalyzed transesterification conditions, except for the N-acylurea derivatives where the cleavage of the N-acyl groups was faster than deprotection. Some of the new compounds exhibited moderate inhibition against rabbit muscle glycogen phosphorylase b and human salivary alpha-amylase. 2009 Elsevier Ltd. All rights reserved.

  15. A practical method for extending the biuret assay to protein determination of corn-based products.

    Science.gov (United States)

    Liu, Zelong; Pan, Junhui

    2017-06-01

    A modified biuret method suitable for protein determination of corn-based products was developed by introducing a combination of an alkaline reagent with sodium dodecyl sulfate (reagent A) and heat treatments. The method was tested on seven corn-based samples. The results showed mostly good agreement (P>0.05) as compared to the Kjeldahl values. The proposed method was found to enhance the accuracy of prediction on zein content using bovine serum albumin as standard. Reagent A and sample treatment were proved to effectively improve protein solubilization for the thermally-dried corn-based products, e.g. corn gluten meal. The absorbance was stable for at least 1-h. Moreover, the whole measurement of protein content only needs 15-20min more than the traditional biuret assay, and can be performed in batches. The findings suggest that the proposed method could be a timesaving alternative for routine protein analyses in corn processing factories. Copyright © 2016 Elsevier Ltd. All rights reserved.

  16. One pot glucose detection by [Fe(III)(biuret-amide)] immobilized on mesoporous silica nanoparticles: an efficient HRP mimic.

    Science.gov (United States)

    Malvi, Bharmana; Panda, Chakadola; Dhar, Basab B; Gupta, Sayam Sen

    2012-05-28

    An [Fe(III)(biuret-amide)] complex has been immobilized onto mesoporous silica nanoparticles via Cu(I) catalyzed azide-alkyne click chemistry. This hybrid material functions as an efficient peroxidase mimic and was successfully used for the quantitative determination of hydrogen peroxide and glucose via a one-pot colorimetric assay. This journal is © The Royal Society of Chemistry 2012

  17. A New Colorimetric Assay of Tabletop Sweeteners Using a Modified Biuret Reagent: An Analytical Chemistry Experiment for the Undergraduate Curriculum

    Science.gov (United States)

    Fenk, Christopher J.; Kaufman, Nathan; Gerbig, Donald G., Jr.

    2007-01-01

    A new, fast and effective colorimetric analysis of the artificial sweetener aspartame is presented for application in undergraduate laboratory courses. This new method incorporates the use of a modified biuret reagent for selective detection and analysis of aspartame in aqueous solutions. The modified reagent is less caustic than the traditional…

  18. The Trimeric Model: A New Model of Periodontal Treatment Planning

    Science.gov (United States)

    Tarakji, Bassel

    2014-01-01

    Treatment of periodontal disease is a complex and multidisciplinary procedure, requiring periodontal, surgical, restorative, and orthodontic treatment modalities. Several authors attempted to formulate models for periodontal treatment that orders the treatment steps in a logical and easy to remember manner. In this article, we discuss two models of periodontal treatment planning from two of the most well-known textbook in the specialty of periodontics internationally. Then modify them to arrive at a new model of periodontal treatment planning, The Trimeric Model. Adding restorative and orthodontic interrelationships with periodontal treatment allows us to expand this model into the Extended Trimeric Model of periodontal treatment planning. These models will provide a logical framework and a clear order of the treatment of periodontal disease for general practitioners and periodontists alike. PMID:25177662

  19. Crystal structures of the TRIC trimeric intracellular cation channel orthologues.

    Science.gov (United States)

    Kasuya, Go; Hiraizumi, Masahiro; Maturana, Andrés D; Kumazaki, Kaoru; Fujiwara, Yuichiro; Liu, Keihong; Nakada-Nakura, Yoshiko; Iwata, So; Tsukada, Keisuke; Komori, Tomotaka; Uemura, Sotaro; Goto, Yuhei; Nakane, Takanori; Takemoto, Mizuki; Kato, Hideaki E; Yamashita, Keitaro; Wada, Miki; Ito, Koichi; Ishitani, Ryuichiro; Hattori, Motoyuki; Nureki, Osamu

    2016-12-01

    Ca 2+ release from the sarcoplasmic reticulum (SR) and endoplasmic reticulum (ER) is crucial for muscle contraction, cell growth, apoptosis, learning and memory. The trimeric intracellular cation (TRIC) channels were recently identified as cation channels balancing the SR and ER membrane potentials, and are implicated in Ca 2+ signaling and homeostasis. Here we present the crystal structures of prokaryotic TRIC channels in the closed state and structure-based functional analyses of prokaryotic and eukaryotic TRIC channels. Each trimer subunit consists of seven transmembrane (TM) helices with two inverted repeated regions. The electrophysiological, biochemical and biophysical analyses revealed that TRIC channels possess an ion-conducting pore within each subunit, and that the trimer formation contributes to the stability of the protein. The symmetrically related TM2 and TM5 helices are kinked at the conserved glycine clusters, and these kinks are important for the channel activity. Furthermore, the kinks of the TM2 and TM5 helices generate lateral fenestrations at each subunit interface. Unexpectedly, these lateral fenestrations are occupied with lipid molecules. This study provides the structural and functional framework for the molecular mechanism of this ion channel superfamily.

  20. Synthesis, Optical Characterization, and Thermal Decomposition of Complexes Based on Biuret Ligand

    Directory of Open Access Journals (Sweden)

    Mei-Ling Wang

    2016-01-01

    Full Text Available Four complexes were synthesized in methanol solution using nickel acetate or nickel chloride, manganese acetate, manganese chloride, and biuret as raw materials. The complexes were characterized by elemental analyses, UV, FTIR, Raman spectra, X-ray powder diffraction, and thermogravimetric analysis. The compositions of the complexes were [Ni(bi2(H2O2](Ac2·H2O (1, [Ni(bi2Cl2] (2, [Mn(bi2(Ac2]·1.5H2O (3, and [Mn(bi2Cl2] (4 (bi = NH2CONHCONH2, respectively. In the complexes, every metal ion was coordinated by oxygen atoms or chlorine ions and even both. The nickel and manganese ions were all hexacoordinated. The thermal decomposition processes of the complexes under air included the loss of water molecule, the pyrolysis of ligands, and the decomposition of inorganic salts, and the final residues were nickel oxide and manganese oxide, respectively.

  1. Hydroxonium triaqua­bis(biuret-κ2 O,O′)dichloridolanthanum(III) dichloride dihydrate

    Science.gov (United States)

    Harrison, William T. A.

    2008-01-01

    In the title compound, (H3O)[LaCl2(C2H5N3O2)2(H2O)3]Cl2·2H2O, the La atom is bonded to seven O atoms (arising from two O,O′-bidentate biuret mol­ecules and three water mol­ecules) and two chloride ions in an irregular arrangement. A network of N—H⋯O, N—H⋯Cl, O—H⋯O and O—H⋯Cl hydrogen bonds helps to establish the packing, leading to a three-dimensional network. The La atom, one Cl atom and four O atoms lie on a crystallographic mirror plane. PMID:21202219

  2. Hydroxonium triaqua-bis(biuret-κO,O')dichloridolanthanum(III) dichloride dihydrate.

    Science.gov (United States)

    Harrison, William T A

    2008-04-18

    In the title compound, (H(3)O)[LaCl(2)(C(2)H(5)N(3)O(2))(2)(H(2)O)(3)]Cl(2)·2H(2)O, the La atom is bonded to seven O atoms (arising from two O,O'-bidentate biuret mol-ecules and three water mol-ecules) and two chloride ions in an irregular arrangement. A network of N-H⋯O, N-H⋯Cl, O-H⋯O and O-H⋯Cl hydrogen bonds helps to establish the packing, leading to a three-dimensional network. The La atom, one Cl atom and four O atoms lie on a crystallographic mirror plane.

  3. Epitope-Independent Purification of Native-Like Envelope Trimers from Diverse HIV-1 Isolates.

    Science.gov (United States)

    Verkerke, Hans P; Williams, James A; Guttman, Miklos; Simonich, Cassandra A; Liang, Yu; Filipavicius, Modestas; Hu, Shiu-Lok; Overbaugh, Julie; Lee, Kelly K

    2016-10-15

    Soluble forms of trimeric HIV-1 envelope glycoprotein (Env) have long been sought as immunogens and as reagents for analysis of Env structure and function. Isolation of trimers that mimic native Env, derived from diverse viruses, however, represents a major challenge. Thus far, the most promising native-like (NL) structures have been obtained by engineering trimer-stabilizing mutations, termed SOSIP, into truncated Env sequences. However, the abundances of NL trimeric conformers vary among Envs, necessitating purification by monoclonal antibodies (MAbs) like PGT145, which target specific epitopes. To surmount this inherent limitation, we developed an approach that uses lectin affinity chromatography, ion-exchange chromatography, hydrophobic-interaction chromatography (HIC), and size exclusion chromatography (SEC) to isolate NL trimers from nonnative Env species. We validated this method with SOSIP trimers from HIV-1 clades A and B. Analyses by SEC, blue native PAGE, SDS-PAGE, and dynamic light scattering indicated that the resulting material was homogeneous (>95% pure), fully cleaved, and of the appropriate molecular weight and size for SOSIP trimers. Negative-stain electron microscopy further demonstrated that our preparations were composed of NL trimeric structures. By hydrogen/deuterium-exchange mass spectrometry, these HIC-pure trimers exhibited structural organization consistent with NL trimers and inconsistent with profiles seen in nonnative Envs. Screened for antigenicity, some Envs, like BS208.b1 and KNH1144 T162A, did not present the glycan/quaternary structure-dependent epitope for PGT145 binding, suggesting that these SOSIPs would be challenging to isolate by existing MAb affinity methods. By selecting based on biochemical rather than antigenic properties, our method offers an epitope-independent alternative to MAbs for isolation of NL Env trimers. The production and purification of diverse soluble Env trimers that maintain native-like (NL) structure

  4. Direct visualization of the trimeric structure of the ASIC1a channel, using AFM imaging

    International Nuclear Information System (INIS)

    Carnally, Stewart M.; Dev, Harveer S.; Stewart, Andrew P.; Barrera, Nelson P.; Van Bemmelen, Miguel X.; Schild, Laurent; Henderson, Robert M.; Edwardson, J.Michael

    2008-01-01

    There has been confusion about the subunit stoichiometry of the degenerin family of ion channels. Recently, a crystal structure of acid-sensing ion channel (ASIC) 1a revealed that it assembles as a trimer. Here, we used atomic force microscopy (AFM) to image unprocessed ASIC1a bound to mica. We detected a mixture of subunit monomers, dimers and trimers. In some cases, triple-subunit clusters were clearly visible, confirming the trimeric structure of the channel, and indicating that the trimer sometimes disaggregated after adhesion to the mica surface. This AFM-based technique will now enable us to determine the subunit arrangement within heteromeric ASICs

  5. The transcriptional form of the phosphoprotein of vesicular stomatitis virus is a trimer: structure and stability.

    Science.gov (United States)

    Gao, Y; Greenfield, N J; Cleverley, D Z; Lenard, J

    1996-11-19

    The phosphoprotein (P) of vesicular stomatitis virus was previously shown to assemble into a homomultimer upon phosphorylation by casein kinase II. It thus acquired transcriptional activity, including the ability to bind to the other two transcriptional components, the polymerase L and the N-RNA template. This multimer has now been found to be a trimer using a His-tag dilution method. Trimer stability was assessed using a variation of this method, by measuring the rate of exchange of monomers between preformed tagged and untagged trimers at different values of pH and ionic strength. Exchange rates increased with increasing ionic strength and were similar at pH 6, 8, and 10, but the trimer was completely dissociated at pH 4. This suggests that the trimer is stabilized by electrostatic interactions, probably involving carboxylate and guanidino groups. Addition of viral L protein stabilized the P trimers, completely preventing subunit exchange under transcription conditions. The association constants (Kass) for trimerization of partially active D and A substitution mutants were also determined by His-tag dilution and found to correlate well with transcriptional activity, further confirming that the active species is the trimer. Circular dichroism spectra were identical for phosphorylated and unphosphorylated wild-type P protein and for D and A mutants known to be predominantly trimeric and monomeric, respectively.

  6. Severe corrosion of copper in a highly alkaline egg white solution due to a biuret corrosion reaction

    International Nuclear Information System (INIS)

    Liu, Xun; Hu, Wenyuan; Yang, Shan; Li, Zhaoqian; Pei, Chonghua; Zhou, Yong; Yin, Guanfu

    2015-01-01

    Highlights: • Egg whites can accelerate the corrosion of copper. • The corrosion rate of copper is closely related to the pH values. • The mechanism of the strong corrosion is due to a biuret corrosion reaction. - Abstract: Through investigating the interaction between copper metal and egg whites as well as the effects of pH, oxygen and the species of alkalis on copper corrosion rate, an unusual severe corrosion of copper metal in a highly alkaline egg white solution was found. The mechanism was investigated through detailed characterizations using a scanning electron microscope, X-ray diffraction, X-ray photoelectron spectroscopy and UV–visible absorption spectroscopy. The severe corrosion was mainly due to the formation of a hydroxyl–peptide–copper multicomponent stable soluble complex, which the biuret reaction also produces

  7. A native-like SOSIP.664 trimer based on an HIV-1 subtype B env gene.

    Science.gov (United States)

    Pugach, Pavel; Ozorowski, Gabriel; Cupo, Albert; Ringe, Rajesh; Yasmeen, Anila; de Val, Natalia; Derking, Ronald; Kim, Helen J; Korzun, Jacob; Golabek, Michael; de Los Reyes, Kevin; Ketas, Thomas J; Julien, Jean-Philippe; Burton, Dennis R; Wilson, Ian A; Sanders, Rogier W; Klasse, P J; Ward, Andrew B; Moore, John P

    2015-03-01

    Recombinant trimeric mimics of the human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein (Env) spike should expose as many epitopes as possible for broadly neutralizing antibodies (bNAbs) but few, if any, for nonneutralizing antibodies (non-NAbs). Soluble, cleaved SOSIP.664 gp140 trimers based on the subtype A strain BG505 approach this ideal and are therefore plausible vaccine candidates. Here, we report on the production and in vitro properties of a new SOSIP.664 trimer derived from a subtype B env gene, B41, including how to make this protein in low-serum media without proteolytic damage (clipping) to the V3 region. We also show that nonclipped trimers can be purified successfully via a positive-selection affinity column using the bNAb PGT145, which recognizes a quaternary structure-dependent epitope at the trimer apex. Negative-stain electron microscopy imaging shows that the purified, nonclipped, native-like B41 SOSIP.664 trimers contain two subpopulations, which we propose represent an equilibrium between the fully closed and a more open conformation. The latter is different from the fully open, CD4 receptor-bound conformation and may represent an intermediate state of the trimer. This new subtype B trimer adds to the repertoire of native-like Env proteins that are suitable for immunogenicity and structural studies. The cleaved, trimeric envelope protein complex is the only neutralizing antibody target on the HIV-1 surface. Many vaccine strategies are based on inducing neutralizing antibodies. For HIV-1, one approach involves using recombinant, soluble protein mimics of the native trimer. At present, the only reliable way to make native-like, soluble trimers in practical amounts is via the introduction of specific sequence changes that confer stability on the cleaved form of Env. The resulting proteins are known as SOSIP.664 gp140 trimers, and the current paradigm is based on the BG505 subtype A env gene. Here, we describe the production and

  8. A mixed Ni(II) ionic complex containing V-shaped water trimer ...

    Indian Academy of Sciences (India)

    A mixed Ni(II) ionic complex containing V-shaped water trimer: Synthesis, spectral, structural and ... bonding interaction between three lattice water molecule forms a V-shaped trimer (H2O)3 which gives rise to a. 1-D polymeric structure in the ... with organic ligands should be handled with care as they can cause explosion.

  9. Allergen Ara h 1 occurs in peanuts as a large oligomer rather than as a trimer

    NARCIS (Netherlands)

    Boxtel, E.L. van; Beers, M.M.C. van; Koppelman, S.J.; Broek, L.A.M. van den; Gruppen, H.

    2006-01-01

    Ara h 1, a major peanut allergen, is known as a stable trimeric protein. Nevertheless, upon purification of native Ara h 1 from peanuts using only size exclusion chromatography, the allergen appeared to exist in an oligomeric structure, rather than as a trimeric structure. The oligomeric structure

  10. Effects of dextran on five biuret-based procedures for total protein in serum.

    Science.gov (United States)

    Barnes, D B; Pierce, G F; Lichti, D; Landt, M; Koenig, J; Chan, K M

    1985-12-01

    We evaluated the effect of dextran on values for total protein in serum as measured by the biuret method with five widely used automated instruments: the American Monitor Parallel; the Du Pont aca II; the Roche Cobas-Bio; the Kodak Ektachem 400; and the Beckman Astra 8. Dextran concentrations as great as 25 or 30 g/L had relatively little or no influence on total protein measurements by the latter three instruments. Dextran concentrations exceeding 6 g/L caused falsely low results with the aca, whereas the Parallel gave falsely high results when the dextran concentration exceeded 2 g/L. The aca total protein procedure could be protected from the interference by dextran concentrations up to 30 g/L by injecting 0.4-0.8 mL of ethylene glycol directly into the reagent pack before sampling. However, we could not eliminate the interference with the Parallel procedure by any simple means; we thus recommend that it not be used for measuring total protein in serum samples from patients who are being treated with dextran.

  11. Chelation Ion Exchange Properties of 2, 4-Dihydroxyacetophenone-Biuret-Formaldehyde Terpolymer Resin

    Directory of Open Access Journals (Sweden)

    Sanjiokumar S. Rahangdale

    2009-01-01

    Full Text Available The terpolymer resin 2, 4-HABF has been synthesized by the condensation of 2, 4-dihydroxyacetophenone (2, 4-HA and biuret (B with formaldehyde (F in 1:1:2 molar ratios in presence of 2 M hydrochloric acid as catalyst. UV-Visible, IR and proton NMR spectral studies have been carried out to elucidate the structure of the resin. A terpolymer (2, 4-HABF proved to be a selective chelating ion exchange polymer for certain metals. Chelating ion-exchange properties of this polymer were studied for Fe3+, Cu2+, Ni2+, Co2+, Zn2+, Cd2+ and Pb2+ ions. A batch equilibrium method was employed in the study of the selectivity of metal ion uptake involving the measurement of the distribution of a given metal ion between the polymer sample and a solution containing the metal ion. The study was carried out over a wide pH range and in media of various ionic strengths. The polymer showed highest selectivity for Fe3+, Cu2+ ions than for Ni2+, Co2+, Zn2+, Cd2+, and Pb2+ ions. Study of distribution ratio as a formation of pH indicates that the amount of metal ion taken by resin is increases with the increasing pH of the medium.

  12. Covalent Linkage of HIV-1 Trimers to Synthetic Liposomes Elicits Improved B Cell and Antibody Responses.

    Science.gov (United States)

    Bale, Shridhar; Goebrecht, Geraldine; Stano, Armando; Wilson, Richard; Ota, Takayuki; Tran, Karen; Ingale, Jidnyasa; Zwick, Michael B; Wyatt, Richard T

    2017-08-15

    We have demonstrated that a liposomal array of well-ordered trimers enhances B cell activation, germinal center formation, and the elicitation of tier-2 autologous neutralizing antibodies. Previously, we coupled well-ordered cleavage-independent NFL trimers via their C-terminal polyhistidine tails to nickel lipids integrated into the lipid bilayer. Despite favorable in vivo effects, concern remained over the potentially longer-term in vivo instability of noncovalent linkage of the trimers to the liposomes. Accordingly, we tested both cobalt coupling and covalent linkage of the trimers to the liposomes by reengineering the polyhistidine tail to include a free cysteine on each protomer of model BG505 NFL trimers to allow covalent linkage. Both cobalt and cysteine coupling resulted in a high-density array of NFL trimers that was stable in both 20% mouse serum and 100 mM EDTA, whereas the nickel-conjugated trimers were not stable under these conditions. Binding analysis and calcium flux with anti-Env-specific B cells confirmed that the trimers maintained conformational integrity following coupling. Following immunization of mice, serologic analysis demonstrated that the covalently coupled trimers elicited Env-directed antibodies in a manner statistically significantly improved compared to soluble trimers and nickel-conjugated trimers. Importantly, the covalent coupling not only enhanced gp120-directed responses compared to soluble trimers, it also completely eliminated antibodies directed to the C-terminal His tag located at the "bottom" of the spike. In contrast, soluble and noncovalent formats efficiently elicited anti-His tag antibodies. These data indicate that covalent linkage of well-ordered trimers to liposomes in high-density array displays multiple advantages in vitro and in vivo IMPORTANCE Enveloped viruses typically encode a surface-bound glycoprotein that mediates viral entry into host cells and is a primary target for vaccine design. Liposomes with

  13. Understanding tantalum-catalyzed ethylene trimerization: When things go wrong

    KAUST Repository

    Chen, Yin

    2013-06-07

    Ethylene oligomerization to linear low-molecular-mass α-olefins is an open industrial challenge. Ta-based catalysts are promising systems, but the unclear understanding of their behavior prevents systematic advances in the field. We demonstrate here that a well-defined (î -SiO)3Ta III species is able to promote ethylene oligo-/polymerization without any cocatalyst, confirming that the active species in Ta systems corresponds to a TaIII species. DFT calculations on a series of Ta systems ranging from ethylene trimerization to ethylene polymerization catalysts highlight the key factors controlling their experimental behavior. Comparison of these Ta systems allows one to set general rules for the rational development of new ethylene Ta oligomerization catalysts. © 2013 American Chemical Society.

  14. Soluble Prefusion Closed DS-SOSIP.664-Env Trimers of Diverse HIV-1 Strains.

    Science.gov (United States)

    Joyce, M Gordon; Georgiev, Ivelin S; Yang, Yongping; Druz, Aliaksandr; Geng, Hui; Chuang, Gwo-Yu; Kwon, Young Do; Pancera, Marie; Rawi, Reda; Sastry, Mallika; Stewart-Jones, Guillaume B E; Zheng, Angela; Zhou, Tongqing; Choe, Misook; Van Galen, Joseph G; Chen, Rita E; Lees, Christopher R; Narpala, Sandeep; Chambers, Michael; Tsybovsky, Yaroslav; Baxa, Ulrich; McDermott, Adrian B; Mascola, John R; Kwong, Peter D

    2017-12-05

    The elicitation of autologous neutralizing responses by immunization with HIV-1 envelope (Env) trimers conformationally stabilized in a prefusion closed state has generated considerable interest in the HIV-1 vaccine field. However, soluble prefusion closed Env trimers have been produced from only a handful of HIV-1 strains, limiting their utility as vaccine antigens and B cell probes. Here, we report the engineering from 81 HIV-1 strains of soluble, fully cleaved, prefusion Env trimers with appropriate antigenicity. We used a 96-well expression-screening format to assess the ability of artificial disulfides and Ile559Pro substitution (DS-SOSIP) to produce soluble cleaved-Env trimers; from 180 Env strains, 20 yielded prefusion closed trimers. We also created chimeras, by utilizing structure-based design to incorporate select regions from the well-behaved BG505 strain; from 180 Env strains, 78 DS-SOSIP-stabilized chimeras, including 61 additional strains, yielded prefusion closed trimers. Structure-based design thus enables the production of prefusion closed HIV-1-Env trimers from dozens of diverse strains. Published by Elsevier Inc.

  15. Application of biuret, dicyandiamide, or urea as a cathode buffer layer toward the efficiency enhancement of polymer solar cells.

    Science.gov (United States)

    Zhao, Xuemei; Xu, Chenhui; Wang, Haitao; Chen, Fei; Zhang, Wenfeng; Zhao, Zhiqiang; Chen, Liwei; Yang, Shangfeng

    2014-03-26

    Three amino-containing small-molecule organic materials-biuret, dicyandiamide (DCDA), and urea-were successfully applied as novel cathode buffer layers (CBLs) in P3HT:PCBM bulk heterojunction polymer solar cells (BHJ-PSCs) for the first time, resulting in obvious efficiency enhancement. Under the optimized condition, the power conversion efficiencies (PCEs) of the CBL-incorporated BHJ-PSC devices are 3.84%, 4.25%, and 4.39% for biuret, DCDA, and urea, which are enhanced by ∼15%, ∼27%, and ∼31%, respectively, compared to the reference poly(3-hexylthiophene-2,5-diyl) : [6,6]-phenyl-C61-butyric acid methyl ester (P3HT:PCBM) BHJ-PSC device without any CBL. The efficiency enhancement is primarily attributed to the increases of both short-circuit current density (Jsc) and fill factor (FF), for which the enhancement ratio is found to be sensitively dependent on the molecular structure of small-molecule organic materials. The surface morphologies and surface potential changes of the CBL-incorporated P3HT:PCBM photoactive layers were studied by atomic force microscopy and scanning Kelvin probe microscopy, respectively, suggesting the formation of an interfacial dipole layer between the photoactive layer and Al cathode, which may decrease the energy level offset between the work function of Al and the lowest unoccipoed molecular orbital level (LUMO) of the PCBM acceptor and consequently facilitate electron extraction by the Al cathode. The difference in the enhancement effect of biuret, DCDA, and urea is due to their difference on the work function matching with P3HT:PCBM. Besides, the coordination interaction between the lone-pair electrons on the N atoms of the amino (-NH2) group and the Al atoms may prohibit interaction between Al and the thiophene rings of P3HT, contributing to the efficiency enhancement of the CBL-incorporated devices as well. In this sense, the different CBL performance of biuret, DCDA, and urea is also proposed to partially originate from the

  16. Structure-Guided Redesign Increases the Propensity of HIV Env To Generate Highly Stable Soluble Trimers.

    Science.gov (United States)

    Guenaga, Javier; Dubrovskaya, Viktoriya; de Val, Natalia; Sharma, Shailendra K; Carrette, Barbara; Ward, Andrew B; Wyatt, Richard T

    2015-12-30

    Due to high viral diversity, an effective HIV-1 vaccine will likely require Envs derived from multiple subtypes to generate broadly neutralizing antibodies (bNAbs). Soluble Env mimics, like the native flexibly linked (NFL) and SOSIP trimers, derived from the subtype A BG505 Env, form homogeneous, stable native-like trimers. However, other Env sequences, such as JRFL and 16055 from subtypes B and C, do so to a lesser degree. The high-resolution BG505 SOSIP crystal structures permit the identification and redesign of Env elements involved in trimer stability. Here, we identified structure trimer-derived (TD) residues that increased the propensity of the subtype B JRFL and subtype C 16055 Env sequences to form well-ordered, homogenous, and highly stable soluble trimers. The generation of these spike mimics no longer required antibody-based selection, positive or negative. Using the redesigned subtype B and C trimer representatives as respective foundations, we further stabilized the NFL TD trimers by engineering an intraprotomer disulfide linkage in the prebridging sheet, I201C-A433C (CC), that locks the gp120 in the receptor nontriggered state. We demonstrated that this disulfide pair prevented CD4 induced-conformational rearrangements in NFL trimers derived from the prototypic subtype A, B, and C representatives. Coupling the TD-based design with the engineered disulfide linkage, CC, increased the propensity of Env to form soluble highly stable spike mimics that are resistant to CD4-induced changes. These advances will allow testing of the hypothesis that such stabilized immunogens will more efficiently elicit neutralizing antibodies in small-animal models and primates. HIV-1 displays unprecedented global diversity circulating in the human population. Since the envelope glycoprotein (Env) is the target of neutralizing antibodies, Env-based vaccine candidates that address such diversity are needed. Soluble well-ordered Env mimics, typified by NFL and SOSIP trimers

  17. Multimeric and trimeric subunit SP-D are interconvertible structures with distinct ligand interaction

    DEFF Research Database (Denmark)

    Sørensen, Grith Lykke; Hoegh, Silje V; Leth-Larsen, Rikke

    2009-01-01

    -D compared to Met11 SP-D. Multimerization has proven important for enhancement of microbial phagocytosis. In the present study defined multimeric forms of Met11Thr SP-D were isolated from human amniotic fluid. Implementation of ManNAc-affinity chromatography allowed high recovery of natural trimeric SP......-D multimers. Trimeric SP-D subunits also showed greater binding to endogenous lipoproteins: LDL, oxLDL, and HDL, than multimeric SP-D. In conclusion, purified trimeric and multimeric SP-D represent separate and only partly interconvertible molecular populations with distinct biochemical properties....

  18. Electrochemical oxidation of 4-morpholinoaniline in aqueous solutions: Synthesis of a new trimer of 4-morpholinoaniline

    International Nuclear Information System (INIS)

    Esmaili, Roya; Nematollahi, Davood

    2011-01-01

    Research highlights: → Electrochemical study of 4-morpholinoaniline in various pHs. → Electrochemical trimerization of 4-morpholinoaniline in aqueous solution. → Green method for the synthesis of '4-morpholinoaniline-trimer'. → Potential-pH diagram for 4-morpholinoaniline. - Abstract: Electrochemical oxidation of 4-morpholinoaniline has been studied in various pHs using cyclic voltammetry and controlled-potential coulometry. The electrochemical trimerization of 4-morpholinoaniline is described and its mechanism has been studied in aqueous solution. This method provides a green, reagent-less, and environmentally friendly procedure with high atom economy, for the synthesis of '4-morpholinoaniline-trimer' using a carbon electrode in an undivided cell in good yield and purity.

  19. Cyclic water-trimer encapsulation into D2 (22)-C84 fullerene

    Science.gov (United States)

    Slanina, Zdeněk; Uhlík, Filip; Nagase, Shigeru; Akasaka, Takeshi; Lu, Xing; Adamowicz, Ludwik

    2018-03-01

    The cyclic water-trimer encapsulations into D2 (22)-C84 fullerene are evaluated. The encapsulation energy is computed at the M06-2X/6-31++G∗∗ level and it is found that the trimer storage in C84 yields the potential-energy gain of 10.4 kcal/mol. The encapsulated trimer can have two different forms, either the conformation known with the free gas-phase water trimer or the arrangement with the three non-hydrogen bonded H atoms on the same side of the O-O-O plane. The latter endohedral isomer is lower in the potential energy by 0.071 kcal/mol and forms about 57% of their equilibrium mixture at room temperature.

  20. Plasticity and Epitope Exposure of the HIV-1 Envelope Trimer.

    Science.gov (United States)

    Powell, Rebecca L R; Totrov, Maxim; Itri, Vincenza; Liu, Xiaomei; Fox, Alisa; Zolla-Pazner, Susan

    2017-09-01

    We recently showed that mutations in the HIV-1 envelope (Env) destabilize the V3 loop, rendering neutralization-resistant viruses sensitive to V3-directed monoclonal antibodies (MAbs). Here, we investigated the propagation of this effect on other Env epitopes, with special emphasis on V2 loop exposure. Wild-type JR-FL and 19 mutant JR-FL pseudoviruses were tested for neutralization sensitivity to 21 MAbs specific for epitopes in V2, the CD4 binding site (CD4bs), and the CD4-induced (CD4i) region. Certain glycan mutants, mutations in the gp120 hydrophobic core, and mutations in residues involved in intraprotomer interactions exposed epitopes in the V2i region (which overlies the α4β7 integrin binding site) and the V3 crown, suggesting general destabilization of the distal region of the trimer apex. In contrast, other glycan mutants, mutations affecting interprotomer interactions, and mutations affecting the CD4bs exposed V3 but not V2i epitopes. These data indicate for the first time that V3 can move independently of V2, with V3 pivoting out from its "tucked" position in the trimer while apparently leaving the V2 apex intact. Notably, none of the mutations exposed V2 epitopes without also exposing V3, suggesting that movement of V2 releases V3. Most mutations increased sensitivity to CD4bs-directed MAbs without exposure of the CD4i epitope, implying these mutations facilitate the trimers' maintenance of an intermediate energy state between open and closed conformations. Taken together, these data indicate that several transient Env epitopes can be rendered more accessible to antibodies (Abs) via specific mutations, and this may facilitate the design of V1V2-targeting immunogens. IMPORTANCE Many epitopes of the HIV envelope (Env) spike are relatively inaccessible to antibodies (Abs) compared to their exposure in the open Env conformation induced by receptor binding. However, the reduced infection rate that resulted from the vaccine used in the RV144 HIV-1 vaccine

  1. Detection of peptides by precolumn derivatization with biuret reagent and preconcentration on capillary liquid chromatography columns with electrochemical detection.

    Science.gov (United States)

    Shen, H; Witowski, S R; Boyd, B W; Kennedy, R T

    1999-03-01

    The separation and detection of biuret complexes of neuropeptides by capillary liquid chromatography with electrochemical detection was explored. Capillaries of 25-micron inner diameter packed with base-resistant, polymer-based reversed-phase particles were used for separation, and C-fiber electrodes were used for detection. Detection at the C-fiber electrode was found to have some differences in relative sensitivity for peptides compared to glassy carbon electrodes used previously. On-column preconcentration of preformed complexes allowed up to 1-microL samples to be injected with minimal band broadening resulting in a 100-fold improvement in concentration detection limit with no effect on mass detection limit. Concentration detection limits ranged from 5 to 59 pM, depending upon the peptide, corresponding to 5-59 amol injected. The low concentration detection limit was possible because of minimal baseline disturbances, minimal formation of unwanted products, and high efficiency of complex formation associated with biuret derivatization. The method was applied to determination of vasopressin and bradykinin in dialysates collected with 5-min sampling frequency from the rat supraoptic nucleus.

  2. Design of lipid nanocapsule delivery vehicles for multivalent display of recombinant Env trimers in HIV vaccination.

    Science.gov (United States)

    Pejawar-Gaddy, Sharmila; Kovacs, James M; Barouch, Dan H; Chen, Bing; Irvine, Darrell J

    2014-08-20

    Immunization strategies that elicit antibodies capable of neutralizing diverse virus strains will likely be an important part of a successful vaccine against HIV. However, strategies to promote robust humoral responses against the native intact HIV envelope trimer structure are lacking. We recently developed chemically cross-linked lipid nanocapsules as carriers of molecular adjuvants and encapsulated or surface-displayed antigens, which promoted follicular helper T-cell responses and elicited high-avidity, durable antibody responses to a candidate malaria antigen. To apply this system to the delivery of HIV antigens, Env gp140 trimers with terminal his-tags (gp140T-his) were anchored to the surface of lipid nanocapsules via Ni-NTA-functionalized lipids. Initial experiments revealed that the large (409 kDa), heavily glycosylated trimers were capable of extracting fluid phase lipids from the membranes of nanocapsules. Thus, liquid-ordered and/or gel-phase lipid compositions were required to stably anchor trimers to the particle membranes. Trimer-loaded nanocapsules combined with the clinically relevant adjuvant monophosphoryl lipid A primed high-titer antibody responses in mice at antigen doses ranging from 5 μg to as low as 100 ng, whereas titers dropped more than 50-fold over the same dose range when soluble trimer was mixed with a strong oil-in-water adjuvant comparator. Nanocapsule immunization also broadened the number of distinct epitopes on the HIV trimer recognized by the antibody response. These results suggest that nanocapsules displaying HIV trimers in an oriented, multivalent presentation can promote key aspects of the humoral response against Env immunogens.

  3. Daily and alternate-day supplementation of urea or biuret to ruminants consuming low-quality forage: II. Effects on site of digestion and microbial efficiency in steers.

    Science.gov (United States)

    Currier, T A; Bohnert, D W; Falck, S J; Schauer, C S; Bartle, S J

    2004-05-01

    Five steers (491 +/- 21 kg BW) were used in an incomplete 5 x 4 Latin square with four 24-d periods to determine the influence of supplemental non-protein N (NPN) source and supplementation frequency (SF) on nutrient intake and site of digestion in steers consuming low-quality grass straw (4% CP). Treatments (TRT) included an unsupplemented control and a urea- or biuret-containing supplement placed directly into the rumen daily (D) or every other day (2D) at 0700. The NPN treatments were formulated to provide 90% of the estimated degradable intake protein requirement. Daily TRT were supplemented CP at 0.04% of BW/d, whereas the 2D TRT were supplemented at 0.08% of BW every other day. Therefore, all supplemented TRT received the same quantity of supplemental CP over a 2-d period. Forage OM intake was not affected (P > 0.05) by NPN supplementation, NPN source, or SF; however, total OM and N intake were increased (P biuret compared with urea (P biuret than for urea. Apparent total-tract N digestibility was increased with NPN supplementation (P biuret can be used effectively as a supplemental N source by steers consuming low-quality forage.

  4. Crystal structures and vibrational spectra of biuret co-crystals with cyanuric and glutaric acids, discussion of hydrogen bonding involving carbonyl groups

    Czech Academy of Sciences Publication Activity Database

    Matulková, I.; Mathauserová, J.; Císařová, I.; Němec, I.; Fábry, Jan

    2016-01-01

    Roč. 231, č. 5 (2016), s. 291-300 ISSN 2194-4946 R&D Projects: GA ČR GA14-05506S Institutional support: RVO:68378271 Keywords : biuret * crystal structure analysis * hydrogen bonding * vibrational spectroscopy * X-ray diffraction Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 3.179, year: 2016

  5. Improving the Immunogenicity of Native-like HIV-1 Envelope Trimers by Hyperstabilization

    Directory of Open Access Journals (Sweden)

    Alba Torrents de la Peña

    2017-08-01

    Full Text Available The production of native-like recombinant versions of the HIV-1 envelope glycoprotein (Env trimer requires overcoming the natural flexibility and instability of the complex. The engineered BG505 SOSIP.664 trimer mimics the structure and antigenicity of native Env. Here, we describe how the introduction of new disulfide bonds between the glycoprotein (gp120 and gp41 subunits of SOSIP trimers of the BG505 and other genotypes improves their stability and antigenicity, reduces their conformational flexibility, and helps maintain them in the unliganded conformation. The resulting next-generation SOSIP.v5 trimers induce strong autologous tier-2 neutralizing antibody (NAb responses in rabbits. In addition, the BG505 SOSIP.v6 trimers induced weak heterologous NAb responses against a subset of tier-2 viruses that were not elicited by the prototype BG505 SOSIP.664. These stabilization methods can be applied to trimers from multiple genotypes as components of multivalent vaccines aimed at inducing broadly NAbs (bNAbs.

  6. Different Infectivity of HIV-1 Strains Is Linked to Number of Envelope Trimers Required for Entry

    Science.gov (United States)

    Brandenberg, Oliver F.; Magnus, Carsten; Rusert, Peter; Regoes, Roland R.; Trkola, Alexandra

    2015-01-01

    HIV-1 enters target cells by virtue of envelope glycoprotein trimers that are incorporated at low density in the viral membrane. How many trimers are required to interact with target cell receptors to mediate virus entry, the HIV entry stoichiometry, still awaits clarification. Here, we provide estimates of the HIV entry stoichiometry utilizing a combined approach of experimental analyses and mathematical modeling. We demonstrate that divergent HIV strains differ in their stoichiometry of entry and require between 1 to 7 trimers, with most strains depending on 2 to 3 trimers to complete infection. Envelope modifications that perturb trimer structure lead to an increase in the entry stoichiometry, as did naturally occurring antibody or entry inhibitor escape mutations. Highlighting the physiological relevance of our findings, a high entry stoichiometry correlated with low virus infectivity and slow virus entry kinetics. The entry stoichiometry therefore directly influences HIV transmission, as trimer number requirements will dictate the infectivity of virus populations and efficacy of neutralizing antibodies. Thereby our results render consideration of stoichiometric concepts relevant for developing antibody-based vaccines and therapeutics against HIV. PMID:25569556

  7. Possible novel therapy for malignant gliomas with secretable trimeric TRAIL.

    Directory of Open Access Journals (Sweden)

    Moonsup Jeong

    Full Text Available Malignant gliomas are the most common primary brain tumors. Despite intensive clinical investigation and many novel therapeutic approaches, average survival for the patients with malignant gliomas is only about 1 year. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL has shown potent and cancer-selective killing activity and drawn considerable attention as a promising therapy for cancers, but concerns over delivery and toxicity have limited progress. We have developed a secretable trimeric TRAIL (stTRAIL and here evaluated the therapeutic potential of this stTRAIL-based gene therapy in brain tumors. An adenovirus (Ad-stTRAIL delivering stTRAIL was injected into intra-cranial human glioma tumors established in nude mice and tumor growth monitored using the magnetic resonance imaging (MRI. Ad-stTRAIL gene therapy showed potent tumor suppressor activity with no toxic side effects at therapeutically effective doses. When compared with 1, 3-bis(2-chloroethyl-1-nitrosourea (BCNU, a conventional therapy for malignant gliomas, Ad-stTRAIL suppressed tumor growth more potently. The combination of Ad-stTRAIL and BCNU significantly increased survival compared to the control mice or mice receiving Ad-stTRAIL alone. Our data indicate that Ad-stTRAIL, either alone or combined with BCNU, has promise as a novel therapy for malignant gliomas.

  8. Molecular beam magnetic deflection behavior of sodium trimers

    International Nuclear Information System (INIS)

    George, A.R.

    1983-01-01

    The observation and characterization of the Stern-Gerlach magnetic deflection behavior of sodium trimers in a supersonic molecular beam is reported. As part of a program to apply molecular beam technique to the study of metal clusters, a molecular beam apparatus designed for magnetic deflection and resonance experiments on selected alkali metal cluster species has been developed and is described. Clusters are produced in a supersonic expansion of a pure metal vapor, and are detected mass selectively by photoionization, quadrupole mass analysis, and an ion counting detector. The deflection profiles reveal peaks corresponding to the one Bohr magneton of magnetic moment of the unpaired electron, but in addition show evidence of a distribution of effective magnetic moments extending the full range between the positive and negative one Bohr magneton peaks. In addition, experiments utilizing multiple magnets and trajectory selecting collimators show evidence for magnetic moment and molecular state changes during traversal through the apparatus. Information from time of flight velocity analysis is used in conjunction with the deflection data and with computer simulations to rule out experimental artifacts and to establish that the observed phenomena can be the result of magnetic moment changes and molecular state changes caused by adiabatic and non-adiabatic traversals of avoided level crossings in the Zeeman energy diagram of these molecules. The phenomena have implications for the application of molecular beam Electron Spin Resonance technique to polyatomic molecules

  9. Structure and immune recognition of trimeric prefusion HIV-1 Env

    Science.gov (United States)

    Pancera, Marie; Zhou, Tongqing; Druz, Aliaksandr; Georgiev, Ivelin S.; Soto, Cinque; Gorman, Jason; Huang, Jinghe; Acharya, Priyamvada; Chuang, Gwo-Yu; Ofek, Gilad; Stewart-Jones, Guillaume B. E.; Stuckey, Jonathan; Bailer, Robert T.; Joyce, M. Gordon; Louder, Mark K.; Tumba, Nancy; Yang, Yongping; Zhang, Baoshan; Cohen, Myron S.; Haynes, Barton F.; Mascola, John R.; Morris, Lynn; Munro, James B.; Blanchard, Scott C.; Mothes, Walther; Connors, Mark; Kwong, Peter D.

    2015-01-01

    The HIV-1-envelope (Env) spike, comprising three gp120 and three gp41 subunits, is a conformational machine that facilitates HIV-1 entry by rearranging from a mature unliganded state, through receptor-bound intermediates, to a postfusion state. As the sole viral antigen on the HIV-1-virion surface, Env is both the target of neutralizing antibodies and a focus of vaccine efforts. Here we report the structure at 3.5-Å resolution for an HIV-1-Env trimer captured in a mature closed state by antibodies PGT122 and 35O22. This structure reveals the prefusion conformation of gp41, indicates rearrangements needed for fusion activation, and defines parameters of immune evasion and immune recognition. Prefusion gp41 encircles N- and C-terminal strands of gp120 with four helices that form a membrane-proximal collar, fastened by insertion of a fusion peptide-proximal methionine into a gp41-tryptophan clasp. Spike rearrangements required for entry likely involve opening the clasp and expelling the termini. N-linked glycosylation and sequence-variable regions cover the prefusion closed spike: we used chronic cohorts to map the prevalence and location of effective HIV-1-neutralizing responses, which were distinguished by their recognition of N-linked glycan and tolerance for epitope-sequence variation. PMID:25296255

  10. Spin and orbital magnetism of coinage metal trimers (Cu3, Ag3, Au3: A relativistic density functional theory study

    Directory of Open Access Journals (Sweden)

    Mahdi Afshar

    2013-11-01

    Full Text Available We have demonstrated electronic structure and magnetic properties of Cu3, Ag3 and Au3 trimers using a full potential local orbital method in the framework of relativistic density functional theory. We have also shown that the non-relativistic generalized gradient approximation for the exchange-correlation energy functional gives reliable magnetic properties in coinage metal trimers compared to experiment. In addition we have indicated that the spin-orbit coupling changes the structure and magnetic properties of gold trimer while the structure and magnetic properties of copper and silver trimers are marginally affected. A significant orbital moment of 0.21μB was found for most stable geometry of the gold trimer whereas orbital magnetism is almost quenched in the copper and silver trimers.

  11. A multivalent clade C HIV-1 Env trimer cocktail elicits a higher magnitude of neutralizing antibodies than any individual component.

    Science.gov (United States)

    Bricault, Christine A; Kovacs, James M; Nkolola, Joseph P; Yusim, Karina; Giorgi, Elena E; Shields, Jennifer L; Perry, James; Lavine, Christy L; Cheung, Ann; Ellingson-Strouss, Katharine; Rademeyer, Cecelia; Gray, Glenda E; Williamson, Carolyn; Stamatatos, Leonidas; Seaman, Michael S; Korber, Bette T; Chen, Bing; Barouch, Dan H

    2015-03-01

    The sequence diversity of human immunodeficiency virus type 1 (HIV-1) presents a formidable challenge to the generation of an HIV-1 vaccine. One strategy to address such sequence diversity and to improve the magnitude of neutralizing antibodies (NAbs) is to utilize multivalent mixtures of HIV-1 envelope (Env) immunogens. Here we report the generation and characterization of three novel, acute clade C HIV-1 Env gp140 trimers (459C, 405C, and 939C), each with unique antigenic properties. Among the single trimers tested, 459C elicited the most potent NAb responses in vaccinated guinea pigs. We evaluated the immunogenicity of various mixtures of clade C Env trimers and found that a quadrivalent cocktail of clade C trimers elicited a greater magnitude of NAbs against a panel of tier 1A and 1B viruses than any single clade C trimer alone, demonstrating that the mixture had an advantage over all individual components of the cocktail. These data suggest that vaccination with a mixture of clade C Env trimers represents a promising strategy to augment vaccine-elicited NAb responses. It is currently not known how to generate potent NAbs to the diverse circulating HIV-1 Envs by vaccination. One strategy to address this diversity is to utilize mixtures of different soluble HIV-1 envelope proteins. In this study, we generated and characterized three distinct, novel, acute clade C soluble trimers. We vaccinated guinea pigs with single trimers as well as mixtures of trimers, and we found that a mixture of four trimers elicited a greater magnitude of NAbs than any single trimer within the mixture. The results of this study suggest that further development of Env trimer cocktails is warranted. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  12. Preparation and properties of a coated slow-release and water-retention biuret phosphoramide fertilizer with superabsorbent.

    Science.gov (United States)

    Jin, Shuping; Yue, Guoren; Feng, Lei; Han, Yuqi; Yu, Xinghai; Zhang, Zenghu

    2011-01-12

    In this investigation, a novel water-insoluble slow-release fertilizer, biuret polyphosphoramide (BPAM), was formulated and synthesized from urea, phosphoric acid (H(3)PO(4)), and ferric oxide (Fe(2)O(3)). The structure of BPAM was characterized by Fourier transform infrared (FTIR) spectroscopy. Subsequently, a coated slow-release BPAM fertilizer with superabsorbent was prepared by ionic cross-linked carboxymethylchitosan (the core), acrylic acid, acrylamide, and active carbon (the coating). The variable influences on the water absorbency were investigated and optimized. Component analysis results showed that the coated slow-release BPAM contained 5.66% nitrogen and 11.7% phosphorus. The property of water retention, the behavior of slow release of phosphorus, and the capacity of adsorption of cations were evaluated, and the results revealed that the product not only had good slow-release property and excellent water retention capacity but also higher adsorption capacities of cations in saline soil.

  13. Spectral flow of trimer states of two heavy impurities and one light condensed boson

    DEFF Research Database (Denmark)

    Zinner, Nikolaj Thomas

    2014-01-01

    The spectral flow of three-body (trimer) states consisting of two heavy (impurity) particles sitting in a condensate of light bosons is considered. Assuming that the condensate is weakly interaction and that an impurity and a boson have a zero-range two-body interaction, we use the Born-Oppenheim......The spectral flow of three-body (trimer) states consisting of two heavy (impurity) particles sitting in a condensate of light bosons is considered. Assuming that the condensate is weakly interaction and that an impurity and a boson have a zero-range two-body interaction, we use the Born......-Oppenheimer approximation to determine the effective three-body potential. We solve the resulting Schr\\"odinger equation numerically and determine the trimer binding energies as a function of the coherence length of the light bosonic condensate particles. The binding energy is found to be suppressed by the presence...... of the condensate when the energy scale corresponding to the coherence length becomes of order the trimer binding energy in the absence of the condensate. We find that the Efimov scaling property is reflected in the critical values of the condensate coherence length at which the trimers are pushed...

  14. Efficient purification protocol for bioengineering allophycocyanin trimer with N-terminus Histag

    Directory of Open Access Journals (Sweden)

    Wenjun Li

    2017-03-01

    Full Text Available Allophycocyanin plays a key role for the photon energy transfer from the phycobilisome to reaction center chlorophylls with high efficiency in cyanobacteria. Previously, the high soluble self-assembled bioengineering allophycocyanin trimer with N-terminus polyhistidine from Synechocystis sp. PCC 6803 had been successfully recombined and expressed in Escherichia coli strain. The standard protocol with immobilized metal-ion affinity chromatography with chelating transition metal ion (Ni2+ was used to purify the recombinant protein. Extensive optimization works were performed to obtain the desired protocol for high efficiency, low disassociation, simplicity and fitting for large-scale purification. In this study, a 33 full factorial response surface methodology was employed to optimize the varied factors such as pH of potassium phosphate (X1, NaCl concentration (X2, and imidazole concentration (X3. A maximum trimerization ratio (Y1 of approximate A650 nm/A620 nm at 1.024 was obtained at these optimum parameters. Further examinations, with absorbance spectra, fluorescence spectra and SDS-PAGE, confirmed the presence of bioengineering allophycocyanin trimer with highly trimeric form. All these results demonstrate that optimized protocol is efficient in purification of bioengineering allophycocyanin trimer with Histag.

  15. Novel metamaterial based on the coupling effect of a dielectric trimer

    Energy Technology Data Exchange (ETDEWEB)

    Fu, Jiahui; Chen, Wan, E-mail: dhtyyobdc@126.com; Lv, Bo; Wang, Zhefei

    2017-01-23

    Highlights: • Novel metamaterial based on the coupling effect of a dielectric trimer is proposed. • The phenomenon of vanishing mode is explained by the zero-sum effect. • Due to the vanishing mode, the bandwidth of the dielectric trimer has been expanded to 37%. - Abstract: In this paper, a novel periodic 2D all-dielectric metamaterial based on dielectric trimer is proposed. The electromagnetic responses are explained by the corrected equations of motion using coupled mode theory (CMT). An abnormal vanishment mode phenomenon is also discovered and explained using the zero-sum effect of magnetic dipole, by which the relative bandwidth of the metamaterial has been improved significantly compared with other structures. The presented design is easy for fabrication and can be applied in microwave region by scaling the dimensions of the cubes.

  16. Symmetry-Breaking Effect on the Electromagnetic Properties of Plasmonic Trimers Composed of Graphene Nanodisks

    Directory of Open Access Journals (Sweden)

    Weibin Qiu

    2018-03-01

    Full Text Available Plasmonic trimers composed of equal-sized graphene nanodisks are proposed in this paper. The symmetry-breaking effect on the electromagnetic properties of the nanostructure is numerically investigated by studying plasmon energy diagrams and optical scattering spectra in mid-infrared range with a gradient vertex angle. The degenerate plasmonic modes are lifted and new modes appear with increased vertex angle. The energy diagrams are consistent with scattering extinction spectra, about which the dipole moment distribution of the proposed structure is discussed to demonstrate the coupling strength of the collective plasmonic modes of the trimer. More specifically, the frequency tunability of the plasmonic trimer is pointed out by modifying the chemical potential of the graphene nanodisks without varying the geometric configuration. The proposed structure might find applications such as light-matter interaction, single molecule detection, and high-sensitivity chemical sensing.

  17. Structure-based Design of Cyclically Permuted HIV-1 gp120 Trimers That Elicit Neutralizing Antibodies.

    Science.gov (United States)

    Kesavardhana, Sannula; Das, Raksha; Citron, Michael; Datta, Rohini; Ecto, Linda; Srilatha, Nonavinakere Seetharam; DiStefano, Daniel; Swoyer, Ryan; Joyce, Joseph G; Dutta, Somnath; LaBranche, Celia C; Montefiori, David C; Flynn, Jessica A; Varadarajan, Raghavan

    2017-01-06

    A major goal for HIV-1 vaccine development is an ability to elicit strong and durable broadly neutralizing antibody (bNAb) responses. The trimeric envelope glycoprotein (Env) spikes on HIV-1 are known to contain multiple epitopes that are susceptible to bNAbs isolated from infected individuals. Nonetheless, all trimeric and monomeric Env immunogens designed to date have failed to elicit such antibodies. We report the structure-guided design of HIV-1 cyclically permuted gp120 that forms homogeneous, stable trimers, and displays enhanced binding to multiple bNAbs, including VRC01, VRC03, VRC-PG04, PGT128, and the quaternary epitope-specific bNAbs PGT145 and PGDM1400. Constructs that were cyclically permuted in the V1 loop region and contained an N-terminal trimerization domain to stabilize V1V2-mediated quaternary interactions, showed the highest homogeneity and the best antigenic characteristics. In guinea pigs, a DNA prime-protein boost regimen with these new gp120 trimer immunogens elicited potent neutralizing antibody responses against highly sensitive Tier 1A isolates and weaker neutralizing antibody responses with an average titer of about 115 against a panel of heterologous Tier 2 isolates. A modest fraction of the Tier 2 virus neutralizing activity appeared to target the CD4 binding site on gp120. These results suggest that cyclically permuted HIV-1 gp120 trimers represent a viable platform in which further modifications may be made to eventually achieve protective bNAb responses. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  18. Reaction of some ambident nucleophiles, thiourea, substituted thioureas, biuret and oxamide with uranyl nitrate and chloride and isolation of some novel adducts

    International Nuclear Information System (INIS)

    Bhattacharyya, R.G.; Doloi, N.C.

    1977-01-01

    The ambident nucleophiles, viz. thiourea (Tu), monobenzoylthiourea (MBTu), oxalythiourea (OxTu), biuret (BuH 2 ) and oxamide (OxamH 2 ) react with UO 2 (NO 3 ) 2 6H 2 O in acetonitrile medium via base hydrolysis affording hydroxo complexes of the types UO 2 (OH) 2 Tu, UO 2 (OH) 2 MBTu, UO 2 (OH) 2 OxTu, UO 2 (OH) 2 3BuH 2 and UO 2 (OH) 2 (OxamH 2 ) 2 . However UO 2 Cl 2 reacts with Tu and BuH 2 forming adducts UO 2 Cl 2 Tu and UO 2 Cl 2 BuH 2 respectively. The IR spectra reveal that thiourea is S-bonded to uranium and in the chloro complex biuret behaves as oxygen donor chelate while in the other case it is probably acting as a unidentate ligand. Oxamide behaves as an oxygen donor neutral chelating ligand. (author)

  19. An improved modification of the biuret method for the determination of protein in turbid materials with high lipid and hemoglobin content.

    Science.gov (United States)

    Steinbrecht, I; Augustin, W

    1983-01-01

    The quantitative determination of protein by means of the biuret method frequently yields erroneous values, especially when applied to turbid, lipid- and hemoglobin-containing materials. These errors can only partially be abolished either by the addition of detergents or by destroying the Cu-protein complex by KCN addition. It was found that most disturbances were almost completely eliminated after prior precipitation of protein by the addition of Triton X-100 to the solubilizing biuret reagent and absorbance measurements being performed at a wavelength of 572 nm before and after the addition of KCN. Values of protein determinations according to the proposed assay and protein concentrations calculated from Kjeldahl nitrogen determinations have been shown to agree fairly satisfactory. The proposed assay represents a relatively simple and versatile approach for the evaluation of protein concentrations in a variety of materials containing lipids, hemoglobin and of some other turbidities as well.

  20. Non-Hermitian trimers: PT-symmetry versus pseudo-Hermiticity

    Science.gov (United States)

    Suchkov, Sergey V.; Fotsa-Ngaffo, Fernande; Kenfack-Jiotsa, Aurelien; Tikeng, Arnaud D.; Kofane, Timoleon C.; Kivshar, Yuri S.; Sukhorukov, Andrey A.

    2016-06-01

    We study a structure composed of three coupled waveguides with gain and loss, a non-Hermitian trimer. We demonstrate that the mode spectrum can be entirely real if the waveguides are placed in a special order and at certain distances between each other. Such structures generally lack a spatial symmetry, in contrast to parity-time symmetric trimers which are known to feature a real spectrum. We also determine a threshold for wave amplification and analyse the scattering properties of such non-conservative systems embedded into a chain of conservative waveguides.

  1. Selected HIV-1 Env trimeric formulations act as potent immunogens in a rabbit vaccination model

    DEFF Research Database (Denmark)

    Heyndrickx, Leo; Stewart-Jones, Guillaume; Jansson, Marianne Bendixen

    2013-01-01

    Ten to 30% of HIV-1 infected subjects develop broadly neutralizing antibodies (bNAbs) during chronic infection. We hypothesized that immunizing rabbits with viral envelope glycoproteins (Envs) from these patients may induce bNAbs, when formulated as a trimeric protein and in the presence of an ad......Ten to 30% of HIV-1 infected subjects develop broadly neutralizing antibodies (bNAbs) during chronic infection. We hypothesized that immunizing rabbits with viral envelope glycoproteins (Envs) from these patients may induce bNAbs, when formulated as a trimeric protein and in the presence...

  2. Structure of the Catalytic Trimer of Methanococcus jannaschii Aspartate Transcarbamoylase in an Orthorhombic Crystal Form

    Energy Technology Data Exchange (ETDEWEB)

    Vitali,J.; Colaneri, M.

    2008-01-01

    Crystals of the catalytic subunit of Methanococcus jannaschii aspartate transcarbamoylase in an orthorhombic crystal form contain four crystallographically independent trimers which associate in pairs to form stable staggered complexes that are similar to each other and to a previously determined monoclinic C2 form. Each subunit has a sulfate in the central channel. The catalytic subunits in these complexes show flexibility, with the elbow angles of the monomers differing by up to 7.4 between crystal forms. Moreover, there is also flexibility in the relative orientation of the trimers around their threefold axis in the complexes, with a difference of 4 between crystal forms.

  3. Structure of the catalytic trimer of Methanococcus jannaschii aspartate transcarbamoylase in an orthorhombic crystal form.

    Science.gov (United States)

    Vitali, Jacqueline; Colaneri, Michael J

    2008-09-01

    Crystals of the catalytic subunit of Methanococcus jannaschii aspartate transcarbamoylase in an orthorhombic crystal form contain four crystallographically independent trimers which associate in pairs to form stable staggered complexes that are similar to each other and to a previously determined monoclinic C2 form. Each subunit has a sulfate in the central channel. The catalytic subunits in these complexes show flexibility, with the elbow angles of the monomers differing by up to 7.4 degrees between crystal forms. Moreover, there is also flexibility in the relative orientation of the trimers around their threefold axis in the complexes, with a difference of 4 degrees between crystal forms.

  4. Glycosylation Benchmark Profile for HIV-1 Envelope Glycoprotein Production Based on Eleven Env Trimers.

    Science.gov (United States)

    Go, Eden P; Ding, Haitao; Zhang, Shijian; Ringe, Rajesh P; Nicely, Nathan; Hua, David; Steinbock, Robert T; Golabek, Michael; Alin, James; Alam, S Munir; Cupo, Albert; Haynes, Barton F; Kappes, John C; Moore, John P; Sodroski, Joseph G; Desaire, Heather

    2017-05-01

    HIV-1 envelope glycoprotein (Env) glycosylation is important because individual glycans are components of multiple broadly neutralizing antibody epitopes, while shielding other sites that might otherwise be immunogenic. The glycosylation on Env is influenced by a variety of factors, including the genotype of the protein, the cell line used for its expression, and the details of the construct design. Here, we used a mass spectrometry (MS)-based approach to map the complete glycosylation profile at every site in multiple HIV-1 Env trimers, accomplishing two goals. (i) We determined which glycosylation sites contain conserved glycan profiles across many trimeric Envs. (ii) We identified the variables that impact Env's glycosylation profile at sites with divergent glycosylation. Over half of the gp120 glycosylation sites on 11 different trimeric Envs have a conserved glycan profile, indicating that a native consensus glycosylation profile does indeed exist among trimers. We showed that some soluble gp120s and gp140s exhibit highly divergent glycosylation profiles compared to trimeric Env. We also assessed the impact of several variables on Env glycosylation: truncating the full-length Env; producing Env, instead of the more virologically relevant T lymphocytes, in CHO cells; and purifying Env with different chromatographic platforms, including nickel-nitrilotriacetic acid (Ni-NTA), 2G12, and PGT151 affinity. This report provides the first consensus glycosylation profile of Env trimers, which should serve as a useful benchmark for HIV-1 vaccine developers. This report also defines the sites where glycosylation may be impacted when Env trimers are truncated or produced in CHO cells. IMPORTANCE A protective HIV-1 vaccine will likely include a recombinant version of the viral envelope glycoprotein (Env). Env is highly glycosylated, and yet vaccine developers have lacked guidance on how to assess whether their immunogens have optimal glycosylation. The following important

  5. Substituted biurets as uranophilic ligands: A facile DMSO-induced conversion of a 1:1 into a 2:1 uranyl-ligand complex

    Energy Technology Data Exchange (ETDEWEB)

    Potts, K.T.; O' Brien, J.J.; Tham, F.S. (Rensselaer Polytechnic Inst., Troy, NY (United States))

    1990-01-01

    1,5-Bis(6-(1-ethoxycarbonyl-3-thioureido)-2-pyridindiyl)biuret and uranyl acetate gave a crystalline 1:1 uranyl-ligand complex which, on crystallization from DMSO, underwent rearrangement to a crystalline 2:1 uranyl-ligand complex and a stoichiometric amount of the uncomplexed ligand. Spectral characteristics of these ligands and their uranyl complexes together with single crystal x-ray data for the uranyl-ligand complexes are described.

  6. Daily and alternate-day supplementation of urea or biuret to ruminants consuming low-quality forage: III. Effects on ruminal fermentation characteristics in steers.

    Science.gov (United States)

    Currier, T A; Bohnert, D W; Falck, S J; Schauer, C S; Bartle, S J

    2004-05-01

    Five ruminally and duodenally cannulated steers (491 +/- 21 kg BW) were used in an incomplete 5 x 4 Latin square with four 24-d periods to determine the influence of supplemental nonprotein N (NPN) source and supplementation frequency (SF) on the dynamics of ruminal fermentation in steers consuming low-quality grass straw (4% CP). Treatments (TRT) included an unsupplemented control (CON) and a urea or biuret supplement that were placed directly into the rumen at 0700 daily (D) or every other day (2D). The NPN treatments were formulated to provide 90% of the estimated degradable intake protein requirement; therefore, the urea and biuret treatments received the same amount of supplemental N over a 2-d period. Daily TRT were supplemented with CP at 0.04% of BW/d, whereas the 2D TRT were supplemented at 0.08% of BW every other day. Forage was provided at 120% of the previous 5-d average intake in two equal portions at 0715 and 1900. Ruminal fluid was collected 0, 3, 6, 9, 12, and 24 h after supplementation on a day of and a day before supplementation for all TRT. Ruminal NH3-N increased (P biuret. Ruminal NH3-N on the day only daily supplements were provided was greater (P = 0.02) for D compared with 2D. On the day all supplements were provided, D increased (P = 0.05) ruminal indigestible acid detergent fiber passage rate and ruminal fluid volume compared with 2D. These results suggest that urea or biuret can be used effectively as a supplemental N source by steers consuming low-quality forage without adversely affecting ruminal fermentation, even when provided every other day.

  7. Application of an improved biuret method to the determination of total protein in urine and cerebrospinal fluid without concentration step by use of Hitachi 7170 auto-analyzer.

    Science.gov (United States)

    Guobing, X; Lili, J; Lihua, Z; Tiean, X

    2001-01-01

    A biuret automated colorimetric assay for total protein in urine and cerebrospinal fluids was established. The procedures were as follows. Acidify all urine sample before analysis. Add precipitant Na(2)WO(4) to urine samples. After 10 min, centrifuge, decant the supernatant fluid, drain the inverted tubes on absorbent tissue, dissolve the precipitation with 0.1 mol/L NaOH, and finally adapt the reconstituted urine to the Hitachi 7170 analyzer. A cell-free cerebrospinal fluid sample produced by centrifugation can be inserted in an auto-analyzer for protein measurement directly. The program: mix 35 microl sample (CSF or reconstituted urine) and standard with 0.2 mol/L NaOH; incurable at 37 degrees C for 5 min, and real A1. Add concentrated biuret reagent, and 10 min later measure absorbance A2 at 546 nm vs. reagent blank. Secondary wavelength was 700 nm. The test results were calculated against a one-point standard. This biuret colorimetric method was relatively simple, fast, and accurate for the determination of protein in urine and cerebrospinal fluid, with a wide linearity extending from 0.125 g/L up to 6 g/L, had a good correlation with Benzethonium chloride turbidimetry technique, and was a practical routine method. Copyright 2001 Wiley-Liss, Inc.

  8. 3-(2,3,5,6,7,8-Hexahydro-1H-cyclo-penta-[b]quinolin-9-yl)-1,5-bis-(4-methoxy-phen-yl)biuret.

    Science.gov (United States)

    Sakurai, Kaori; Noguchi, Keiichi; Nishibe, Koichiro

    2010-02-20

    Ipidacrine (2,3,5,6,7,8-hexa-hydro-1H-cyclo-penta-[b]quinolin-9-amine) was reacted with 4-methoxy-phenyl isocyanate to give the title compound, C(28)H(30)N(4)O(4). An intra-molecular N-H⋯O hydrogen bond results in an essentially planar [r.m.s. deviation from the mean plane is 0.126 (1) Å] conformation for the biuret unit. The central ring of the quinoline unit is twisted by 78.2 (1)° with respect to the biuret mean plane, whereas the two 4-methoxy-benzene rings are twisted out of this plane by 24.3 (1)° and 48.5 (1)°, resulting in an overall propeller-like structure. An inter-molecular N-H⋯N hydrogen bond between the biuret NH atom and the quinoline ring nitro-gen defines the crystal packing.

  9. Structure of Acidic pH Dengue Virus Showing the Fusogenic Glycoprotein Trimers

    NARCIS (Netherlands)

    Zhang, Xinzheng; Sheng, Ju; Austin, S. Kyle; Hoornweg, Tabitha E.; Smit, Jolanda M.; Kuhn, Richard J.; Diamond, Michael S.; Rossmann, Michael G.

    Flaviviruses undergo large conformational changes during their life cycle. Under acidic pH conditions, the mature virus forms transient fusogenic trimers of E glycoproteins that engage the lipid membrane in host cells to initiate viral fusion and nucleocapsid penetration into the cytoplasm. However,

  10. Unusual H-Bond Topology and Bifurcated H-bonds in the 2-Fluoroethanol Trimer.

    Science.gov (United States)

    Thomas, Javix; Liu, Xunchen; Jäger, Wolfgang; Xu, Yunjie

    2015-09-28

    By using a combination of rotational spectroscopy and ab initio calculations, an unusual H-bond topology was revealed for the 2-fluoroethanol trimer. The trimer exhibits a strong heterochiral preference and adopts an open OH⋅⋅⋅OH H-bond topology while utilizing two types of bifurcated H-bonds involving organic fluorine. This is in stark contrast to the cyclic OH⋅⋅⋅OH H-bond topology adopted by trimers of water and other simple alcohols. The strengths of different H-bonds in the trimer were analyzed by using the quantum theory of atoms in molecules. The study showcases a remarkable example of a chirality-induced switch in H-bond topology in a simple transient chiral fluoroalcohol. It provides important insight into the H-bond topologies of small fluoroalcohol aggregates, which are proposed to play a key role in protein folding and in enantioselective reactions and separations where fluoroalcohols serve as a (co)solvent. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  11. Two mechanisms for dissipation of excess light in monomeric and trimeric light-harvesting complexes

    Energy Technology Data Exchange (ETDEWEB)

    Dall' Osto, Luca [Univ. di Verona, Verona (Italy). Dipartimento di Biotecnologie; Cazzaniga, Stefano [Univ. di Verona, Verona (Italy). Dipartimento di Biotecnologie; Bressan, Mauro [Univ. di Verona, Verona (Italy). Dipartimento di Biotecnologie; Paleček, David [Lund Univ. (Sweden). Dept. of Chemical Physics; Židek, Karel [Lund Univ. (Sweden). Dept. of Chemical Physics; Niyogi, Krishna K. [Univ. of California, Berkeley, CA (United States). Howard Hughes Medical Inst., Dept. of Plant and Microbial Biology; Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Molecular Biophysics and Integrated Bioimaging Division; Fleming, Graham R. [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Molecular Biophysics and Integrated Bioimaging Division; Univ. of California, Berkeley, CA (United States). Dept. of Chemistry, Graduate Group in Applied Science and Technology; Zigmantas, Donatas [Lund Univ. (Sweden). Dept. of Chemical Physics; Bassi, Roberto [Univ. di Verona, Verona (Italy). Dipartimento di Biotecnologie; Consiglio Nazionale delle Ricerche (CNR), Firenze (Italy). Istituto per la Protezione delle Piante (IPP)

    2017-04-10

    Oxygenic photoautotrophs require mechanisms for rapidly matching the level of chlorophyll excited states from light harvesting with the rate of electron transport from water to carbon dioxide. These photoprotective reactions prevent formation of reactive excited states and photoinhibition. The fastest response to excess illumination is the so-called non-photochemical quenching which, in higher plants, requires the luminal pH sensor PsbS and other yet unidentified components of the photosystem II antenna. Both trimeric light-harvesting complex II (LHCII) and monomeric LHC proteins have been indicated as site(s) of the heat-dissipative reactions. Different mechanisms have been proposed: Energy transfer to a lutein quencher in trimers, formation of a zeaxanthin radical cation in monomers. Here, we report on the construction of a mutant lacking all monomeric LHC proteins but retaining LHCII trimers. Its non-photochemical quenching induction rate was substantially slower with respect to the wild type. A carotenoid radical cation signal was detected in the wild type, although it was lost in the mutant. Here, we conclude that non-photochemical quenching is catalysed by two independent mechanisms, with the fastest activated response catalysed within monomeric LHC proteins depending on both zeaxanthin and lutein and on the formation of a radical cation. Trimeric LHCII was responsible for the slowly activated quenching component whereas inclusion in supercomplexes was not required. Finally, this latter activity does not depend on lutein nor on charge transfer events, whereas zeaxanthin was essential.

  12. Selected HIV-1 Env Trimeric Formulations Act as Potent Immunogens in a Rabbit Vaccination Model

    NARCIS (Netherlands)

    Heyndrickx, Leo; Stewart-Jones, Guillaume; Jansson, Marianne; Schuitemaker, Hanneke; Bowles, Emma; Buonaguro, Luigi; Grevstad, Berit; Vinner, Lasse; Vereecken, Katleen; Parker, Joe; Ramaswamy, Meghna; Biswas, Priscilla; Vanham, Guido; Scarlatti, Gabriella; Fomsgaard, Anders

    2013-01-01

    Ten to 30% of HIV-1 infected subjects develop broadly neutralizing antibodies (bNAbs) during chronic infection. We hypothesized that immunizing rabbits with viral envelope glycoproteins (Envs) from these patients may induce bNAbs, when formulated as a trimeric protein and in the presence of an

  13. CD4-induced activation in a soluble HIV-1 Env trimer

    NARCIS (Netherlands)

    Guttman, Miklos; Garcia, Natalie K.; Cupo, Albert; Matsui, Tsutomu; Julien, Jean-Philippe; Sanders, Rogier W.; Wilson, Ian A.; Moore, John P.; Lee, Kelly K.

    2014-01-01

    The HIV envelope glycoprotein (Env) trimer undergoes receptor-induced conformational changes that drive fusion of the viral and cellular membranes. Env conformational changes have been observed using low-resolution electron microscopy, but only large-scale rearrangements have been visible. Here, we

  14. Cleavage-independent HIV-1 Env trimers engineered as soluble native spike mimetics for vaccine design.

    Science.gov (United States)

    Sharma, Shailendra Kumar; de Val, Natalia; Bale, Shridhar; Guenaga, Javier; Tran, Karen; Feng, Yu; Dubrovskaya, Viktoriya; Ward, Andrew B; Wyatt, Richard T

    2015-04-28

    Viral glycoproteins mediate entry by pH-activated or receptor-engaged activation and exist in metastable pre-fusogenic states that may be stabilized by directed rational design. As recently reported, the conformationally fixed HIV-1 envelope glycoprotein (Env) trimers in the pre-fusion state (SOSIP) display molecular homogeneity and structural integrity at relatively high levels of resolution. However, the SOSIPs necessitate full Env precursor cleavage, which requires endogenous furin overexpression. Here, we developed an alternative strategy using flexible peptide covalent linkage of Env subdomains to produce soluble, homogeneous, and cleavage-independent Env mimics, called native flexibly linked (NFL) trimers, as vaccine candidates. This simplified design avoids the need for furin co-expression and, in one case, antibody affinity purification to accelerate trimer scale-up for preclinical and clinical applications. We have successfully translated the NFL design to multiple HIV-1 subtypes, establishing the potential to become a general method of producing native-like, well-ordered Env trimers for HIV-1 or other viruses. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

  15. Woodfordin D and oenothein A, trimeric hydrolyzable tannins of macro-ring structure with antitumor activity.

    Science.gov (United States)

    Yoshida, T; Chou, T; Matsuda, M; Yasuhara, T; Yazaki, K; Hatano, T; Nitta, A; Okuda, T

    1991-05-01

    Two new antitumor trimeric hydrolyzable tannins, woodfordin D (5) and oenothein A (13), were isolated from the dried flowers of Woodfordia fruticosa, and their macrocyclic structures, which have a novel constituent unit (woodfordinoyl group) connecting the monomers, have been elucidated on the basis of spectral and chemical evidence. Oenothein A (13) was also isolated from the leaves of Oenothera biennis.

  16. A mixed Ni(II) ionic complex containing V-shaped water trimer ...

    Indian Academy of Sciences (India)

    A mixed Ni(II) ionic complex containing V-shaped water trimer: Synthesis, spectral, structural and thermal properties of. {[Ni(2,2. ′. -bpy)3][Ni(2-cpida)(2,2. ′. -bpy)]} (ClO4).3H2O. NALLASAMY PALANISAMIa, KABALI SENTHILKUMARb, MOHAN GOPALAKRISHNANb and IL-SHIK MOONa,∗. aDepartment of Chemical ...

  17. Reduced influenza viral neutralizing activity of natural human trimers of surfactant protein D

    DEFF Research Database (Denmark)

    Hartshorn, Kevan L; White, Mitchell R; Tecle, Tesfaldet

    2007-01-01

    human SP-D multimers as well as reduced hemagglutination inhibiting activity against several strains of IAV. Natural SP-D trimers also had different interactions with human neutrophil peptide defensins (HNPs) in viral neutralization assays as compared to multimeric SP-D. CONCLUSION: These studies...

  18. Selective ethylene trimerization : A study into the mechanism and the reduction of PE formation

    NARCIS (Netherlands)

    Hagen, H.; Kretschmer, WP; van Buren, FR; Hessen, B; Oeffelen, Dominicus A. van

    2006-01-01

    In the present study, the titanium-catalyzed ethylene trimerization in general, and more specifically, the concomitant PE formation have been studied. The polymer formation is undesirable as it not only will lead to lower I-hexene yields, but it can cause reactor fouling under the applied conditions

  19. Selected HIV-1 Env trimeric formulations act as potent immunogens in a rabbit vaccination model.

    Science.gov (United States)

    Heyndrickx, Leo; Stewart-Jones, Guillaume; Jansson, Marianne; Schuitemaker, Hanneke; Bowles, Emma; Buonaguro, Luigi; Grevstad, Berit; Vinner, Lasse; Vereecken, Katleen; Parker, Joe; Ramaswamy, Meghna; Biswas, Priscilla; Vanham, Guido; Scarlatti, Gabriella; Fomsgaard, Anders

    2013-01-01

    Ten to 30% of HIV-1 infected subjects develop broadly neutralizing antibodies (bNAbs) during chronic infection. We hypothesized that immunizing rabbits with viral envelope glycoproteins (Envs) from these patients may induce bNAbs, when formulated as a trimeric protein and in the presence of an adjuvant. Based on in vitro neutralizing activity in serum, patients with bNAbs were selected for cloning of their HIV-1 Env. Seven stable soluble trimeric gp140 proteins were generated from sequences derived from four adults and two children infected with either clade A or B HIV-1. From one of the clade A Envs both the monomeric and trimeric Env were produced for comparison. Rabbits were immunized with soluble gp120 or trimeric gp140 proteins in combination with the adjuvant dimethyl dioctadecyl ammonium/trehalose dibehenate (CAF01). Env binding in rabbit immune serum was determined using ELISAs based on gp120-IIIB protein. Neutralizing activity of IgG purified from rabbit immune sera was measured with the pseudovirus-TZMbl assay and a PBMC-based neutralization assay for selected experiments. It was initially established that gp140 trimers induce better antibody responses over gp120 monomers and that the adjuvant CAF01 was necessary for such strong responses. Gp140 trimers, based on HIV-1 variants from patients with bNAbs, were able to elicit both gp120IIIB specific IgG and NAbs to Tier 1 viruses of different subtypes. Potency of NAbs closely correlated with titers, and an gp120-binding IgG titer above a threshold of 100,000 was predictive of neutralization capability. Finally, peptide inhibition experiments showed that a large fraction of the neutralizing IgG was directed against the gp120 V3 region. Our results indicate that the strategy of reverse immunology based on selected Env sequences is promising when immunogens are delivered as stabilized trimers in CAF01 adjuvant and that the rabbit is a valuable model for HIV vaccine studies.

  20. Selected HIV-1 Env trimeric formulations act as potent immunogens in a rabbit vaccination model.

    Directory of Open Access Journals (Sweden)

    Leo Heyndrickx

    Full Text Available BACKGROUND: Ten to 30% of HIV-1 infected subjects develop broadly neutralizing antibodies (bNAbs during chronic infection. We hypothesized that immunizing rabbits with viral envelope glycoproteins (Envs from these patients may induce bNAbs, when formulated as a trimeric protein and in the presence of an adjuvant. METHODS: Based on in vitro neutralizing activity in serum, patients with bNAbs were selected for cloning of their HIV-1 Env. Seven stable soluble trimeric gp140 proteins were generated from sequences derived from four adults and two children infected with either clade A or B HIV-1. From one of the clade A Envs both the monomeric and trimeric Env were produced for comparison. Rabbits were immunized with soluble gp120 or trimeric gp140 proteins in combination with the adjuvant dimethyl dioctadecyl ammonium/trehalose dibehenate (CAF01. Env binding in rabbit immune serum was determined using ELISAs based on gp120-IIIB protein. Neutralizing activity of IgG purified from rabbit immune sera was measured with the pseudovirus-TZMbl assay and a PBMC-based neutralization assay for selected experiments. RESULTS: It was initially established that gp140 trimers induce better antibody responses over gp120 monomers and that the adjuvant CAF01 was necessary for such strong responses. Gp140 trimers, based on HIV-1 variants from patients with bNAbs, were able to elicit both gp120IIIB specific IgG and NAbs to Tier 1 viruses of different subtypes. Potency of NAbs closely correlated with titers, and an gp120-binding IgG titer above a threshold of 100,000 was predictive of neutralization capability. Finally, peptide inhibition experiments showed that a large fraction of the neutralizing IgG was directed against the gp120 V3 region. CONCLUSIONS: Our results indicate that the strategy of reverse immunology based on selected Env sequences is promising when immunogens are delivered as stabilized trimers in CAF01 adjuvant and that the rabbit is a valuable model

  1. Targeting protein-protein interactions with trimeric ligands: high affinity inhibitors of the MAGUK protein family.

    Science.gov (United States)

    Nissen, Klaus B; Haugaard-Kedström, Linda M; Wilbek, Theis S; Nielsen, Line S; Åberg, Emma; Kristensen, Anders S; Bach, Anders; Jemth, Per; Strømgaard, Kristian

    2015-01-01

    PDZ domains in general, and those of PSD-95 in particular, are emerging as promising drug targets for diseases such as ischemic stroke. We have previously shown that dimeric ligands that simultaneously target PDZ1 and PDZ2 of PSD-95 are highly potent inhibitors of PSD-95. However, PSD-95 and the related MAGUK proteins contain three consecutive PDZ domains, hence we envisioned that targeting all three PDZ domains simultaneously would lead to more potent and potentially more specific interactions with the MAGUK proteins. Here we describe the design, synthesis and characterization of a series of trimeric ligands targeting all three PDZ domains of PSD-95 and the related MAGUK proteins, PSD-93, SAP-97 and SAP-102. Using our dimeric ligands targeting the PDZ1-2 tandem as starting point, we designed novel trimeric ligands by introducing a PDZ3-binding peptide moiety via a cysteine-derivatized NPEG linker. The trimeric ligands generally displayed increased affinities compared to the dimeric ligands in fluorescence polarization binding experiments and optimized trimeric ligands showed low nanomolar inhibition towards the four MAGUK proteins, thus being the most potent inhibitors described. Kinetic experiments using stopped-flow spectrometry showed that the increase in affinity is caused by a decrease in the dissociation rate of the trimeric ligand as compared to the dimeric ligands, likely reflecting the lower probability of simultaneous dissociation of all three PDZ ligands. Thus, we have provided novel inhibitors of the MAGUK proteins with exceptionally high affinity, which can be used to further elucidate the therapeutic potential of these proteins.

  2. Receptor Activation of HIV-1 Env Leads to Asymmetric Exposure of the gp41 Trimer.

    Science.gov (United States)

    Khasnis, Mukta D; Halkidis, Konstantine; Bhardwaj, Anshul; Root, Michael J

    2016-12-01

    Structural rearrangements of HIV-1 glycoprotein Env promote viral entry through membrane fusion. Env is a symmetric homotrimer with each protomer composed of surface subunit gp120 and transmembrane subunit gp41. Cellular CD4- and chemokine receptor-binding to gp120 coordinate conformational changes in gp41, first to an extended prehairpin intermediate (PHI) and, ultimately, into a fusogenic trimer-of-hairpins (TOH). HIV-1 fusion inhibitors target gp41 in the PHI and block TOH formation. To characterize structural transformations into and through the PHI, we employed asymmetric Env trimers containing both high and low affinity binding sites for individual fusion inhibitors. Asymmetry was achieved using engineered Env heterotrimers composed of protomers deficient in either CD4- or chemokine receptor-binding. Linking receptor engagement to inhibitor affinity allowed us to assess conformational changes of individual Env protomers in the context of a functioning trimer. We found that the transition into the PHI could occur symmetrically or asymmetrically depending on the stoichiometry of CD4 binding. Sequential engagement of multiple CD4s promoted progressive exposure of individual fusion inhibitor binding sites in a CD4-dependent fashion. By contrast, engagement of only a single CD4 molecule led to a delayed, but symmetric, exposure of the gp41 trimer. This complex coupling between Env-CD4 interaction and gp41 exposure explained the multiphasic fusion-inhibitor titration observed for a mutant Env homotrimer with a naturally asymmetric gp41. Our results suggest that the spatial and temporal exposure of gp41 can proceed in a nonconcerted, asymmetric manner depending on the number of CD4s that engage the Env trimer. The findings have important implications for the mechanism of viral membrane fusion and the development of vaccine candidates designed to elicit neutralizing antibodies targeting gp41 in the PHI.

  3. Temperature dependent dynamics of DegP-trimer: A molecular dynamics study

    Directory of Open Access Journals (Sweden)

    Nivedita Rai

    2015-01-01

    Full Text Available DegP is a heat shock protein from high temperature requirement protease A family, which reacts to the environmental stress conditions in an ATP independent way. The objective of the present analysis emerged from the temperature dependent functional diversity of DegP between chaperonic and protease activities at temperatures below and above 28 °C, respectively. DegP is a multimeric protein and the minimal functional unit, DegP-trimer, is of great importance in understanding the DegP pathway. The structural aspects of DegP-trimer with respect to temperature variation have been studied using molecular dynamics simulations (for 100 ns and principal component analysis to highlight the temperature dependent dynamics facilitating its functional diversity. The DegP-trimer revealed a pronounced dynamics at both 280 and 320 K, when compared to the dynamics observed at 300 K. The LA loop is identified as the highly flexible region during dynamics and at extreme temperatures, the residues 46–80 of LA loop express a flip towards right (at 280 and left ( at 320 K with respect to the fixed β-sheet connecting the LA loop of protease for which Phe46 acts as one of the key residues. Such dynamics of LA loop facilitates inter-monomeric interaction with the PDZ1 domain of the neighbouring monomer and explains its active participation when DegP exists as trimer. Hence, the LA loop mediated dynamics of DegP-trimer is expected to provide further insight into the temperature dependent dynamics of DegP towards the understanding of its assembly and functional diversity in the presence of substrate.

  4. Homogeneous photochemical water oxidation by biuret-modified Fe-TAML: evidence of Fe(V)(O) intermediate.

    Science.gov (United States)

    Panda, Chakadola; Debgupta, Joyashish; Díaz Díaz, David; Singh, Kundan K; Sen Gupta, Sayam; Dhar, Basab B

    2014-09-03

    Water splitting, leading to hydrogen and oxygen in a process that mimics natural photosynthesis, is extremely important for devising a sustainable solar energy conversion system. Development of earth-abundant, transition metal-based catalysts that mimic the oxygen-evolving complex of photosystem II, which is involved in oxidation of water to O2 during natural photosynthesis, represents a major challenge. Further, understanding the exact mechanism, including elucidation of the role of active metal-oxo intermediates during water oxidation (WO), is critical to the development of more efficient catalysts. Herein, we report Fe(III) complexes of biuret-modified tetra-amidomacrocyclic ligands (Fe-TAML; 1a and 1b) that catalyze fast, homogeneous, photochemical WO to give O2, with moderate efficiency (maximum TON = 220, TOF = 0.76 s(-1)). Previous studies on photochemical WO using iron complexes resulted in demetalation of the iron complexes with concomitant formation of iron oxide nanoparticles (NPs) that were responsible for WO. Herein, we show for the first time that a high valent Fe(V)(O) intermediate species is photochemically generated as the active intermediate for the oxidation of water to O2. To the best of our knowledge, this represents the first example of a molecular iron complex catalyzing photochemical WO through a Fe(V)(O) intermediate.

  5. Ion Exchange Study of Some New Copolymer Resins Derived from 8-Hydroxyquinoline-5-sulphonic Acid, Biuret and Formaldehyde

    Directory of Open Access Journals (Sweden)

    P. A. Dhakite

    2011-01-01

    Full Text Available Copolymer resins (8-HQSABF were synthesized by the condensation of 8-hydroxyquinoline-5-sulphonic acid and biuret with formaldehyde in the presence of hydrochloric acid as catalyst, proved to be selective chelation ion exchange copolymer resins for certain metals. Chelation ion exchange properties to these polymers were studied for Cu2+, Cd2+, Co2+ and Zn2+ ions. A batch equilibrium method was employed in the study of the selectivity of the distribution of a given metal ions between the polymer sample and a solution containing the metal ion. The study was carried out over a wide pH range and in a media of various ions strengths. The polymer showed a higher selectivity for Cu2+ ions than for Cd2+, Co2+ and Zn2+ ions. Hence on the basis of above studies these copolymer may be used as semiconductors, surface coating, ion-exchangers, materials for rechargeable battery cell in various electronic industries, plastic materials, elastomers and in boiler plants

  6. Ethanol water azeotrope Study. Molecular characterization of ethanol dimers, hetero dimers and hetero trimers of ethanol water

    International Nuclear Information System (INIS)

    Mejia, Sol M; Espinal, Juan F; Mondragon, Fanor

    2006-01-01

    In this investigation, as a first stage, we studied the structure and stability of the dimers,hetero dimers and hetero trimers like aggregates of ethanol and ethanol water respectively. Molecular modelling using hybrid B3LYP of the density functional theory (DFT) was used in this research. O-H bond lengths, hydrogen bond distances, CCOH dihedral angles, enthalpies and Gibbs free energies of dimerization and trimerization and vibrational frequencies of stretching O-H for the dimers and hetero dimers in function of the redshift undergone by proton donor molecules were analyzed. The results shows that the monomers arrange into aggregates which undergo geometric changes induced by the hydrogen bonds. For hetero dimers and hetero trimers the enhancement of the hydrogen bonds where the proton donor molecule corresponds to water was observed. In general, the dimers are more stable than the hetero dimers. We propose the formation of C-H-O hydrogen bonds in some hetero trimers

  7. Immunogenicity of a Prefusion HIV-1 Envelope Trimer in Complex with a Quaternary-Structure-Specific Antibody.

    Science.gov (United States)

    Cheng, Cheng; Pancera, Marie; Bossert, Adam; Schmidt, Stephen D; Chen, Rita E; Chen, Xuejun; Druz, Aliaksandr; Narpala, Sandeep; Doria-Rose, Nicole A; McDermott, Adrian B; Kwong, Peter D; Mascola, John R

    2015-12-30

    The HIV-1 envelope trimer (Env) is the target of broadly neutralizing antibodies and is being explored as a vaccine candidate to elicit protective antibodies. One of the most promising antigenic and structural mimics of HIV-1 Env is the SOSIP.664-stabilized soluble trimer from the clade A strain BG505, which is preferentially recognized by broadly neutralizing antibodies. Trimer immunization elicits high-titer neutralization of the autologous tier 2 BG505 strain; however, breadth is limited, and substantial interest has focused on understanding and improving trimer immunogenicity. We sought to improve the antigenic specificity of BG505 SOSIP.664 by reducing recognition of the variable loop 3 (V3) region, which elicits only weakly neutralizing antibodies. To stabilize the trimer in its prefusion closed conformation, we complexed trimeric BG505 SOSIP.664 with the antigen-binding fragment (Fab) of PGT145, a broadly neutralizing quaternary-structure-specific antibody. Compared to the ligand-free trimer, the PGT145 Fab-BG505 SOSIP.664 complex displayed increased melting temperature stability and reduced V3 recognition. In guinea pigs, immunization with the PGT145 Fab-BG505 SOSIP.664 complex elicited ∼100-fold lower V3-directed binding and neutralization titers than those obtained with ligand-free BG505 SOSIP.664. Both complexed and ligand-free BG505 SOSIP.664 elicited comparable neutralization of the autologous BG505 virus, and in both cases, BG505 neutralization mapped to the outer domain of gp120 for some guinea pigs. Our results indicate that it is possible to reduce immune recognition of the V3 region of the trimer while maintaining the antigenic profile needed to induce autologous neutralizing antibodies. These data suggest that appropriate modifications of trimer immunogens could further focus the immune response on key neutralization epitopes. HIV-1 Env trimers have been proposed as preferred HIV-1 vaccine immunogens. One version, BG505 SOSIP.664, a soluble

  8. Supermolecular bent configuration composed of achiral flexible liquid crystal trimers exhibiting chiral domains with opposite handedness.

    Science.gov (United States)

    Sasaki, Haruna; Takanishi, Yoichi; Yamamoto, Jun; Yoshizawa, Atsushi

    2015-03-26

    Chirality's effects on physical properties of materials and how chirality arises have persisted as attractive issues in chemistry. We prepared a homologous series of achiral liquid crystal trimers in which three phenylpyrimidine units are connected via flexible heptamethylene spacers. An equimolecular mixture of a trimer with a nematic (N) phase and that with smectic A (SmA), smectic C (SmC), and smectic B phases was found to exhibit an N phase, a SmC phase, and a B4 phase composed of chiral domains with opposite handedness. The chiral characteristics of the B4 phase were confirmed by uncrossing the polarizers in opposite directions. XRD measurements reveal that both SmC and B4 phases have an interdigitated layer structure. That molecular interdigitation might form a supermolecular bent configuration that can produce saddle splay curvature to drive the B4 phase.

  9. Targeting Protein-Protein Interactions with Trimeric Ligands: High Affinity Inhibitors of the MAGUK Protein Family

    DEFF Research Database (Denmark)

    Nissen, Klaus B; Kedström, Linda Maria Haugaard; Wilbek, Theis S

    2015-01-01

    PDZ domains in general, and those of PSD-95 in particular, are emerging as promising drug targets for diseases such as ischemic stroke. We have previously shown that dimeric ligands that simultaneously target PDZ1 and PDZ2 of PSD-95 are highly potent inhibitors of PSD-95. However, PSD-95 and the ......PDZ domains in general, and those of PSD-95 in particular, are emerging as promising drug targets for diseases such as ischemic stroke. We have previously shown that dimeric ligands that simultaneously target PDZ1 and PDZ2 of PSD-95 are highly potent inhibitors of PSD-95. However, PSD-95...... linker. The trimeric ligands generally displayed increased affinities compared to the dimeric ligands in fluorescence polarization binding experiments and optimized trimeric ligands showed low nanomolar inhibition towards the four MAGUK proteins, thus being the most potent inhibitors described. Kinetic...

  10. Activation and Inactivation of Primary Human Immunodeficiency Virus Envelope Glycoprotein Trimers by CD4-Mimetic Compounds

    Science.gov (United States)

    Madani, Navid; Princiotto, Amy M.; Zhao, Connie; Jahanbakhshsefidi, Fatemeh; Mertens, Max; Herschhorn, Alon; Melillo, Bruno; Smith, Amos B.

    2016-01-01

    ABSTRACT Human immunodeficiency virus type 1 (HIV-1) entry into cells is mediated by the viral envelope glycoproteins (Env), a trimer of three gp120 exterior glycoproteins, and three gp41 transmembrane glycoproteins. The metastable Env is triggered to undergo entry-related conformational changes when gp120 binds sequentially to the receptors, CD4 and CCR5, on the target cell. Small-molecule CD4-mimetic compounds (CD4mc) bind gp120 and act as competitive inhibitors of gp120-CD4 engagement. Some CD4mc have been shown to trigger Env prematurely, initially activating Env function, followed by rapid and irreversible inactivation. Here, we study CD4mc with a wide range of anti-HIV-1 potencies and demonstrate that all tested CD4mc are capable of activating as well as inactivating Env function. Biphasic dose-response curves indicated that the occupancy of the protomers in the Env trimer governs viral activation versus inactivation. One CD4mc bound per Env trimer activated HIV-1 infection. Envs with two CD4mc bound were activated for infection of CD4-negative, CCR5-positive cells, but the infection of CD4-positive, CCR5-positive cells was inhibited. Virus was inactivated when all three Env protomers were occupied by the CD4mc, and gp120 shedding from the Env trimer was increased in the presence of some CD4mc. Env reactivity and the on rates of CD4mc binding to the Env trimer were found to be important determinants of the potency of activation and entry inhibition. Cross-sensitization of Env protomers that do not bind the CD4mc to neutralization by an anti-V3 antibody was not evident. These insights into the mechanism of antiviral activity of CD4mc should assist efforts to optimize their potency and utility. IMPORTANCE The trimeric envelope glycoproteins of human immunodeficiency virus type 1 (HIV-1) mediate virus entry into host cells. Binding to the host cell receptors, CD4 and CCR5, triggers changes in the conformation of the HIV-1 envelope glycoprotein trimer important

  11. Trimerization catalysis of phenylisocyanate in the presence of phenolic mannich bases

    Energy Technology Data Exchange (ETDEWEB)

    Selivanov, A.V.; Zenitova, L.A.; Bakirova, I.N.; Kirpichnikov, P.A.

    1988-11-01

    The kinetics of the cyclic trimerization of phenylisocyanate in the presence of phenolic Mannich bases have been studied by IR spectroscopy; a catalysis mechanism for the reaction is proposed. It was found that in order for trimerization to occur the catalyst molecule must contain both a hydroxyl group and a tertiary nitrogen atom, which leads to reaction at the OH group of the catalyst and its conversion via a urethane derivative to an allophanate; the latter species undergoes tautomeric rearrangement to a bipolar ion, which is the actual catalysis site. The effects of the number of aminomethyl, hydroxyl, and other electron-donating and electron-withdrawing substituents on the structure of Mannich bases have also been investigated.

  12. Analysis of an unusual hetero-halogen bonded trimer using charge ...

    Indian Academy of Sciences (India)

    Special Issue on CHEMICAL BONDING. DOI 10.1007/s12039-016-1149-2. Analysis of an unusual hetero-halogen bonded trimer using charge density analysis: A case of concerted type I Br···Br and type II. Br···Cl interactions. MYSORE S PAVAN and TAYUR N GURU ROW. ∗. Solid State and Structural Chemistry Unit, ...

  13. Surface Induced Dissociations and Reactions of Acetonitrile Monomer, Dimer and Trimer Ions

    Czech Academy of Sciences Publication Activity Database

    Mair, C.; Herman, Zdeněk; Fedor, J.; Lezius, M.; Märk, T. D.

    2003-01-01

    Roč. 118, č. 3 (2003), s. 1479-1486 ISSN 0021-9606 R&D Projects: GA ČR GA203/00/0632 Grant - others:Cooperation PST(AT) 2000-06 Institutional research plan: CEZ:AV0Z4040901 Keywords : surface-induced * acetonitrile monomer * trimer ions Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 2.950, year: 2003

  14. Homogeneous and heterogeneous cyclopentadienyl-arene titanium catalysts for selective ethylene trimerization to 1-hexene

    Czech Academy of Sciences Publication Activity Database

    Varga, Vojtěch; Hodík, Tomáš; Lamač, Martin; Horáček, Michal; Zukal, Arnošt; Žilková, Naděžda; Parker Jr., W. O.; Pinkas, Jiří

    2015-01-01

    Roč. 777, FEB 2015 (2015), s. 57-66 ISSN 0022-328X R&D Projects: GA ČR GBP106/12/G015; GA ČR(CZ) GAP207/12/2368; GA ČR(CZ) GA14-08531S Institutional support: RVO:61388955 Keywords : Cyclopentadienyl-arene complexes * Ethylene trimerization * SBA-15 Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 2.336, year: 2015

  15. Recognition of HIV-inactivating peptide triazoles by the recombinant soluble Env trimer, BG505 SOSIP.664.

    Science.gov (United States)

    Acharya, Kriti; Rashad, Adel A; Moraca, Francesca; Klasse, Per Johan; Moore, John P; Abrams, Cameron; Chaiken, Irwin

    2017-05-01

    Peptide triazole (PT) antagonists interact with gp120 subunits of HIV-1 Env trimers to block host cell receptor interactions, trigger gp120 shedding, irreversibly inactivate virus and inhibit infection. Despite these enticing functions, understanding the structural mechanism of PT-Env trimer encounter has been limited. In this work, we combined competition interaction analysis and computational simulation to demonstrate PT binding to the recombinant soluble trimer, BG505 SOSIP.664, a stable variant that resembles native virus spikes in binding to CD4 receptor as well as known conformationally-dependent Env antibodies. Binding specificity and computational modeling fit with encounter through complementary PT pharmacophore Ile-triazolePro-Trp interaction with a 2-subsite cavity in the Env gp120 subunit of SOSIP trimer similar to that in monomeric gp120. These findings argue that PTs are able to recognize and bind a closed prefusion state of Env trimer upon HIV-1 encounter. The results provide a structural model of how PTs exert their function on virion trimeric spike protein and a platform to inform future antagonist design. Proteins 2017; 85:843-851. © 2016 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  16. Effect of trimerization motifs on quaternary structure, antigenicity, and immunogenicity of a noncleavable HIV-1 gp140 envelope glycoprotein

    International Nuclear Information System (INIS)

    Du, Sean X.; Idiart, Rebecca J.; Mariano, Ellaine B.; Chen, Helen; Jiang Peifeng; Xu Li; Ostrow, Kristin M.; Wrin, Terri; Phung, Pham; Binley, James M.; Petropoulos, Christos J.; Ballantyne, John A.; Whalen, Robert G.

    2009-01-01

    The external domains of the HIV-1 envelope glycoprotein (gp120 and the gp41 ectodomain, collectively known as gp140) contain all known viral neutralization epitopes. Various strategies have been used to create soluble trimers of the envelope to mimic the structure of the native viral protein, including mutation of the gp120-gp41 cleavage site, introduction of disulfide bonds, and fusion to heterologous trimerization motifs. We compared the effects on quaternary structure, antigenicity, and immunogenicity of three such motifs: T4 fibritin, a GCN4 variant, and the Escherichia coli aspartate transcarbamoylase catalytic subunit. Fusion of each motif to the C-terminus of a noncleavable JRCSF gp140(-) envelope protein led to enhanced trimerization but had limited effects on the antigenic profile and CD4-binding ability of the trimers. Immunization of rabbits provided no evidence that the trimerized gp140(-) constructs induced significantly improved neutralizing antibodies to several HIV-1 pseudoviruses, compared to gp140 lacking a trimerization motif. However, modest differences in both binding specificity and neutralizing antibody responses were observed among the various immunogens.

  17. [Sensitive method for continuous detection of peptides and proteins using the biuret reaction and a copper-Sephadex reactor (author's transl)].

    Science.gov (United States)

    Later, R; Quincy, C

    1979-06-01

    We describe a detection method relying both on the copper displacement from a Sephadex gel by peptides and proteins, and on the subsequent colorimetric determination of the complexed copper. The system described is fully automated and it permits a continuous analysis of column effluents. The choice of cuprizone as a detecting reagent for copper, enables one to bring the detection limit down to 200 ng for albumin and 60 ng for alanylglycylglycin. The specificity of the method is the same as the biuret reaction. Some examples of the possible applications are given.

  18. Gram-negative trimeric porins have specific LPS binding sites that are essential for porin biogenesis

    Science.gov (United States)

    Arunmanee, Wanatchaporn; Pathania, Monisha; Solovyova, Alexandra S.; Le Brun, Anton P.; Ridley, Helen; Baslé, Arnaud; van den Berg, Bert; Lakey, Jeremy H.

    2016-01-01

    The outer membrane (OM) of gram-negative bacteria is an unusual asymmetric bilayer with an external monolayer of lipopolysaccharide (LPS) and an inner layer of phospholipids. The LPS layer is rigid and stabilized by divalent cation cross-links between phosphate groups on the core oligosaccharide regions. This means that the OM is robust and highly impermeable to toxins and antibiotics. During their biogenesis, OM proteins (OMPs), which function as transporters and receptors, must integrate into this ordered monolayer while preserving its impermeability. Here we reveal the specific interactions between the trimeric porins of Enterobacteriaceae and LPS. Isolated porins form complexes with variable numbers of LPS molecules, which are stabilized by calcium ions. In earlier studies, two high-affinity sites were predicted to contain groups of positively charged side chains. Mutation of these residues led to the loss of LPS binding and, in one site, also prevented trimerization of the porin, explaining the previously observed effect of LPS mutants on porin folding. The high-resolution X-ray crystal structure of a trimeric porin–LPS complex not only helps to explain the mutagenesis results but also reveals more complex, subtle porin–LPS interactions and a bridging calcium ion. PMID:27493217

  19. Trimeric HIV-1-Env Structures Define Glycan Shields from Clades A, B, and G.

    Science.gov (United States)

    Stewart-Jones, Guillaume B E; Soto, Cinque; Lemmin, Thomas; Chuang, Gwo-Yu; Druz, Aliaksandr; Kong, Rui; Thomas, Paul V; Wagh, Kshitij; Zhou, Tongqing; Behrens, Anna-Janina; Bylund, Tatsiana; Choi, Chang W; Davison, Jack R; Georgiev, Ivelin S; Joyce, M Gordon; Kwon, Young Do; Pancera, Marie; Taft, Justin; Yang, Yongping; Zhang, Baoshan; Shivatare, Sachin S; Shivatare, Vidya S; Lee, Chang-Chun D; Wu, Chung-Yi; Bewley, Carole A; Burton, Dennis R; Koff, Wayne C; Connors, Mark; Crispin, Max; Baxa, Ulrich; Korber, Bette T; Wong, Chi-Huey; Mascola, John R; Kwong, Peter D

    2016-05-05

    The HIV-1-envelope (Env) trimer is covered by a glycan shield of ∼90 N-linked oligosaccharides, which comprises roughly half its mass and is a key component of HIV evasion from humoral immunity. To understand how antibodies can overcome the barriers imposed by the glycan shield, we crystallized fully glycosylated Env trimers from clades A, B, and G, visualizing the shield at 3.4-3.7 Å resolution. These structures reveal the HIV-1-glycan shield to comprise a network of interlocking oligosaccharides, substantially ordered by glycan crowding, that encase the protein component of Env and enable HIV-1 to avoid most antibody-mediated neutralization. The revealed features delineate a taxonomy of N-linked glycan-glycan interactions. Crowded and dispersed glycans are differently ordered, conserved, processed, and recognized by antibody. The structures, along with glycan-array binding and molecular dynamics, reveal a diversity in oligosaccharide affinity and a requirement for accommodating glycans among known broadly neutralizing antibodies that target the glycan-shielded trimer. Copyright © 2016 Elsevier Inc. All rights reserved.

  20. Composition and Antigenic Effects of Individual Glycan Sites of a Trimeric HIV-1 Envelope Glycoprotein

    Directory of Open Access Journals (Sweden)

    Anna-Janina Behrens

    2016-03-01

    Full Text Available The HIV-1 envelope glycoprotein trimer is covered by an array of N-linked glycans that shield it from immune surveillance. The high density of glycans on the trimer surface imposes steric constraints limiting the actions of glycan-processing enzymes, so that multiple under-processed structures remain on specific areas. These oligomannose glycans are recognized by broadly neutralizing antibodies (bNAbs that are not thwarted by the glycan shield but, paradoxically, target it. Our site-specific glycosylation analysis of a soluble, recombinant trimer (BG505 SOSIP.664 maps the extremes of simplicity and diversity of glycan processing at individual sites and reveals a mosaic of dense clusters of oligomannose glycans on the outer domain. Although individual sites usually minimally affect the global integrity of the glycan shield, we identify examples of how deleting some glycans can subtly influence neutralization by bNAbs that bind at distant sites. The network of bNAb-targeted glycans should be preserved on vaccine antigens.

  1. Ammonium nitrate, urea, and biuret fertilizers increase volume growth of 57-year-old douglas-fir trees within a gradient of nitrogen deficiency. Forest Service research paper

    Energy Technology Data Exchange (ETDEWEB)

    Miller, R.E.; Reukema, D.L.; Hazard, J.W.

    1996-03-01

    In a nitrogen-deficient plantation in southwest Washington, the authors (1) compared effects of 224 kg N/ha as ammonium nitrate, urea, and biuret on volume growth of dominant and codominant Douglas-fir (Pseudotsuga menziesii var. menziesii (Mirb.) Franco); (2) determined how 8-year response of these trees to fertilization was related to their distance from a strip of the plantation interplanted with nitrogen-fixing red alder (alnus rubra Bong.); and (3) observed effects of biuret on understory vegetation. On both sides of the strip centerline, the authors grouped subject trees into 30 plots of 4 trees each, based on slope position and distance from alder. The authors randomly assigned three fertilizers and a control within each plot. They analyzed separately data from east and west of the mixed stand certerline. Initial volume differed greatly among the 120 trees on each side, so they used covariance analysis to adjust observed treatment means. Adjusted mean volume growth was increased (p equal to or less than 0.10) by 22 to 28 percent on the east side and by 11 to 14 percent on the west side, with no significant difference in response to the three fertilizers.

  2. Validation for the determination of biuret in water-soluble, urea-based commercial inorganic fertilizer materials, urea solutions, and sulfur-coated urea products by reversed-phase liquid chromatography: single-laboratory validation of an extension of AOAC official methods 2003.14.

    Science.gov (United States)

    Hojjatie, Michael M; Abrams, Dean

    2014-01-01

    A single-laboratory validation (SLV) study for the LC determination of biuret in dry and liquid urea-based commercial fertilizers was conducted. A total of 23 samples were used: 11 commercial dry urea products, two urea ammonium nitrate products, eight liquid urea-based commercial fertilizers, and four sulfur-coated urea samples from different sources. In addition, one biuret standard from Aldrich and one sample from the Magruder check sample program were used as validation samples. The proposed method is an extension of AOAC Official Method 2003.14 and is based on dissolution of the test portion in the LC mobile phase and determination by HPLC. The system is linear over a concentration range of 1.00-4.50 mg/L biuret, with a correlation coefficient > or = 0.9999. The biuret was well- separated from urea in the commercial urea samples, and from other constituents in the commercial liquid fertilizer with no observed interferences. Recoveries were determined by spiking four of the validation materials with a known amount of biuret standard and measuring the biuret level according to the method. The averaged recovery was 97%. Method precision was determined by quadruplicate analyses of four of the liquid and six of the commercially available dry urea validation materials using three and four replicate analyses. For the liquid fertilizer analyses, the RSD ranged from 7.04 to 13.31%. For the dry urea analyses, the RSDs ranged from 5.68 to 14.34%. Instrument precision was evaluated at the test initiation by using seven injections of five biuret standard solutions. SD varied from 0.27 to 1.02%, with RSDs averaging 1.14%. The LOD was determined to be 0.009% biuret in material, while the LOQ was determined to be 0.031% biuret in material. In addition to the intralaboratory study, interlaboratory studies were performed by two other outside laboratories using this method. Over a concentration range of 0.2 to 0.9% biuret, the average SD was 0.11%, the average RSD was 21

  3. Development and Validation of a High-Performance Liquid Chromatography Method With UV Detection for Determination of 1-(2-phenylethyl)-5- (quinaldin-4-yl) Biuret in Rat Plasma.

    Science.gov (United States)

    Adibpour, Neda; Ahmadnasr, Maryam; Khodayar, Mohammad Javad; Rezaee, Saeed

    2013-01-01

    Recently, biuret derivatives have been reported as showing moderate to good cytotoxic effect against certain cancer cell lines. In this study, a high-performance liquid chromatography method was developed for determination of 1-(2-phenylethyl)-5-(quinaldin-4-yl) biuret (PEQB) in rat plasma to use in future studies on this compound and related derivatives. In this study, we describe a simple and sensitive high-performance liquid chromatography method with UV detection for determination of 1-(2-phenylethyl)-6-(quinaldin-4-yl) biuret (PEQB) in rat plasma. Separations were performed on a Nucleosil-100 CN HPLC column (125 × 4.0 mm) (5 µm), using a mixture of acetonitrile: methanol: potassium dihydrogen phosphate buffer (0.05 M, pH 3.5) (10:10:80) as mobile phase delivered at a flow rate of 1 mL/minute. Detection of PEQB and internal standard (1-([[3-(1,3-benzothiazol-2-ylsulfanyl)propyl]carbamoyl]amino)-N-phenylformamide) was performed at 235 nm and ambient temperature. Plasma samples (200 µL) were prepared by addition of 40 µL internal standard (100 µg/mL), and 400 µL acetonitrile. After vortex mixing and centrifugation at 10000 g, 50 µL of the clear supernatant was directly injected onto the chromatography column. Calibration curves were constructed by fitting the peak area ratio of the biuret to internal standard against concentration of biuret to a power model using generalized least squares nonlinear regression method. Under the above chromatography condition, biuret compound (PEQB) and the internal standard were detected at 4.5 and 13.5 minutes, respectively. Limit of quantitation of the PEQB was 0.1 µg/mL. Accuracy of the method over the concentration range of 0.1-100 µg/mL was between 88-109%. Inter- and intraday precisions were 4-19% and 6-8%, respectively. A good relationship in the form of a power model was found for two separate concentration ranges of 0.1-1 and 2.5-100 µg/mL (R (2)> 0.99). The presented simple HPLC method is sufficiently accurate

  4. Trimeric transmembrane domain interactions in paramyxovirus fusion proteins: roles in protein folding, stability, and function.

    Science.gov (United States)

    Smith, Everett Clinton; Smith, Stacy E; Carter, James R; Webb, Stacy R; Gibson, Kathleen M; Hellman, Lance M; Fried, Michael G; Dutch, Rebecca Ellis

    2013-12-13

    Paramyxovirus fusion (F) proteins promote membrane fusion between the viral envelope and host cell membranes, a critical early step in viral infection. Although mutational analyses have indicated that transmembrane (TM) domain residues can affect folding or function of viral fusion proteins, direct analysis of TM-TM interactions has proved challenging. To directly assess TM interactions, the oligomeric state of purified chimeric proteins containing the Staphylococcal nuclease (SN) protein linked to the TM segments from three paramyxovirus F proteins was analyzed by sedimentation equilibrium analysis in detergent and buffer conditions that allowed density matching. A monomer-trimer equilibrium best fit was found for all three SN-TM constructs tested, and similar fits were obtained with peptides corresponding to just the TM region of two different paramyxovirus F proteins. These findings demonstrate for the first time that class I viral fusion protein TM domains can self-associate as trimeric complexes in the absence of the rest of the protein. Glycine residues have been implicated in TM helix interactions, so the effect of mutations at Hendra F Gly-508 was assessed in the context of the whole F protein. Mutations G508I or G508L resulted in decreased cell surface expression of the fusogenic form, consistent with decreased stability of the prefusion form of the protein. Sedimentation equilibrium analysis of TM domains containing these mutations gave higher relative association constants, suggesting altered TM-TM interactions. Overall, these results suggest that trimeric TM interactions are important driving forces for protein folding, stability and membrane fusion promotion.

  5. Screening-level risk assessment for styrene-acrylonitrile (SAN) trimer detected in soil and groundwater.

    Science.gov (United States)

    Kirman, C R; Gargas, M L; Collins, J J; Rowlands, J C

    2012-01-01

    A screening-level risk assessment was conducted for styrene-acrylonitrile (SAN) Trimer detected at the Reich Farm Superfund site in Toms River, NJ. Consistent with a screening-level approach, on-site and off-site exposure scenarios were evaluated using assumptions that are expected to overestimate actual exposures and hazards at the site. Environmental sampling data collected for soil and groundwater were used to estimate exposure point concentrations. Several exposure scenarios were evaluated to assess potential on-site and off-site exposures, using parameter values for exposures to soil (oral, inhalation of particulates, and dermal contact) and groundwater (oral, dermal contact) to reflect central tendency exposure (CTE) and reasonable maximum exposure (RME) conditions. Three reference dose (RfD) values were derived for SAN Trimer for short-term, subchronic, and chronic exposures, based upon its effects on the liver in exposed rats. Benchmark (BMD) methods were used to assess the relationship between exposure and response, and to characterize appropriate points of departure (POD) for each RfD. An uncertainty factor of 300 was applied to each POD to yield RfD values of 0.1, 0.04, and 0.03 mg/kg-d for short-term, subchronic, and chronic exposures, respectively. Because a chronic cancer bioassay for SAN Trimer in rats (NTP 2011a) does not provide evidence of carcinogenicity, a cancer risk assessment is not appropriate for this chemical. Potential health hazards to human health were assessed using a hazard index (HI) approach, which considers the ratio of exposure dose (i.e., average daily dose, mg/kg-d) to toxicity dose (RfD, mg/kg-d) for each scenario. All CTE and RME HI values are well below 1 (where the average daily dose is equivalent to the RfD), indicating that there is no concern for potential noncancer effects in exposed populations even under the conservative assumptions of this screening-level assessment.

  6. A Transition Metal-Binding, Trimeric βγ-Crystallin from Methane-Producing Thermophilic Archaea, Methanosaeta thermophila.

    Science.gov (United States)

    Srivastava, Shanti Swaroop; Jamkhindikar, Aditya Anand; Raman, Rajeev; Jobby, Maroor K; Chadalawada, Swathi; Sankaranarayanan, Rajan; Sharma, Yogendra

    2017-03-07

    βγ-Crystallins are important constituents of the vertebrate eye lens, whereas in microbes, they are prevalent as Ca 2+ -binding proteins. In archaea, βγ-crystallins are conspicuously confined to two methanogens, viz., Methanosaeta and Methanosarcina. One of these, i.e., M-crystallin from Methanosarcina acetivorans, has been shown to be a typical Ca 2+ -binding βγ-crystallin. Here, with the aid of a high-resolution crystal structure and isothermal titration calorimetry, we report that "Methallin", a βγ-crystallin from Methanosaeta thermophila, is a trimeric, transition metal-binding protein. It binds Fe, Ni, Co, or Zn ion with nanomolar affinity, which is consistent even at 55 °C, the optimal temperature for the methanogen's growth. At the center of the protein trimer, the metal ion is coordinated by six histidines, two from each protomer, leading to an octahedral geometry. Small-angle X-ray scattering analysis confirms that the trimer seen in the crystal lattice is a biological assembly; this assembly dissociates to monomers upon removal of the metal ion. The introduction of two histidines (S17H/S19H) into a homologous βγ-crystallin, Clostrillin, allows it to bind nickel at the introduced site, though with micromolar affinity. However, because of the lack of a compatible interface, nickel binding could not induce trimerization, affirming that Methallin is a naturally occurring trimer for high-affinity transition metal binding. While βγ-crystallins are known to bind Ca 2+ and form homodimers and oligomers, the transition metal-binding, trimeric Methallin is a new paradigm for βγ-crystallins. The distinct features of Methallin, such as nickel or iron binding, are also possible imprints of biogeochemical changes during the period of its origin.

  7. Relevance of Simultaneous Mono-Ubiquitinations of Multiple Units of PCNA Homo-Trimers in DNA Damage Tolerance

    Science.gov (United States)

    Kanao, Rie; Masuda, Yuji; Deguchi, Saori; Yumoto-Sugimoto, Mayumi; Hanaoka, Fumio; Masutani, Chikahide

    2015-01-01

    DNA damage tolerance (DDT) pathways, including translesion synthesis (TLS) and additional unknown mechanisms, enable recovery from replication arrest at DNA lesions. DDT pathways are regulated by post-translational modifications of proliferating cell nuclear antigen (PCNA) at its K164 residue. In particular, mono-ubiquitination by the ubiquitin ligase RAD18 is crucial for Polη-mediated TLS. Although the importance of modifications of PCNA to DDT pathways is well known, the relevance of its homo-trimer form, in which three K164 residues are present in a single ring, remains to be elucidated. Here, we show that multiple units of a PCNA homo-trimer are simultaneously mono-ubiquitinated in vitro and in vivo. RAD18 catalyzed sequential mono-ubiquitinations of multiple units of a PCNA homo-trimer in a reconstituted system. Exogenous PCNA formed hetero-trimers with endogenous PCNA in WI38VA13 cell transformants. When K164R-mutated PCNA was expressed in these cells at levels that depleted endogenous PCNA homo-trimers, multiple modifications of PCNA complexes were reduced and the cells showed defects in DDT after UV irradiation. Notably, ectopic expression of mutant PCNA increased the UV sensitivities of Polη-proficient, Polη-deficient, and REV1-depleted cells, suggesting the disruption of a DDT pathway distinct from the Polη- and REV1-mediated pathways. These results suggest that simultaneous modifications of multiple units of a PCNA homo-trimer are required for a certain DDT pathway in human cells. PMID:25692884

  8. A procyanidin type A trimer from cinnamon extract attenuates glial cell swelling and the reduction in glutamate uptake following ischemic injury in vitro

    Science.gov (United States)

    Dietary polyphenols exert neuroprotective effects in ischemic injury. The protective effects of a procyanidin type A trimer (trimer 1) isolated from a water soluble cinnamon extract (CE) were investigated on key features of ischemic injury including cell swelling, increased free radical production, ...

  9. Evidence for a gold trimer on the Si(111)-√{ 21} ×√{ 21}-(Ag + Au) surface

    Science.gov (United States)

    Takahashi, Toshio; Yamaguchi, Yudai; Shirasawa, Tetsuroh; Voegeli, Wolfgang; Tajiri, Hiroo

    2018-02-01

    The structure of the Si(111)-√{ 21} ×√{ 21}-(Ag + Au) surface is studied using surface X-ray diffraction and fluorescence yield measurements. The fluorescence measurements indicate that there are three gold atoms in the unit cell. From the analysis of the diffraction data, it is shown that the gold atoms form a trimer with an interatomic distance of 2.8 Å and that the sides of the trimer are almost parallel to the Si(111)-1 × 1 fundamental vectors, similar to that in the Si(111)-√{ 3} ×√{ 3}-Au structure.

  10. Computational design of trimeric influenza-neutralizing proteins targeting the hemagglutinin receptor binding site

    Energy Technology Data Exchange (ETDEWEB)

    Strauch, Eva-Maria; Bernard, Steffen M.; La, David; Bohn, Alan J.; Lee, Peter S.; Anderson, Caitlin E.; Nieusma, Travis; Holstein, Carly A.; Garcia, Natalie K.; Hooper, Kathryn A.; Ravichandran, Rashmi; Nelson, Jorgen W.; Sheffler, William; Bloom, Jesse D.; Lee, Kelly K.; Ward, Andrew B.; Yager, Paul; Fuller, Deborah H.; Wilson, Ian A.; Baker , David (UWASH); (Scripps); (FHCRC)

    2017-06-12

    Many viral surface glycoproteins and cell surface receptors are homo-oligomers1, 2, 3, 4, and thus can potentially be targeted by geometrically matched homo-oligomers that engage all subunits simultaneously to attain high avidity and/or lock subunits together. The adaptive immune system cannot generally employ this strategy since the individual antibody binding sites are not arranged with appropriate geometry to simultaneously engage multiple sites in a single target homo-oligomer. We describe a general strategy for the computational design of homo-oligomeric protein assemblies with binding functionality precisely matched to homo-oligomeric target sites5, 6, 7, 8. In the first step, a small protein is designed that binds a single site on the target. In the second step, the designed protein is assembled into a homo-oligomer such that the designed binding sites are aligned with the target sites. We use this approach to design high-avidity trimeric proteins that bind influenza A hemagglutinin (HA) at its conserved receptor binding site. The designed trimers can both capture and detect HA in a paper-based diagnostic format, neutralizes influenza in cell culture, and completely protects mice when given as a single dose 24 h before or after challenge with influenza.

  11. Advanced environmentally friendly coatings prepared from amine-capped aniline trimer-based waterborne electroactive polyurethane

    Energy Technology Data Exchange (ETDEWEB)

    Huang, Hsiu-Ying; Huang, Tsao-Cheng [Department of Chemistry, Center for Nanotechnology at CYCU, Chung Yuan Christian University, Chung Li 32023, Taiwan, ROC (China); Lin, Jui-Chi [Department of Products at TTRI, Taiwan Textile Research Institute, Tucheng, Taipei 23674, Taiwan, ROC (China); Chang, Jung-Hsiang; Lee, Yu-Ting [Department of Chemistry, Center for Nanotechnology at CYCU, Chung Yuan Christian University, Chung Li 32023, Taiwan, ROC (China); Yeh, Jui-Ming, E-mail: juiming@cycu.edu.tw [Department of Chemistry, Center for Nanotechnology at CYCU, Chung Yuan Christian University, Chung Li 32023, Taiwan, ROC (China)

    2013-01-15

    Electroactive waterborne polyurethane (EWPU) containing conjugated segments of electroactive amino-capped aniline trimer (ACAT) unit was successfully prepared and characterized. Iodine-doped EWPU with conductivity 1.2 Multiplication-Sign 10{sup -6} S cm{sup -1} was detected. The electroactivity of EWPU was evaluated by performing electrochemical cyclic voltammetry studies. It was worth noting that the EWPU coating was found to exhibit enhanced corrosion protection effects on cold-rolled steel (CRS) electrodes as compared to the corresponding non-electroactive waterborne polyurethane (NEWPU) coating based on a series of electrochemical measurements in 3.5 wt% NaCl electrolyte. A possible mechanism for the enhanced corrosion protection ability of the EWPU coatings on the CRS electrode could be interpreted as the redox catalytic capabilities of the aniline trimer units existed in EWPU induce the formation of passive metal oxide layers on the CRS electrode, as further evidenced by SEM and XPS studies. The thermal stability of EWPU was also investigated by thermogravimetric analysis (TGA). Highlights: Black-Right-Pointing-Pointer Electroactive waterborne polyurethane (EWPU) was successfully synthesized. Black-Right-Pointing-Pointer Electroactivity of EWPU was investigated by electrochemical CV analysis. Black-Right-Pointing-Pointer Corrosion protection of EWPU coating was better than NEWPU.

  12. Managing light polarization via plasmon-molecule interactions within an asymmetric metal nanoparticle trimer

    International Nuclear Information System (INIS)

    Shegai, Timur; Li, Zhipeng; Zhang, Zhenyu; Xu, Hongxing; Haran, Gilad

    2008-01-01

    The interaction of light with metal nanoparticles leads to novel phenomena mediated by surface plasmon excitations. In this paper we use single molecules to characterize the interaction of surface plasmons with light, and show that such interaction can strongly modulate the polarization of the emitted light. The simplest nanostructures that enable such polarization modulation are asymmetric silver nanocrystal trimers, where individual Raman scattering molecules are located in the gap between two of the nanoparticles. The third particle breaks the dipolar symmetry of the two-particle junction, generating a wavelength-dependent polarization pattern. Indeed, the scattered light becomes elliptically polarized and its intensity pattern is rotated in the presence of the third particle. We use a combination of spectroscopic observations on single molecules, scanning electron microscope imaging, and generalized Mie theory calculations to provide a full picture of the effect of particles on the polarization of the emitted light. Furthermore, our theoretical analysis allows us to show that the observed phenomenon is very sensitive to the size of the trimer particles and their relative position, suggesting future means for precise control of light polarization on the nanoscale.

  13. Procyanidin trimer C1 derived from Theobroma cacao reactivates latent human immunodeficiency virus type 1 provirus.

    Science.gov (United States)

    Hori, Takanori; Barnor, Jacob; Huu, Tung Nguyen; Morinaga, Osamu; Hamano, Akiko; Ndzinu, Jerry; Frimpong, Angela; Minta-Asare, Keren; Amoa-Bosompem, Mildred; Brandful, James; Odoom, John; Bonney, Joseph; Tuffour, Isaac; Owusu, Baffour-Awuah; Ofosuhene, Mark; Atchoglo, Philip; Sakyiamah, Maxwell; Adegle, Richard; Appiah-Opong, Regina; Ampofo, William; Koram, Kwadwo; Nyarko, Alexander; Okine, Laud; Edoh, Dominic; Appiah, Alfred; Uto, Takuhiro; Yoshinaka, Yoshiyuki; Uota, Shin; Shoyama, Yukihiro; Yamaoka, Shoji

    2015-04-03

    Despite remarkable advances in combination antiretroviral therapy (cART), human immunodeficiency virus type 1 (HIV-1) infection remains incurable due to the incomplete elimination of the replication-competent virus, which persists in latent reservoirs. Strategies for targeting HIV reservoirs for eradication that involves reactivation of latent proviruses while protecting uninfected cells by cART are urgently needed for cure of HIV infection. We screened medicinal plant extracts for compounds that could reactivate the latent HIV-1 provirus and identified a procyanidin trimer C1 derived from Theobroma cacao as a potent activator of the provirus in human T cells latently infected with HIV-1. This reactivation largely depends on the NF-κB and MAPK signaling pathways because either overexpression of a super-repressor form of IκBα or pretreatment with a MEK inhibitor U0126 diminished provirus reactivation by C1. A pan-PKC inhibitor significantly blocked the phorbol ester-induced but not the C1-induced HIV-1 reactivation. Although C1-induced viral gene expression persisted for as long as 48 h post-stimulation, NF-κB-dependent transcription peaked at 12 h post-stimulation and then quickly declined, suggesting Tat-mediated self-sustainment of HIV-1 expression. These results suggest that procyanidin C1 trimer is a potential compound for reactivation of latent HIV-1 reservoirs. Copyright © 2015 Elsevier Inc. All rights reserved.

  14. X-ray and EM structures of a natively glycosylated HIV-1 envelope trimer.

    Science.gov (United States)

    Gristick, Harry B; Wang, Haoqing; Bjorkman, Pamela J

    2017-10-01

    The structural and biochemical characterization of broadly neutralizing anti-HIV-1 antibodies (bNAbs) has been essential in guiding the design of potential vaccines to prevent infection by HIV-1. While these studies have revealed critical mechanisms by which bNAbs recognize and/or accommodate N-glycans on the trimeric envelope glycoprotein (Env), they have been limited to the visualization of high-mannose glycan forms only, since heterogeneity introduced from the presence of complex glycans makes it difficult to obtain high-resolution structures. 3.5 and 3.9 Å resolution crystal structures of the HIV-1 Env trimer with fully processed and native glycosylation were solved, revealing a glycan shield of high-mannose and complex-type N-glycans that were used to define the complete epitopes of two bNAbs. Here, the refinement of the N-glycans in the crystal structures is discussed and comparisons are made with glycan densities in glycosylated Env structures derived by single-particle cryo-electron microscopy.

  15. Structure of a trimeric variant of the Epstein-Barr virus glycoprotein B

    Energy Technology Data Exchange (ETDEWEB)

    Backovic, Marija [Northwestern Univ., Evanston, IL (United States); Longnecker, Richard [Northwestern Univ., Chicago, IL (United States); Jardetzky, Theodore S [Northwestern Univ., Evanston, IL (United States)

    2009-03-16

    Epstein-Barr virus (EBV) is a herpesvirus that is associated with development of malignancies of lymphoid tissue. EBV infections are life-long and occur in >90% of the population. Herpesviruses enter host cells in a process that involves fusion of viral and cellular membranes. The fusion apparatus is comprised of envelope glycoprotein B (gB) and a heterodimeric complex made of glycoproteins H and L. Glycoprotein B is the most conserved envelope glycoprotein in human herpesviruses, and the structure of gB from Herpes simplex virus 1 (HSV-1) is available. Here, we report the crystal structure of the secreted EBV gB ectodomain, which forms 16-nm long spike-like trimers, structurally homologous to the postfusion trimers of the fusion protein G of vesicular stomatitis virus (VSV). Comparative structural analyses of EBV gB and VSV G, which has been solved in its pre and postfusion states, shed light on gB residues that may be involved in conformational changes and membrane fusion. Also, the EBV gB structure reveals that, despite the high sequence conservation of gB in herpesviruses, the relative orientations of individual domains, the surface charge distributions, and the structural details of EBV gB differ from the HSV-1 protein, indicating regions and residues that may have important roles in virus-specific entry.

  16. The four-transmembrane protein IP39 of Euglena forms strands by a trimeric unit repeat

    Science.gov (United States)

    Suzuki, Hiroshi; Ito, Yasuyuki; Yamazaki, Yuji; Mineta, Katsuhiko; Uji, Masami; Abe, Kazuhiro; Tani, Kazutoshi; Fujiyoshi, Yoshinori; Tsukita, Sachiko

    2013-01-01

    Euglenoid flagellates have striped surface structures comprising pellicles, which allow the cell shape to vary from rigid to flexible during the characteristic movement of the flagellates. In Euglena gracilis, the pellicular strip membranes are covered with paracrystalline arrays of a major integral membrane protein, IP39, a putative four-membrane-spanning protein with the conserved sequence motif of the PMP-22/EMP/MP20/Claudin superfamily. Here we report the three-dimensional structure of Euglena IP39 determined by electron crystallography. Two-dimensional crystals of IP39 appear to form a striated pattern of antiparallel double-rows in which trimeric IP39 units are longitudinally polymerised, resulting in continuously extending zigzag-shaped lines. Structural analysis revealed an asymmetric molecular arrangement in the trimer, and suggested that at least four different interactions between neighbouring protomers are involved. A combination of such multiple interactions would be important for linear strand formation of membrane proteins in a lipid bilayer. PMID:23612307

  17. Apa is a trimeric autotransporter adhesin of Actinobacillus pleuropneumoniae responsible for autoagglutination and host cell adherence.

    Science.gov (United States)

    Xiao, Longwen; Zhou, Liang; Sun, Changjiang; Feng, Xin; Du, ChongTao; Gao, Yu; Ji, Qun; Yang, Shuxin; Wang, Yu; Han, Wenyu; Langford, P R; Lei, Liancheng

    2012-10-01

    Actinobacillus pleuropneumoniae is the causative agent of porcine pleuropneumonia, and adherence to host cells is a key step in the pathogenic process. Although trimeric autotransporter adhesins (TAAs) were identified in many pathogenic bacteria in recent years, none in A. pleuropneumoniae have been characterized. In this study, we identified a TAA from A. pleuropneumoniae, Apa, and characterized the contribution of its amino acid residues to the adhesion process. Sequence analysis of the C-terminal amino acid residues of Apa revealed the presence of a putative translocator domain and six conserved HsfBD1-like or HsfBD2-like binding domains. Western blot analysis revealed that the 126 C-terminal amino acids of Apa could form trimeric molecules. By confocal laser scanning microscopy, one of these six domains (ApaBD3) was determined to mediate adherence to epithelial cells. Adherence assays and adherence inhibition assays using a recombinant E. coli- ApaBD3 strain which expressed ApaBD3 on the surface of E. coli confirmed that this domain was responsible for the adhesion activity. Moreover, cellular enzyme-linked immunosorbent assays demonstrated that ApaBD3 mediated high-level adherence to epithelial cell lines. Intriguingly, autoagglutination was observed with the E. coli- ApaBD3 strain, and this phenomenon was dependent upon the association of the expressed ApaBD3 with the C-terminal translocator domain. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  18. Rate Constant Change of Photo Reaction of Bacteriorhodopsin Observed in Trimeric Molecular System.

    Science.gov (United States)

    Tsujiuchi, Yutaka; Masumoto, Hiroshi; Goto, Takashi

    2016-04-01

    To elucidate the time evolution of photo reaction of bacteriorhodopsin in glycerol mixed purple membrane at around 196 K under irradiation by red light, a kinetic model was constructed. The change of absorption with irradiation at times of 560 nm and 412 nm was analyzed for the purpose of determining reaction rates of photo reaction of bacteriorhodopsin and its product M intermediate. In this study it is shown that reaction rates of conversion from bacteriorhodopsin to the M intermediate can be explained by a set of linear differential equations. This model analysis concludes that bacteriorhodopsin in which constitutes a trimer unit with other two bacteriorhodopsin molecules changes into M intermediates in the 1.73 of reaction rate, in the initial step, and according to the number of M intermediate in a trimer unit, from three to one, the reaction rate of bacteriorhodopsin into M intermediates smaller as 1.73, 0.80, 0.19 which caused by influence of inter-molecular interaction between bacteriorhodopsin.

  19. CD4-induced activation in a soluble HIV-1 Env trimer.

    Science.gov (United States)

    Guttman, Miklos; Garcia, Natalie K; Cupo, Albert; Matsui, Tsutomu; Julien, Jean-Philippe; Sanders, Rogier W; Wilson, Ian A; Moore, John P; Lee, Kelly K

    2014-07-08

    The HIV envelope glycoprotein (Env) trimer undergoes receptor-induced conformational changes that drive fusion of the viral and cellular membranes. Env conformational changes have been observed using low-resolution electron microscopy, but only large-scale rearrangements have been visible. Here, we use hydrogen-deuterium exchange and oxidative labeling to gain a more precise understanding of the unliganded and CD4-bound forms of soluble Env trimers (SOSIP.664), including their glycan composition. CD4 activation induces the reorganization of bridging sheet elements, V1/V2 and V3, much of the gp120 inner domain, and the gp41 fusion subunit. Two CD4 binding site-targeted inhibitors have substantially different effects: NBD-556 partially mimics CD4-induced destabilization of the V1/V2 and V3 crown, whereas BMS-806 only affects regions around the gp120/gp41 interface. The structural information presented here increases our knowledge of CD4- and small molecule-induced conformational changes in Env and the allosteric pathways that lead to membrane fusion. Copyright © 2014 Elsevier Ltd. All rights reserved.

  20. Cryo-EM structure of a native, fully glycosylated, cleaved HIV-1 envelope trimer.

    Science.gov (United States)

    Lee, Jeong Hyun; Ozorowski, Gabriel; Ward, Andrew B

    2016-03-04

    The envelope glycoprotein trimer (Env) on the surface of HIV-1 recognizes CD4(+) T cells and mediates viral entry. During this process, Env undergoes substantial conformational rearrangements, making it difficult to study in its native state. Soluble stabilized trimers have provided valuable insights into the Env structure, but they lack the hydrophobic membrane proximal external region (MPER, an important target of broadly neutralizing antibodies), the transmembrane domain, and the cytoplasmic tail. Here we present (i) a cryogenic electron microscopy (cryo-EM) structure of a clade B virus Env, which lacks only the cytoplasmic tail and is stabilized by the broadly neutralizing antibody PGT151, at a resolution of 4.2 angstroms and (ii) a reconstruction of this form of Env in complex with PGT151 and MPER-targeting antibody 10E8 at a resolution of 8.8 angstroms. These structures provide new insights into the wild-type Env structure. Copyright © 2016, American Association for the Advancement of Science.

  1. Therapeutic TNF Inhibitors can Differentially Stabilize Trimeric TNF by Inhibiting Monomer Exchange

    Science.gov (United States)

    van Schie, Karin A.; Ooijevaar-de Heer, Pleuni; Dijk, Lisanne; Kruithof, Simone; Wolbink, Gertjan; Rispens, Theo

    2016-01-01

    Tumor necrosis factor (TNF) is a homotrimeric cytokine that is a key mediator of inflammation. It is unstable at physiological concentrations and slowly converts into an inactive form. Here, we investigated the mechanism of this process by using a Förster resonance energy transfer (FRET) assay that allowed monitoring of monomeric subunit exchange in time. We observed continuous exchange of monomeric subunits even at concentrations of TNF high enough to maintain its bioactivity. The kinetics of this process closely corresponds with the appearance of monomeric subunits and disappearance of trimeric TNF in time at ng/ml concentrations as monitored by high-performance size-exclusion chromatography (HP-SEC). Furthermore, of the five therapeutic TNF inhibitors that are currently used in the clinic, three (adalimumab, infliximab, etanercept) were found to completely inhibit the monomer exchange reaction and stabilize TNF trimers, whereas golimumab and certolizumab could not prevent monomer exchange, but did slow down the exchange process. These differences were not correlated with the affinities of the TNF inhibitors, measured with both surface plasmon resonance (SPR) and in fluid phase using fluorescence-assisted HP-SEC. The stabilizing effect of these TNF inhibitors might result in prolonged residual TNF bioactivity under conditions of incomplete blocking, as observed in vitro for adalimumab. PMID:27605058

  2. AAFreqCoil: a new classifier to distinguish parallel dimeric and trimeric coiled coils.

    Science.gov (United States)

    Wang, Xiaofeng; Zhou, Yuan; Yan, Renxiang

    2015-07-01

    Coiled coils are characteristic rope-like protein structures, constituted by one or more heptad repeats. Native coiled-coil structures play important roles in various biological processes, while the designed ones are widely employed in medicine and industry. To date, two major oligomeric states (i.e. dimeric and trimeric states) of a coiled-coil structure have been observed, plausibly exerting different biological functions. Therefore, exploration of the relationship between heptad repeat sequences and coiled coil structures is highly important. In this paper, we develop a new method named AAFreqCoil to classify parallel dimeric and trimeric coiled coils. Our method demonstrated its competitive performance when benchmarked based on 10-fold cross validation and jackknife cross validation. Meanwhile, the rules that can explicitly explain the prediction results of the test coiled coil can be extracted from the AAFreqCoil model for a better explanation of user predictions. A web server and stand-alone program implementing the AAFreqCoil algorithm are freely available at .

  3. Destabilization of the PCNA trimer mediated by its interaction with the NEIL1 DNA glycosylase

    Energy Technology Data Exchange (ETDEWEB)

    Prakash, Aishwarya; Moharana, Kedar; Wallace, Susan S.; Doublié, Sylvie

    2016-12-19

    The base excision repair (BER) pathway repairs oxidized lesions in the DNA that result from reactive oxygen species generated in cells. If left unrepaired, these damaged DNA bases can disrupt cellular processes such as replication. NEIL1 is one of the 11 human DNA glycosylases that catalyze the first step of the BER pathway, i.e. recognition and excision of DNA lesions. NEIL1 interacts with essential replication proteins such as the ring-shaped homotrimeric proliferating cellular nuclear antigen (PCNA). We isolated a complex formed between NEIL1 and PCNA (±DNA) using size exclusion chromatography (SEC). This interaction was confirmed using native gel electrophoresis and mass spectrometry. Stokes radii measured by SEC hinted that PCNA in complex with NEIL1 (±DNA) was no longer a trimer. Height measurements and images obtained by atomic force microscopy also demonstrated the dissociation of the PCNA homotrimer in the presence of NEIL1 and DNA, while small-angle X-ray scattering analysis confirmed the NEIL1 mediated PCNA trimer dissociation and formation of a 1:1:1 NEIL1-DNA-PCNA(monomer) complex. Furthermore, ab initio shape reconstruction provides insights into the solution structure of this previously unreported complex. Together, these data point to a potential mechanistic switch between replication and BER.

  4. The sensitivity of approved Ninhydrin and Biuret tests in the assessment of protein contamination on surgical steel as an aid to prevent iatrogenic prion transmission.

    Science.gov (United States)

    Lipscomb, I P; Pinchin, H E; Collin, R; Harris, K; Keevil, C W

    2006-11-01

    Regulations recommend the routine application of biochemical tests, such as the Ninhydrin or Biuret tests, to confirm the efficacy of hospital sterile service department (SSD) washer-disinfector cycles in removing proteinaceous material, particularly with respect to prions. The effectiveness of these methods relies on both the effective sampling of the instruments and the sensitivity of the tests employed. Two commercially available contamination assessment tests were evaluated for their sensitivity to ME7 brain homogenate on surgical-grade stainless steel surfaces. Controls were visualized by the application of episcopic differential interference contrast/Epi-fluorecence microscopy (EDIC/EF) combined with the sensitive fluorescent reagent, SYPRO Ruby, which has been shown previously to rapidly visualize and assess low levels of contamination on medical devices. The Ninhydrin test displayed a minimum level of detection observed by 75% of volunteers (MLD(75)) of 9.25 microg [95% confidence interval (95% CI) 8.6-10.0 microg]. The Biuret test provided better sensitivity, with a MLD(75) of 6.7 microg (95% CI 5.4-8.2 microg). However, much lower concentrations of proteinaceous soiling (pg) were visualized using the EDIC/EF microscopy method. From these findings, it is clear that these approved colorimetric tests of cleaning are relatively insensitive. This investigation demonstrates how large amounts (up to 6.5 microg) of proteinaceous brain contamination could remain undetected and the instruments deemed clean using such methods. The application of more sensitive cleanliness evaluation methods should be applied to reduce the risk of iatrogenic transmission of prion disease in 'high-risk' instruments such as neurosurgical devices.

  5. Stabilization of the soluble, cleaved, trimeric form of the envelope glycoprotein complex of human immunodeficiency virus type 1

    NARCIS (Netherlands)

    Sanders, Rogier W.; Vesanen, Mika; Schuelke, Norbert; Master, Aditi; Schiffner, Linnea; Kalyanaraman, Roopa; Paluch, Maciej; Berkhout, Ben; Maddon, Paul J.; Olson, William C.; Lu, Min; Moore, John P.

    2002-01-01

    The envelope glycoprotein (Env) complex of human immunodeficiency virus type I has evolved a structure that is minimally immunogenic while retaining its natural function of receptor-mediated virus-cell fusion. The Env complex is trimeric; its six individual subunits (three gp120 and three gp41

  6. Characterization of humoral responses to soluble trimeric HIV gp140 from a clade A Ugandan field isolate

    DEFF Research Database (Denmark)

    Visciano, Maria Luisa; Tagliamonte, Maria; Stewart-Jones, Guillaume

    2013-01-01

    with neutralizing activity against homologous and heterologous viruses. In the present study, we generated a recombinant trimeric soluble gp140, derived from a previously identified Ugandan A-clade HIV field isolate (gp14094UG018). Antibodies elicited in immunized rabbits show a broad binding pattern to HIV...

  7. Photon emission statistics and photon tracking in single-molecule spectroscopy of molecular aggregates : Dimers and trimers

    NARCIS (Netherlands)

    Bloemsma, E. A.; Knoester, J.

    2012-01-01

    Based on the generating function formalism, we investigate broadband photon statistics of emission for single dimers and trimers driven by a continuous monochromatic laser field. In particular, we study the first and second moments of the emission statistics, which are the fluorescence excitation

  8. Intramolecular trimerization, a novel strategy for making multispecific antibodies with controlled orientation of the antigen binding domains

    DEFF Research Database (Denmark)

    Álvarez-Cienfuegos, Ana; Alanes, Natalia Nuñez del Prado; Compte, Marta

    2016-01-01

    Here, we describe a new strategy that allows the rapid and efficient engineering of mono and multispecific trivalent antibodies. By fusing single-domain antibodies from camelid heavy-chain-only immunoglobulins (VHHs) to the N-terminus of a human collagen XVIII trimerization domain (TIEXVIII) we p...

  9. Direct Probing of Germinal Center Responses Reveals Immunological Features and Bottlenecks for Neutralizing Antibody Responses to HIV Env Trimer

    NARCIS (Netherlands)

    Havenar-Daughton, Colin; Carnathan, Diane G.; Torrents de la Peña, Alba; Pauthner, Matthias; Briney, Bryan; Reiss, Samantha M.; Wood, Jennifer S.; Kaushik, Kirti; van Gils, Marit J.; Rosales, Sandy L.; van der Woude, Patricia; Locci, Michela; Le, Khoa M.; de Taeye, Steven W.; Sok, Devin; Mohammed, Ata Ur Rasheed; Huang, Jessica; Gumber, Sanjeev; Garcia, AnaPatricia; Kasturi, Sudhir P.; Pulendran, Bali; Moore, John P.; Ahmed, Rafi; Seumois, Grégory; Burton, Dennis R.; Sanders, Rogier W.; Silvestri, Guido; Crotty, Shane

    2016-01-01

    Generating tier 2 HIV-neutralizing antibody (nAb) responses by immunization remains a challenging problem, and the immunological barriers to induction of such responses with Env immunogens remain unclear. Here, some rhesus monkeys developed autologous tier 2 nAbs upon HIV Env trimer immunization

  10. Triphenylene discotic liquid crystal trimers synthesized by Co2(CO8-catalyzed terminal alkyne [2 + 2 + 2] cycloaddition

    Directory of Open Access Journals (Sweden)

    Bin Han

    2013-12-01

    Full Text Available The synthesis of star-shaped discotic liquid crystal trimers using Co2(CO8-catalyzed terminal alkyne [2 + 2 + 2] cycloaddition reaction is reported. The trimers consist of three triphenylene discotic units linked to a central 1,2,4-trisubstituted benzene ring via flexible spacers. The trimers were synthesized in the yields up to 70% by mixing the monomers with 10 mol % of Co2(CO8 as the catalyst in refluxing 1,4-dioxane. The liquid crystalline properties were investigated by using polarizing optical microscopy (POM, differential scanning calorimetry (DSC and X-ray diffraction (XRD. Trimer 4 with an ester connecting group and a longer spacer exhibited a rectangular columnar mesophase, while 5b and 5c possessing an ether linkage and a shorter spacer display a hexagonal columnar mesophase. The connecting functional group and the length of the flexible spacer between the central benzene ring and the triphenylene units have pivotal influence on the mesomorphism.

  11. Subcellular redistribution of trimeric G-proteins – potential mechanism of desensitization of hormone response: internalisation, solubilization, down-regulation

    Czech Academy of Sciences Publication Activity Database

    Drastichová, Zdeňka; Bouřová, Lenka; Lisý, Václav; Hejnová, L.; Rudajev, Vladimír; Stöhr, Jiří; Durchánková, Dana; Ostašov, Pavel; Teisinger, Jan; Soukup, Tomáš; Novotný, Jiří; Svoboda, Petr

    2008-01-01

    Roč. 57, Suppl.3 (2008), S1-S10 ISSN 0862-8408 R&D Projects: GA MŠk(CZ) LC554; GA ČR(CZ) GA309/06/0121 Institutional research plan: CEZ:AV0Z50110509 Keywords : brain * subcellular fractionation * trimeric G-proteins Subject RIV: CE - Biochemistry Impact factor: 1.653, year: 2008

  12. Differential binding of neutralizing and non-neutralizing antibodies to native-like soluble HIV-1 Env trimers, uncleaved Env proteins, and monomeric subunits.

    Science.gov (United States)

    Yasmeen, Anila; Ringe, Rajesh; Derking, Ronald; Cupo, Albert; Julien, Jean-Philippe; Burton, Dennis R; Ward, Andrew B; Wilson, Ian A; Sanders, Rogier W; Moore, John P; Klasse, Per Johan

    2014-05-29

    The trimeric envelope glycoproteins (Env) on the surface of HIV-1 virions are the targets for neutralizing antibodies (NAbs). No candidate HIV-1 immunogen has yet induced potent, broadly active NAbs (bNAbs). Part of the explanation may be that previously tested Env proteins inadequately mimic the functional, native Env complex. Trimerization and the proteolytic processing of Env precursors into gp120 and gp41 profoundly alter antigenicity, but soluble cleaved trimers are too unstable to serve as immunogens. By introducing stabilizing mutations (SOSIP), we constructed soluble, cleaved Env trimers derived from the HIV-1 subtype A isolate BG505 that resemble native Env spikes on virions both structurally and antigenically. We used surface plasmon resonance (SPR) to quantify antibody binding to different forms of BG505 Env: the proteolytically cleaved SOSIP.664 trimers, cleaved gp120-gp41ECTO protomers, and gp120 monomers. Non-NAbs to the CD4-binding site bound only marginally to the trimers but equally well to gp120-gp41ECTO protomers and gp120 monomers, whereas the bNAb VRC01, directed to the CD4bs, bound to all three forms. In contrast, bNAbs to V1V2 glycan-dependent epitopes bound preferentially (PG9 and PG16) or exclusively (PGT145) to trimers. We also explored the antigenic consequences of three different features of SOSIP.664 gp140 trimers: the engineered inter-subunit disulfide bond, the trimer-stabilizing I559P change in gp41ECTO, and proteolytic cleavage at the gp120-gp41ECTO junction. Each of these three features incrementally promoted native-like trimer antigenicity. We compared Fab and IgG versions of bNAbs and validated a bivalent model of IgG binding. The NAbs showed widely divergent binding kinetics and degrees of binding to native-like BG505 SOSIP.664. High off-rate constants and low stoichiometric estimates of NAb binding were associated with large amounts of residual infectivity after NAb neutralization of the corresponding BG505.T332N pseudovirus

  13. Antigenic and 3D structural characterization of soluble X4 and hybrid X4-R5 HIV-1 Env trimers.

    Science.gov (United States)

    Arnold, Philipp; Himmels, Patricia; Weiß, Svenja; Decker, Tim-Michael; Markl, Jürgen; Gatterdam, Volker; Tampé, Robert; Bartholomäus, Patrick; Dietrich, Ursula; Dürr, Ralf

    2014-05-30

    HIV-1 is decorated with trimeric glycoprotein spikes that enable infection by engaging CD4 and a chemokine coreceptor, either CCR5 or CXCR4. The variable loop 3 (V3) of the HIV-1 envelope protein (Env) is the main determinant for coreceptor usage. The predominant CCR5 using (R5) HIV-1 Env has been intensively studied in function and structure, whereas the trimeric architecture of the less frequent, but more cytopathic CXCR4 using (X4) HIV-1 Env is largely unknown, as are the consequences of sequence changes in and near V3 on antigenicity and trimeric Env structure. Soluble trimeric gp140 Env constructs were used as immunogenic mimics of the native spikes to analyze their antigenic properties in the context of their overall 3D structure. We generated soluble, uncleaved, gp140 trimers from a prototypic T-cell line-adapted (TCLA) X4 HIV-1 strain (NL4-3) and a hybrid (NL4-3/ADA), in which the V3 spanning region was substituted with that from the primary R5 isolate ADA. Compared to an ADA (R5) gp140, the NL4-3 (X4) construct revealed an overall higher antibody accessibility, which was most pronounced for the CD4 binding site (CD4bs), but also observed for mAbs against CD4 induced (CD4i) epitopes and gp41 mAbs. V3 mAbs showed significant binding differences to the three constructs, which were refined by SPR analysis. Of interest, the NL4-3/ADA construct with the hybrid NL4-3/ADA CD4bs showed impaired CD4 and CD4bs mAb reactivity despite the presence of the essential elements of the CD4bs epitope. We obtained 3D reconstructions of the NL4-3 and the NL4-3/ADA gp140 trimers via electron microscopy and single particle analysis, which indicates that both constructs inherit a propeller-like architecture. The first 3D reconstruction of an Env construct from an X4 TCLA HIV-1 strain reveals an open conformation, in contrast to recently published more closed structures from R5 Env. Exchanging the X4 V3 spanning region for that of R5 ADA did not alter the open Env architecture as

  14. Fragmentation dynamics of ionized neon trimer inside helium nanodroplets: a theoretical study.

    Science.gov (United States)

    Bonhommeau, David; Viel, Alexandra; Halberstadt, Nadine

    2004-06-22

    We report a theoretical study of the fragmentation dynamics of Ne(3) (+) inside helium nanodroplets, following vertical ionization of the neutral neon trimer. The motion of the neon atoms is treated classically, while transitions between the electronic states of the ionic cluster are treated quantum mechanically. A diatomics-in-molecules description of the potential energy surfaces is used, in a minimal basis set consisting of three effective p orbitals on each neon atom for the missing electron. The helium environment is modeled by a friction force acting on the neon atoms when their speed exceeds the Landau velocity. A reasonable range of values for the corresponding friction coefficient is obtained by comparison with existing experimental measurements. (c) 2004 American Institute of Physics.

  15. Spatial dependence of the super-exchange interactions for transition-metal trimers in graphene

    Science.gov (United States)

    Crook, Charles B.; Houchins, Gregory; Zhu, Jian-Xin; Balatsky, Alexander V.; Constantin, Costel; Haraldsen, Jason T.

    2018-01-01

    This study examines the magnetic interactions between spatially variable manganese and chromium trimers substituted into a graphene superlattice. Using density functional theory, we calculate the electronic band structure and magnetic populations for the determination of the electronic and magnetic properties of the system. To explore the super-exchange coupling between the transition-metal atoms, we establish the magnetic ground states through a comparison of multiple magnetic and spatial configurations. Through an analysis of the electronic and magnetic properties, we conclude that the presence of transition-metal atoms can induce a distinct magnetic moment in the surrounding carbon atoms as well as produce a Ruderman-Kittel-Kasuya-Yosida-like super-exchange coupling. It is hoped that these simulations can lead to the realization of spintronic applications in graphene through electronic control of the magnetic clusters.

  16. Diverse Regulation of Temperature Sensation by Trimeric G-Protein Signaling in Caenorhabditis elegans.

    Directory of Open Access Journals (Sweden)

    Tomoyo Ujisawa

    Full Text Available Temperature sensation by the nervous system is essential for life and proliferation of animals. The molecular-physiological mechanisms underlying temperature signaling have not been fully elucidated. We show here that diverse regulatory machinery underlies temperature sensation through trimeric G-protein signaling in the nematode Caenorhabditis elegans. Molecular-genetic studies demonstrated that cold tolerance is regulated by additive functions of three Gα proteins in a temperature-sensing neuron, ASJ, which is also known to be a light-sensing neuron. Optical recording of calcium concentration in ASJ upon temperature-changes demonstrated that three Gα proteins act in different aspects of temperature signaling. Calcium concentration changes in ASJ upon temperature change were unexpectedly decreased in a mutant defective in phosphodiesterase, which is well known as a negative regulator of calcium increase. Together, these data demonstrate commonalities and differences in the molecular components concerned with light and temperature signaling in a single sensory neuron.

  17. Diverse Regulation of Temperature Sensation by Trimeric G-Protein Signaling in Caenorhabditis elegans

    Science.gov (United States)

    Ujisawa, Tomoyo; Ohta, Akane; Uda-Yagi, Misato

    2016-01-01

    Temperature sensation by the nervous system is essential for life and proliferation of animals. The molecular-physiological mechanisms underlying temperature signaling have not been fully elucidated. We show here that diverse regulatory machinery underlies temperature sensation through trimeric G-protein signaling in the nematode Caenorhabditis elegans. Molecular-genetic studies demonstrated that cold tolerance is regulated by additive functions of three Gα proteins in a temperature-sensing neuron, ASJ, which is also known to be a light-sensing neuron. Optical recording of calcium concentration in ASJ upon temperature-changes demonstrated that three Gα proteins act in different aspects of temperature signaling. Calcium concentration changes in ASJ upon temperature change were unexpectedly decreased in a mutant defective in phosphodiesterase, which is well known as a negative regulator of calcium increase. Together, these data demonstrate commonalities and differences in the molecular components concerned with light and temperature signaling in a single sensory neuron. PMID:27788246

  18. Solution Synthesis of Semiconducting Two-Dimensional Polymer via Trimerization of Carbonitrile.

    Science.gov (United States)

    Liu, Jingjing; Zan, Wu; Li, Ke; Yang, Yang; Bu, Fanxing; Xu, Yuxi

    2017-08-30

    The synthesis of crystalline two-dimensional polymers (2DPs) with proper bandgaps and well-defined repeating units presents a great challenge to synthetic chemists. Here we report the first solution synthesis of a single-layer/few-layer triazine-based 2DP via trimerization of carbonitrile at the interface of dichloromethane and trifluoromethanesulfonic acid. The processable triazine-based 2DP can be assembled into mechanically strong layered free-standing films with a high specific surface area via filtration. Moreover, the highly crystalline triazine-based 2DP can function as the active semiconducting layer in a field-effect transistor via drop coating and exhibits slightly bipolar behavior with a high on/off ratio of 10 3 and a remarkable mobility of 0.15 cm 2 V -1 s -1 .

  19. Esterase variation in Turkish white-toothed shrews (Crocidura: Record of a trimeric esterase

    Directory of Open Access Journals (Sweden)

    Tez C.

    2009-01-01

    Full Text Available This study focuses on esterase variation of the genus Crocidura in Turkey. A total of 248 white-toothed shrews were analyzed by means of cellulose acetate gel electrophoresis. Liver tissue and alfa naphthyl acetate were used to investigate esterase variation in Turkish white-toothed shrews. A different esterase banding pattern was found in one Crocidura individual. This phenotype had four anodally migrated bands on cellulose acetate gel. The Crocidura individual displaying the given phenotype was identified as Crocidura suaveolens. The different esterase banding pattern observed in this study is considered to be a result of the trimeric structure of esterase in the lesser white-toothed shrew (Crocidura suaveolens.

  20. 3-(2,3,5,6,7,8-Hexahydro-1H-cyclo­penta­[b]quinolin-9-yl)-1,5-bis­(4-methoxy­phen­yl)biuret

    Science.gov (United States)

    Sakurai, Kaori; Noguchi, Keiichi; Nishibe, Koichiro

    2010-01-01

    Ipidacrine (2,3,5,6,7,8-hexa­hydro-1H-cyclo­penta­[b]quinolin-9-amine) was reacted with 4-methoxy­phenyl isocyanate to give the title compound, C28H30N4O4. An intra­molecular N—H⋯O hydrogen bond results in an essentially planar [r.m.s. deviation from the mean plane is 0.126 (1) Å] conformation for the biuret unit. The central ring of the quinoline unit is twisted by 78.2 (1)° with respect to the biuret mean plane, whereas the two 4-methoxy­benzene rings are twisted out of this plane by 24.3 (1)° and 48.5 (1)°, resulting in an overall propeller-like structure. An inter­molecular N—H⋯N hydrogen bond between the biuret NH atom and the quinoline ring nitro­gen defines the crystal packing. PMID:21580406

  1. Characterization of bacterial diversity in an atrazine degrading enrichment culture and degradation of atrazine, cyanuric acid and biuret in industrial wastewater.

    Science.gov (United States)

    Dutta, Anirban; Vasudevan, Venugopal; Nain, Lata; Singh, Neera

    2016-01-01

    An enrichment culture was used to study atrazine degradation in mineral salt medium (MSM) (T1), MSM+soil extract (1:1, v/v) (T2) and soil extract (T3). Results suggested that enrichment culture required soil extract to degrade atrazine, as after second sequential transfer only partial atrazine degradation was observed in T1 treatment while atrazine was completely degraded in T2 and T3 treatments even after fourth transfer. Culture independent polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) technique confirmed selective enrichment of genus Bacillus along with Pseudomonas and Burkholderia. Degradation of atrazine/metabolites in the industrial wastewater was studied at different initial concentrations of the contaminants [wastewater-water (v/v) ratio: T1, 1:9; T2, 2:8; T3, 3:7; T4, 5:5 and T5, undiluted effluent]. The initial concentrations of atrazine, cyanuric acid and biuret ranged between 5.32 and 53.92 µg mL(-1), 265.6 and 1805.2 µg mL(-1) and 1.85 and 16.12 µg mL(-1), respectively. The enrichment culture was able to completely degrade atrazine, cyanuric acid and biuret up to T4 treatment, while no appreciable degradation of contaminants was observed in the undiluted effluent (T5). Inability of enrichment culture to degrade atrazine/metabolites might be due to high concentrations of cyanuric acid. Therefore, a separate study on cyanuric acid degradation suggested: (i) no appreciable cyanuric acid degradation with accumulation of an unidentified metabolite in the medium where cyanuric acid was supplemented as the sole source of carbon and nitrogen; (ii) partial cyanuric acid degradation with accumulation of unidentified metabolite in the medium containing additional nitrogen source; and (iii) complete cyanuric acid degradation in the medium supplemented with an additional carbon source. This unidentified metabolite observed during cyanuric acid degradation and also detected in the enrichment culture inoculated wastewater samples

  2. Hydrolyzable tannins of tamaricaceous plants. V. Structures of monomeric-trimeric tannins and cytotoxicity of macrocyclic-type tannins isolated from Tamarix nilotica (1).

    Science.gov (United States)

    Orabi, Mohamed A A; Taniguchi, Shoko; Sakagami, Hiroshi; Yoshimura, Morio; Yoshida, Takashi; Hatano, Tsutomu

    2013-05-24

    Three new ellagitannin monomers, nilotinins M5-M7 (1-3), a dimer, nilotinin D10 (4), and a trimer, nilotinin T1 (5), together with three known dimers, hirtellin D (7) and tamarixinins B (8) and C (9), and a trimer, hirtellin T2 (6), were isolated from Tamarix nilotica dried leaves. The structures of the tannins were elucidated by intensive spectroscopic methods and chemical conversions into known tannins. The new trimer (5) is a unique macrocyclic type whose monomeric units are linked together by an isodehydrodigalloyl and two dehydrodigalloyl moieties. Additionally, dimeric and trimeric macrocyclic-type tannins isolated from T. nilotica in this study were assessed for possible cytotoxic activity against four human tumor cell lines. Tumor-selective cytotoxicities of the tested compounds were higher than those of synthetic and natural potent cytotoxic compounds, including polyphenols, and comparable with those of 5-fluorouracil and melphalan.

  3. A computational study of dimers and trimers of nitrosyl hydride: Blue shift of NH bonds that are involved in H-bond and orthogonal interactions

    International Nuclear Information System (INIS)

    Solimannejad, Mohammad; Massahi, Shokofeh; Alkorta, Ibon

    2009-01-01

    Ab initio calculations at MP2/aug-cc-pVTZ level were used to analyze the interactions between nitrosyl hydride (HNO) dimers and trimers. The structures obtained have been analyzed with the Atoms in Molecules (AIMs) and Natural Bond Orbital (NBO) methodologies. Four minima were located on the potential energy surface of the dimers. Nine different structures have been obtained for the trimers. Three types of interactions are observed, NH···N and NH···O hydrogen bonds and orthogonal interaction between the lone pair of the oxygen with the electron-deficient region of the nitrogen atom. Stabilization energies of dimers and trimers including BSSE and ZPE are in the range 4-8 kJ mol -1 and 12-19 kJ mol -1 , respectively. Blue shift of NH bond upon complex formation in the ranges between 30-80 and 14,114 cm -1 is predicted for dimers and trimers, respectively.

  4. Two scandium-biuret complexes: [Sc(C2H5N3O2)(H2O)5]Cl3 x H2O and [Sc(C2H5N3O2)4](NO3)3.

    Science.gov (United States)

    Harrison, William T A

    2008-05-01

    The scandium(III) cations in the structures of pentaaqua(biuret-kappa(2)O,O')scandium(III) trichloride monohydrate, [Sc(C(2)H(5)N(3)O(2))(H(2)O)(5)]Cl(3) x H(2)O, (I), and tetrakis(biuret-kappa(2)O,O')scandium(III) trinitrate, [Sc(C(2)H(5)N(3)O(2))(4)](NO(3))(3), (II), are found to adopt very different coordinations with the same biuret ligand. The roles of hydrogen bonding and the counter-ion in the establishment of the structures are described. In (I), the Sc(3+) cation adopts a fairly regular pentagonal bipyramidal coordination geometry arising from one O,O'-bidentate biuret molecule and five water molecules. A dense network of N-H...Cl, O-H...O and O-H...Cl hydrogen bonds help to establish the packing, resulting in dimeric associations of two cations and two water molecules. In (II), the Sc(3+) cation (site symmetry 2) adopts a slightly squashed square-antiprismatic geometry arising from four O,O'-bidentate biuret molecules. A network of N-H...O hydrogen bonds help to establish the packing, which features [010] chains of cations. One of the nitrate ions is disordered about an inversion centre. Both structures form three-dimensional hydrogen-bond networks.

  5. Design and structure of two HIV-1 clade C SOSIP.664 trimers that increase the arsenal of native-like Env immunogens.

    Science.gov (United States)

    Julien, Jean-Philippe; Lee, Jeong Hyun; Ozorowski, Gabriel; Hua, Yuanzi; Torrents de la Peña, Alba; de Taeye, Steven W; Nieusma, Travis; Cupo, Albert; Yasmeen, Anila; Golabek, Michael; Pugach, Pavel; Klasse, P J; Moore, John P; Sanders, Rogier W; Ward, Andrew B; Wilson, Ian A

    2015-09-22

    A key challenge in the quest toward an HIV-1 vaccine is design of immunogens that can generate a broadly neutralizing antibody (bnAb) response against the enormous sequence diversity of the HIV-1 envelope glycoprotein (Env). We previously demonstrated that a recombinant, soluble, fully cleaved SOSIP.664 trimer based on the clade A BG505 sequence is a faithful antigenic and structural mimic of the native trimer in its prefusion conformation. Here, we sought clade C native-like trimers with comparable properties. We identified DU422 and ZM197M SOSIP.664 trimers as being appropriately thermostable (Tm of 63.4 °C and 62.7 °C, respectively) and predominantly native-like, as determined by negative-stain electron microscopy (EM). Size exclusion chromatography, ELISA, and surface plasmon resonance further showed that these trimers properly display epitopes for all of the major bnAb classes, including quaternary-dependent, trimer-apex (e.g., PGT145) and gp120/gp41 interface (e.g., PGT151) epitopes. A cryo-EM reconstruction of the ZM197M SOSIP.664 trimer complexed with VRC01 Fab against the CD4 binding site at subnanometer resolution revealed a striking overall similarity to its BG505 counterpart with expected local conformational differences in the gp120 V1, V2, and V4 loops. These stable clade C trimers contribute additional diversity to the pool of native-like Env immunogens as key components of strategies to induce bnAbs to HIV-1.

  6. Controllable supramolecular structures and luminescent properties of unique trimeric Zn(II) 8-hydroxyquinolinates tuned by functional substituents.

    Science.gov (United States)

    Yuan, Guozan; Huo, Yanping; Nie, Xiaoli; Jiang, Hong; Liu, Bin; Fang, Xiaoming; Zhao, Fenghua

    2013-02-28

    We reported here the self-assembly of two supramolecular structures based on similar trimeric Zn(II) units that are built from novel 2-substituted 8-hydroxyquinoline ligands and coordination Zn(II) ions. The aggregation behavior of zinc salt and ligand in solution was investigated by a variety of techniques, including (1)H NMR, UV-vis and photoluminescence (PL). In the solid state, the supramolecular structures can be controlled by the substituted groups (-NO(2) and -F) via intermolecular interaction, such as π···π stacking, C-H···O, and C-F···F-C interactions. As a result, the two trimeric Zn(II) complexes exhibit disparate photophysical properties. The present research holds great promise in the development of novel multinuclear Zn(II) materials, and may contribute to the understanding of structure-property relationships.

  7. Energy transfer dynamics in trimers and aggregates of light-harvesting complex II probed by 2D electronic spectroscopy.

    Science.gov (United States)

    Enriquez, Miriam M; Akhtar, Parveen; Zhang, Cheng; Garab, Győző; Lambrev, Petar H; Tan, Howe-Siang

    2015-06-07

    The pathways and dynamics of excitation energy transfer between the chlorophyll (Chl) domains in solubilized trimeric and aggregated light-harvesting complex II (LHCII) are examined using two-dimensional electronic spectroscopy (2DES). The LHCII trimers and aggregates exhibit the unquenched and quenched excitonic states of Chl a, respectively. 2DES allows direct correlation of excitation and emission energies of coupled states over population time delays, hence enabling mapping of the energy flow between Chls. By the excitation of the entire Chl b Qy band, energy transfer from Chl b to Chl a states is monitored in the LHCII trimers and aggregates. Global analysis of the two-dimensional (2D) spectra reveals that energy transfer from Chl b to Chl a occurs on fast and slow time scales of 240-270 fs and 2.8 ps for both forms of LHCII. 2D decay-associated spectra resulting from the global analysis identify the correlation between Chl states involved in the energy transfer and decay at a given lifetime. The contribution of singlet-singlet annihilation on the kinetics of Chl energy transfer and decay is also modelled and discussed. The results show a marked change in the energy transfer kinetics in the time range of a few picoseconds. Owing to slow energy equilibration processes, long-lived intermediate Chl a states are present in solubilized trimers, while in aggregates, the population decay of these excited states is significantly accelerated, suggesting that, overall, the energy transfer within the LHCII complexes is faster in the aggregated state.

  8. Fractional Mott insulator-to-superfluid transition of Bose-Hubbard model in a trimerized Kagomé optical lattice

    Science.gov (United States)

    Chen, Qi-Hui; Li, Peng; Su, Haibin

    2016-06-01

    By generalizing the traditional single-site strong coupling expansion approach to a cluster one, we study the zero-temperature phase diagram of bosonic atoms in a trimerized Kagomé optical lattice. Some new features are present in this system. Due to the strong intra-trimer hopping interaction, there will be a new Mott insulator (MI), which is by definition incompressible but with a fractional filling per trimer. This is different from the traditional MI, which has an integral filling and originates only from the repulsive interaction between particles. We investigate the MI-to-superfluid transition and the nature of the fractional MI by calculating the critical exponents of phase transitions and the low-lying energy excitation spectra of quasiparticles (quasihole). We will show how the low-energy properties of this system can be understood qualitatively as a Bose-Hubbard model in triangular lattice from the point of view of the cluster strong coupling expansion. We also discuss how our results are related to experiment by studying the Bragg spectroscopy.

  9. HIV-1 Nef responsiveness is determined by Env variable regions involved in trimer association and correlates with neutralization sensitivity.

    Science.gov (United States)

    Usami, Yoshiko; Göttlinger, Heinrich

    2013-11-14

    HIV-1 Nef and the unrelated murine leukemia virus glycoGag similarly enhance the infectivity of HIV-1 virions. We now show that the effects of Nef and glycoGag are similarly determined by variable regions of HIV-1 gp120 that control Env trimer association and neutralization sensitivity. Whereas neutralization-sensitive X4-tropic Env proteins conferred high responsiveness to Nef and glycoGag, particles bearing neutralization-resistant R5-tropic Envs were considerably less affected. The profoundly different Nef/glycoGag responsiveness of a neutralization-resistant and a neutralization-sensitive R5-tropic Env could be switched by exchanging their gp120 V1/V2 regions, which also switches their neutralization sensitivity. Within V1/V2, the same determinants governed Nef/glycoGag responsiveness and neutralization sensitivity, indicating that these phenotypes are mechanistically linked. The V1/V2 and V3 regions, which form an apical trimer-association domain, together determined the Nef and glycoGag responsiveness of an X4-tropic Env. Our results suggest that Nef and glycoGag counteract the inactivation of Env spikes with relatively unstable apical trimer-association domains. Copyright © 2013 The Authors. Published by Elsevier Inc. All rights reserved.

  10. Structure of an HIV-1-neutralizing antibody target, the lipid-bound gp41 envelope membrane proximal region trimer.

    Science.gov (United States)

    Reardon, Patrick N; Sage, Harvey; Dennison, S Moses; Martin, Jeffrey W; Donald, Bruce R; Alam, S Munir; Haynes, Barton F; Spicer, Leonard D

    2014-01-28

    The membrane proximal external region (MPER) of HIV-1 glycoprotein (gp) 41 is involved in viral-host cell membrane fusion. It contains short amino acid sequences that are binding sites for the HIV-1 broadly neutralizing antibodies 2F5, 4E10, and 10E8, making these binding sites important targets for HIV-1 vaccine development. We report a high-resolution structure of a designed MPER trimer assembled on a detergent micelle. The NMR solution structure of this trimeric domain, designated gp41-M-MAT, shows that the three MPER peptides each adopt symmetric α-helical conformations exposing the amino acid side chains of the antibody binding sites. The helices are closely associated at their N termini, bend between the 2F5 and 4E10 epitopes, and gradually separate toward the C termini, where they associate with the membrane. The mAbs 2F5 and 4E10 bind gp41-M-MAT with nanomolar affinities, consistent with the substantial exposure of their respective epitopes in the trimer structure. The traditional structure determination of gp41-M-MAT using the Xplor-NIH protocol was validated by independently determining the structure using the DISCO sparse-data protocol, which exploits geometric arrangement algorithms that guarantee to compute all structures and assignments that satisfy the data.

  11. Structural and biochemical characterization of the biuret hydrolase (BiuH) from the cyanuric acid catabolism pathway of Rhizobium leguminasorum bv. viciae 3841.

    Science.gov (United States)

    Esquirol, Lygie; Peat, Thomas S; Wilding, Matthew; Lucent, Del; French, Nigel G; Hartley, Carol J; Newman, Janet; Scott, Colin

    2018-01-01

    Biuret deamination is an essential step in cyanuric acid mineralization. In the well-studied atrazine degrading bacterium Pseudomonas sp. strain ADP, the amidase AtzE catalyzes this step. However, Rhizobium leguminosarum bv. viciae 3841 uses an unrelated cysteine hydrolase, BiuH, instead. Herein, structures of BiuH, BiuH with bound inhibitor and variants of BiuH are reported. The substrate is bound in the active site by a hydrogen bonding network that imparts high substrate specificity. The structure of the inactive Cys175Ser BiuH variant with substrate bound in the active site revealed that an active site cysteine (Cys175), aspartic acid (Asp36) and lysine (Lys142) form a catalytic triad, which is consistent with biochemical studies of BiuH variants. Finally, molecular dynamics simulations highlighted the presence of three channels from the active site to the enzyme surface: a persistent tunnel gated by residues Val218 and Gln215 forming a potential substrate channel and two smaller channels formed by Val28 and a mobile loop (including residues Phe41, Tyr47 and Met51) that may serve as channels for co-product (ammonia) or co-substrate (water).

  12. Structural and biochemical characterization of the biuret hydrolase (BiuH from the cyanuric acid catabolism pathway of Rhizobium leguminasorum bv. viciae 3841.

    Directory of Open Access Journals (Sweden)

    Lygie Esquirol

    Full Text Available Biuret deamination is an essential step in cyanuric acid mineralization. In the well-studied atrazine degrading bacterium Pseudomonas sp. strain ADP, the amidase AtzE catalyzes this step. However, Rhizobium leguminosarum bv. viciae 3841 uses an unrelated cysteine hydrolase, BiuH, instead. Herein, structures of BiuH, BiuH with bound inhibitor and variants of BiuH are reported. The substrate is bound in the active site by a hydrogen bonding network that imparts high substrate specificity. The structure of the inactive Cys175Ser BiuH variant with substrate bound in the active site revealed that an active site cysteine (Cys175, aspartic acid (Asp36 and lysine (Lys142 form a catalytic triad, which is consistent with biochemical studies of BiuH variants. Finally, molecular dynamics simulations highlighted the presence of three channels from the active site to the enzyme surface: a persistent tunnel gated by residues Val218 and Gln215 forming a potential substrate channel and two smaller channels formed by Val28 and a mobile loop (including residues Phe41, Tyr47 and Met51 that may serve as channels for co-product (ammonia or co-substrate (water.

  13. Excitonic Coupling in Linear and Trefoil Trimer Perylenediimide Molecules Probed by Single-Molecule Spectroscopy

    KAUST Repository

    Yoo, Hyejin

    2012-10-25

    Perylenediimide (PDI) molecules are promising building blocks for photophysical studies of electronic interactions within multichromophore arrays. Such PDI arrays are important materials for fabrication of molecular nanodevices such as organic light-emitting diodes, organic semiconductors, and biosensors because of their high photostability, chemical and physical inertness, electron affinity, and high tinctorial strength over the entire visible spectrum. In this work, PDIs have been organized into linear (L3) and trefoil (T3) trimer molecules and investigated by single-molecule fluorescence microscopy to probe the relationship between molecular structures and interchromophoric electronic interactions. We found a broad distribution of coupling strengths in both L3 and T3 and hence strong/weak coupling between PDI units by monitoring spectral peak shifts in single-molecule fluorescence spectra upon sequential photobleaching of each constituent chromophore. In addition, we used a wide-field defocused imaging technique to resolve heterogeneities in molecular structures of L3 and T3 embedded in a PMMA polymer matrix. A systematic comparison between the two sets of experimental results allowed us to infer the correlation between intermolecular interactions and molecular structures. Our results show control of the PDI intermolecular interactions using suitable multichromophoric structures. © 2012 American Chemical Society.

  14. A Rac1--GDP trimer complex binds zinc with tetrahedral and octahedral coordination, displacing magnesium

    Energy Technology Data Exchange (ETDEWEB)

    Prehna, G.; Stebbins, C

    2007-01-01

    The Rho family of small GTPases represent well characterized signaling molecules that regulate many cellular functions such as actin cytoskeletal arrangement and the cell cycle by acting as molecular switches. A Rac1-GDP-Zn complex has been crystallized in space group P3221 and its crystal structure has been solved at 1.9 {angstrom} resolution. These trigonal crystals reveal the unexpected ability of Rac1 to coordinate Zn atoms in a tetrahedral fashion by use of its biologically relevant switch I and switch II regions. Upon coordination of zinc, the switch I region is stabilized in the GDP-bound conformation and contributes to a Rac1 trimer in the asymmetric unit. Zinc coordination causes switch II to adopt a novel conformation with a symmetry-related molecule. Additionally, zinc was found to displace magnesium from its octahedral coordination at switch I, although GDP binding remained stable. This structure represents the first reported Rac1-GDP-Zn complex, which further underscores the conformational flexibility and versatility of the small GTPase switch regions.

  15. A Rac1-GDP Trimer Complex Binds Zinc with Tetrahedral and Octahedral Coordination, Displacing Magnesium

    Energy Technology Data Exchange (ETDEWEB)

    Prehna,G.; Stebbins, E.

    2007-01-01

    The Rho family of small GTPases represent well characterized signaling molecules that regulate many cellular functions such as actin cytoskeletal arrangement and the cell cycle by acting as molecular switches. A Rac1-GDP-Zn complex has been crystallized in space group P3{sub 2}21 and its crystal structure has been solved at 1.9 {angstrom} resolution. These trigonal crystals reveal the unexpected ability of Rac1 to coordinate Zn atoms in a tetrahedral fashion by use of its biologically relevant switch I and switch II regions. Upon coordination of zinc, the switch I region is stabilized in the GDP-bound conformation and contributes to a Rac1 trimer in the asymmetric unit. Zinc coordination causes switch II to adopt a novel conformation with a symmetry-related molecule. Additionally, zinc was found to displace magnesium from its octahedral coordination at switch I, although GDP binding remained stable. This structure represents the first reported Rac1-GDP-Zn complex, which further underscores the conformational flexibility and versatility of the small GTPase switch regions.

  16. Linear Trimeric Hafnium Clusters in Hf0.86(1I3

    Directory of Open Access Journals (Sweden)

    Jan Beekhuizen

    2011-05-01

    Full Text Available The reduction of hafnium tetraiodide, HfI4, with aluminum at 600 °C or 850 °C in the presence of a NaI flux resulted in black single crystals of Hf0.86(1I3. This composition corresponds well to the upper end of the non-stoichiometry range 0.89 ≤ x ≤ 1.00 previously reported for HfxI3. The crystal structure (a = 1250.3(2, c = 1999.6(3 pm, R-3m, Z = 18 is made up of hexagonal closest packed layers of iodide ions. One third of the octahedral holes would be filled as in TiI3 or ZrI3 if it were Hf1.00I3. In Hf0.86(1I3, one out of six octahedral holes along [001] are, however, only occupied by 16%. In contrast to TiI3-I and ZrI3, one striking structural feature is in the formation of linear hafnium trimers with identical Hf―Hf distances of 318.3(2 pm rather than the formation of dimers. These may be associated with Hf―Hf bonding although only 2.64 electrons are available for one Hf5.16I18 column.

  17. Bioinformatic Characterization of the Trimeric Intracellular Cation-Specific Channel Protein Family

    Science.gov (United States)

    Silverio, Abe L. F.

    2014-01-01

    Trimeric intracellular cation-specific (TRIC) channels are integral to muscle excitation–contraction coupling. TRIC channels provide counter-ionic flux when calcium is rapidly transported from intracellular stores to the cell cytoplasm. Until recently, knowledge of the presence of these proteins was limited to animals. We analyzed the TRIC family and identified a profusion of prokaryotic family members with topologies and motifs similar to those of their eukaryotic counterparts. Prokaryotic members far outnumber eukaryotic members, and although none has been functionally characterized, the evidence suggests that they function as secondary carriers. The presence of fused N- or C-terminal domains of known biochemical functions as well as genomic context analyses provide clues about the functions of these prokaryotic homologs. They are proposed to function in metabolite (e.g., amino acid/ nucleotide) efflux. Phylogenetic analysis revealed that TRIC channel homologs diverged relatively early during evolutionary history and that horizontal gene transfer was frequent in prokaryotes but not in eukaryotes. Topological analyses of TRIC channels revealed that these proteins possess seven putative transmembrane segments (TMSs), which arose by intragenic duplication of a three-TMS polypeptide-encoding genetic element followed by addition of a seventh TMS at the C terminus to give the precursor of all current TRIC family homologs. We propose that this family arose in prokaryotes. PMID:21519847

  18. Crystal structure of the catalytic trimer of Methanococcus jannaschii aspartate transcarbamoylase.

    Science.gov (United States)

    Vitali, Jacqueline; Colaneri, Michael J; Kantrowitz, Evan

    2008-05-15

    The catalytic trimer of Methanococcus jannaschii aspartate transcarbamoylase is extremely heat stable, maintaining 75% of its activity after heat treatment for 60 min at 75 degrees C. We undertook its structural analysis in order to understand the molecular basis of its thermostability and gain insight on how its catalytic function adapts to high temperature. Several structural elements potentially contributing to thermostability were identified. These include: (i) changes in the amino acid composition such as a decrease in the thermolabile residues Gln and Asn, an increase in the charged residues Lys and Glu, an increase in Tyr and a decrease in Ala residues; (ii) a larger number of salt bridges, in particular, the improvement of ion-pair networks; (iii) shortening of the N-terminus and shortening of three loops. An interesting feature of the crystal structure is the association of two crystallographically independent catalytic subunits into a staggered complex with an intertrimer distance of 33.8 A. The active site appears similar to Escherichia coli. Upon substrate binding, smaller changes in the global orientation of domains and larger conformational changes of the active site residues are expected as compared to E. coli. 2007 Wiley-Liss, Inc.

  19. Complementation of diverse HIV-1 Env defects through cooperative subunit interactions: a general property of the functional trimer

    Directory of Open Access Journals (Sweden)

    Salzwedel Karl

    2009-08-01

    Full Text Available Abstract Background The HIV-1 Env glycoprotein mediates virus entry by catalyzing direct fusion between the virion membrane and the target cell plasma membrane. Env is composed of two subunits: gp120, which binds to CD4 and the coreceptor, and gp41, which is triggered upon coreceptor binding to promote the membrane fusion reaction. Env on the surface of infected cells is a trimer consisting of three gp120/gp41 homo-dimeric protomers. An emerging question concerns cooperative interactions between the protomers in the trimer, and possible implications for Env function. Results We extended studies on cooperative subunit interactions within the HIV-1 Env trimer, using analysis of functional complementation between coexpressed inactive variants harboring different functional deficiencies. In assays of Env-mediated cell fusion, complementation was observed between variants with a wide range of defects in both the gp120 and gp41 subunits. The former included gp120 subunits mutated in the CD4 binding site or incapable of coreceptor interaction due either to mismatched specificity or V3 loop mutation. Defective gp41 variants included point mutations at different residues within the fusion peptide or heptad repeat regions, as well as constructs with modifications or deletions of the membrane proximal tryptophan-rich region or the transmembrane domain. Complementation required the defective variants to be coexpressed in the same cell. The observed complementation activities were highly dependent on the assay system. The most robust activities were obtained with a vaccinia virus-based expression and reporter gene activation assay for cell fusion. In an alternative system involving Env expression from integrated provirus, complementation was detected in cell fusion assays, but not in virus particle entry assays. Conclusion Our results indicate that Env function does not require every subunit in the trimer to be competent for all essential activities. Through

  20. Stabilizing the native trimer of HIV-1 Env by destabilizing the heterodimeric interface of the gp41 postfusion six-helix bundle.

    Science.gov (United States)

    Kesavardhana, Sannula; Varadarajan, Raghavan

    2014-09-01

    The HIV-1 envelope glycoprotein (Env) is a trimer of gp120-gp41 heterodimers and is essential for viral entry. The gp41 subunit in native, prefusion trimeric Env exists in a metastable conformation and attains a stable six-helix bundle (6-HB) conformation comprised of a trimer of N-heptad repeat (NHR) and C-heptad repeat (CHR) heterodimers, that drives the fusion of viral and cellular membranes. We attempted to stabilize native Env trimers by incorporation of mutations at the NHR-CHR interface that disrupt the postfusion 6-HB of gp41. The mutations V570D and I573D stabilize native Env of the HIV-1 JRFL strain and occlude nonneutralizing epitopes to a greater extent than the previously identified I559P mutation that is at the interface of the NHR trimers in the 6-HB. The mutations prevent soluble-CD4 (sCD4)-induced gp120 shedding and 6-HB formation. In the context of cell surface-expressed JRFL Env, introduction of a previously reported additional disulfide between residues A501 and T605 perturbs the native conformation, though this effect is partially alleviated by furin coexpression. The data suggest that positions 570 and 573 are surface proximal in native Env and that the NHR homotrimeric coiled coil in native Env terminates before or close to residue 573. Aspartic acid substitutions at these positions stabilize native trimers through destabilization of the postfusion 6-HB conformation. These mutations can be used to stabilize Env in a DNA vaccine format. The major protein on the surface of HIV-1 is the envelope (Env) glycoprotein. Env is a trimer of gp120-gp41 heterodimers. gp120 is involved in receptor/coreceptor binding and gp41 in the fusion of viral and cellular membranes. Like many other viral fusion proteins, the gp41 subunit in native trimeric Env exists in a metastable conformation. gp41 readily forms a stable six-helix bundle (6-HB) conformation comprised of a trimer of N-heptad repeat (NHR) and C-heptad repeat (CHR) heterodimers that drives fusion of

  1. Clarifying and illustrating the electronic energy transfer pathways in trimeric and hexameric aggregation state of cyanobacteria allophycocyanin within the framework of Förster theory.

    Science.gov (United States)

    Ren, Yanliang; Melhem, Osama; Li, Yongjian; Chi, Bo; Han, Xinya; Zhu, Hao; Feng, Lingling; Wan, Jian; Xu, Xin

    2015-01-30

    Within the framework of the Förster theory, the electronic excitation energy transfer pathways in the cyanobacteria allophycocyanin (APC) trimer and hexamer were studied. The associated physical quantities (i.e., excitation energy, oscillator strength, and transition dipole moments) of the phycocyanobilins (PCBs) located in APC were calculated at time-dependent density functional theory (TDDFT) level of theory. To estimate the influence of protein environment on the preceding calculated physical quantities, the long-range interactions were approximately considered with the polarizable continuum model at the TDDFT level of theory, and the short-range interaction caused by surrounding aspartate residue of PCBs were taken into account as well. The shortest energy transfer time calculated in the framework of the Förster model at TDDFT/B3LYP/6-31+G* level of theory are about 0.10 ps in the APC trimer and about 170 ps in the APC monomer, which are in qualitative agreement with the experimental finding that a very fast lifetime of 0.43-0.44 ps in APC trimers, whereas its monomers lacked any corresponding lifetime. These results suggest that the lifetime of 0.43-0.44 ps in the APC trimers determined by Sharkov et al. was most likely attributed to the energy transfer of α(1) -84 ↔ β(3) -84 (0.23 ps), β(1) -84 ↔ α(2) -84 (0.11 ps) or β(2) -84 ↔ α(3) -84 (0.10 ps). So far, no experimental or theoretical energy transfer rates between two APC trimmers were reported, our calculations predict that the predominate energy transfer pathway between APC trimers is likely to occur from α(3) -84 in one trimer to α(5) -84 in an adjacent trimer with a rate of 32.51 ps. © 2014 Wiley Periodicals, Inc.

  2. Daily and alternate day supplementation of urea or biuret to ruminants consuming low-quality forage: I. Effects on cow performance and the efficiency of nitrogen use in wethers.

    Science.gov (United States)

    Currier, T A; Bohnert, D W; Falck, S J; Bartle, S J

    2004-05-01

    Two experiments were conducted to determine the influence of supplemental nonprotein N (NPN) provided daily (D) or every other day (2D) on ruminant performance and N efficiency. Treatments included an unsupplemented control (CON) and a urea (28.7% CP) or biuret (28.6% CP) supplement provided D or 2D at 0700. In Exp. 1, five wethers (39 +/- 1 kg BW) were used in an incomplete 5 x 4 Latin square with four 24-d periods to determine the influence of supplemental NPN source and supplementation frequency (SF) on the efficiency of N use in lambs consuming low-quality grass straw (4% CP). The amount of CP supplied by each supplement was approximately 0.10% of BW/d (averaged over a 2-d period). In Exp. 2, 80 Angus x Hereford cows (540 +/- 8 kg BW) in the last third of gestation were used to determine the effect of NPN source and SF on cow performance. The NPN treatments were formulated to provide 90% of the estimated degradable intake protein requirement. The supplemented treatments received the same amount of supplemental N over a 2-d period; therefore, the 2D treatments received double the quantity of supplemental N on their respective supplementation day than the D treatments. In Exp. 1, total DM, OM, and N intake; DM, OM, and N digestibility; N balance; and digested N retained were greater (P 0.05) between NPN sources or SF. Plasma urea-N (PUN) was increased with N supplementation compared with CON (P biuret (P biuret as the primary source of supplemental N can be effectively used by lambs and cows consuming low-quality forage, even when provided every other day.

  3. Novel Anti-Nicotine Vaccine Using a Trimeric Coiled-Coil Hapten Carrier.

    Directory of Open Access Journals (Sweden)

    Keith D Miller

    Full Text Available Tobacco addiction represents one of the largest public health problems in the world and is the leading cause of cancer and heart disease, resulting in millions of deaths a year. Vaccines for smoking cessation have shown considerable promise in preclinical models, although functional antibody responses induced in humans are only modestly effective in preventing nicotine entry into the brain. The challenge in generating serum antibodies with a large nicotine binding capacity is made difficult by the fact that this drug is non-immunogenic and must be conjugated as a hapten to a protein carrier. To circumvent the limitations of traditional carriers like keyhole limpet hemocyanin (KLH, we have synthesized a short trimeric coiled-coil peptide (TCC that creates a series of B and T cell epitopes with uniform stoichiometry and high density. Here we compared the relative activities of a TCC-nic vaccine and two control KLH-nic vaccines using Alum as an adjuvant or GLA-SE, which contains a synthetic TLR4 agonist formulated in a stable oil-in-water emulsion. The results showed that the TCC's high hapten density correlated with a better immune response in mice as measured by anti-nicotine Ab titer, affinity, and specificity, and was responsible for a reduction in anti-carrier immunogenicity. The Ab responses achieved with this synthetic vaccine resulted in a nicotine binding capacity in serum that could prevent >90% of a nicotine dose equivalent to three smoked cigarettes (0.05 mg/kg from reaching the brain.

  4. Antigenic and immunosuppressive properties of a trimeric recombinant transmembrane envelope protein gp41 of HIV-1.

    Directory of Open Access Journals (Sweden)

    Michael Mühle

    Full Text Available The transmembrane envelope (TM protein gp41 of the human immunodeficiency virus-1 (HIV-1 plays an important role during virus infection inducing the fusion of the viral and cellular membranes. In addition, there are indications that the TM protein plays a role in the immunopathogenesis leading to the acquired immunodeficiency syndrome (AIDS. Inactivated virus particles and recombinant gp41 have been reported to inhibit lymphocyte proliferation, as well as to alter cytokine release and gene expression. The same was shown for a peptide corresponding to a highly conserved domain of all retroviral TM proteins, the immunosuppressive domain. Due to its propensity to aggregate and to be expressed at low levels, studies comprising authentic gp41 produced in eukaryotic cells are extremely rare. Here we describe the production of a secreted, soluble recombinant gp41 in 293 cells. The antigen was purified to homogeneity and characterised thoroughly by various biochemical and immunological methods. It was shown that the protein was glycosylated and assembled into trimers. Binding studies by ELISA and surface plasmon resonance using conformation-specific monoclonal antibodies implied a six-helix bundle conformation. The low binding of broadly neutralising antibodies (bnAb directed against the membrane proximal external region (MPER suggested that this gp41 is probably not suited as vaccine to induce such bnAb. Purified gp41 bound to monocytes and to a lesser extent to lymphocytes and triggered the production of specific cytokines when added to normal peripheral blood mononuclear cells. In addition, gp41 expressed on target cells inhibited the antigen-specific response of murine CD8+ T cells by drastically impairing their IFNγ production. To our knowledge, this is the first comprehensive analysis of a gp41 produced in eukaryotic cells including its immunosuppressive properties. Our data provide another line of evidence that gp41 might be directly involved in

  5. Interactions of cationic trimeric, gemini and monomeric surfactants with trianionic curcumin in aqueous solution.

    Science.gov (United States)

    Wang, Meina; Wu, Chunxian; Tang, Yongqiang; Fan, Yaxun; Han, Yuchun; Wang, Yilin

    2014-05-21

    Interactions of trianionic curcumin (Cur(3-)) with a series of cationic surfactants, monomeric surfactant dodecyl trimethylammonium bromide (DTAB), dimeric surfactant hexamethylene-1,6-bis(dodecyldimethylammonium bromide) (12-6-12) and trimeric surfactant tri(dodecyldimethylammonioacetoxy)diethyltriamine trichloride (DTAD), have been investigated in aqueous solution of pH 13.0. Surface tension and spectral measurements indicate that the cationic surfactants display a similar surfactant concentration dependent interaction process with Cur(3-), involving three interaction stages. At first the three cationic surfactants electrostatically bind on Cur(3-) to form the surfactant-Cur(3-) complex. Then the bound and unbound cationic surfactants with Cur(3-) aggregate into surfactant-Cur(3-) mixed micelles through hydrophobic interactions above the critical micelle concentration of the surfactants (CMCC) in the presence of Cur(3-). Finally excess unbound surfactants self-assemble into micelles like those without Cur(3-). For all the three surfactants, the addition of Cur(3-) only decreases the critical micelle concentration of 12-6-12 but does not affect the critical micelle concentration of DTAB and DTAD. As the oligomeric degree of surfactants increases, the intermolecular interaction of the cationic surfactants with Cur(3-) increases and the surfactant amount needed for Cur(3-) encapsulation decreases. Compared with 12-6-12, either the weaker interaction of DTAB with Cur(3-) or stronger interaction of DTAD with Cur(3-) limits the stability or solubility of Cur(3-) in surfactant micelles. Therefore, gemini surfactant 12-6-12 is the best choice to effectively suppress Cur(3-) degradation at very low concentrations. Isothermal titration microcalorimetry, surface tension and (1)H NMR results reveal that 12-6-12 and Cur(3-) form a (12-6-12)2-Cur(3-) complex and start to form micelles at extremely decreased concentrations, where either 12-6-12 or Cur(3-) works as a bridge

  6. Comparative assessment of early acute lung injury in mice and rats exposed to 1,6-hexamethylene diisocyanate-polyisocyanate aerosols.

    Science.gov (United States)

    Pauluhn, Jürgen

    2008-05-02

    The aliphatic diisocyanate monomer 1,6-hexamethylene diisocyanate (HDI) is used as a building block for non-volatile polycondensation products, such as HDI-isocyanurate (HDI-IC) and HDI-biuret (HDI-BT). This paper describes the results from acute inhalation studies with these types of polyisocyanate aerosols in OF1 and C57BL/6J mice and in Wistar rats. The modifying role of different concentrations of residual HDI in HDI-BT on pulmonary irritation was also addressed. These data close data gaps for acute mouse inhalation studies in direct comparison with rats. The sensory irritant potency was examined in OF1 mice during a 3h nose-only exposure to the polyisocyanate aerosols. Concurrent with exposure, breathing patterns suitable to distinguish upper/lower respiratory tract irritation where examined. Functional measurements in barometric plethysmographs (Penh) addressed changes in respiratory function in C57BL/6J mice exposed for 6h up to 16h postexposure. These measurements revealed that these polyisocyanates elicit changes slow in onset suggestive of pulmonary irritation rather than upper airway irritation. This conclusion was supported by similarly exposed OF1 mice exposed to non-irritant, surface active respirable particles of amorphous silica. In C57BL/6J mice and Wistar rats, nose-only exposed for 6h to 10mg/m(3) of aerosolized HDI-BT HDI (0.1% or 2% residual HDI), the pulmonary irritation potency was comparatively assessed by bronchoalveolar lavage (BAL) on postexposure day 1. Similarly air-exposed animals served as concurrent controls. Most changes in BAL suggestive of acute pulmonary irritation were more pronounced in Wistar rats than in C57BL/6J mice. A conclusive dependence of BAL endpoints on the residual content of residual HDI monomer in the polyisocyanate was not found. The results of this study show that mice may be particularly suitable to functionally analyze at which location of the respiratory tract predominant irritation may occur. However, with

  7. {Ba[Au(SCN2]2}n: a three-dimensional net comprised of monomeric and trimeric gold(I units

    Directory of Open Access Journals (Sweden)

    A. Darren Back

    2010-07-01

    Full Text Available The noteworthy structural feature of the title complex, poly[acetonitriletetra-μ2-thiocyanato-barium(IIdigold(I], {[Au2Ba(SCN4(CH3CN]}n, is that the bis(thiocyanatoaurate(I anion adopts both monomeric and trimeric motifs. The trimer unit has an Au...Au distance of 3.1687 (3 Å. In both the monomeric and trimeric units, the AuI atoms are also bonded to two S atoms. Within the trimeric unit, the AuI atoms exist in differing environments; one Au atom has a T-shaped three-coordinate geometry while the other has a square-planar four-coordinate geometry. The AuI atom of the monomer adopts a linear two-coordinate geometry. The extended structure can be described as a three-dimensional coordination polymer consisting of chains of Ba atoms bridged by thiocyanate N atoms. These chains are cross-linked via the gold monomeric and trimeric units.

  8. Lateral manipulation of small clusters on the Cu and Ag(1 1 1) surfaces with the single-atom and trimer-apex tips: Reliability study

    International Nuclear Information System (INIS)

    Xie Yiqun; Liu Fen; Huang Lei

    2010-01-01

    We study the reliability of the lateral manipulation of small Cu clusters (dimer and trimer) on the flat Cu(1 1 1) surface with both the single-atom and trimer-apex tips and that for the Ag/Ag(1 1 1) system, and compare the results between the two systems as well as with the single-atom manipulation on these surfaces. Manipulations are simulated using molecular statics method with semi-empirical potentials. The dependence of the manipulation reliability on the tip height and tip orientation are investigated. Overall, the manipulation reliability increases with decreasing tip height although it depends obviously on the tip orientation. For the Cu/Cu(1 1 1) system, the manipulation of the dimmer and trimer can be successful with both tips. The manipulation reliability can be improved by the trimer-apex tip, and the tip-height range for the successful manipulation is also broader, as compared to the single-atom apex tip. Differently from the single-atom manipulation, the tip orientation has a noticeable influence on the manipulation reliability even for the single-atom tip due to the stronger tip-cluster and surface-adatom interactions in cluster manipulation. For the Ag/Ag(1 1 1) system, successful manipulations only be achieved with the trimer-apex tip, and the manipulation reliability is worse than that of the Cu/Cu(1 1 1) system, indicating the difference in mechanic properties between the two surfaces at the atomic level.

  9. Particulate Array of Well-Ordered HIV Clade C Env Trimers Elicits Neutralizing Antibodies that Display a Unique V2 Cap Approach.

    Science.gov (United States)

    Martinez-Murillo, Paola; Tran, Karen; Guenaga, Javier; Lindgren, Gustaf; Àdori, Monika; Feng, Yu; Phad, Ganesh E; Vázquez Bernat, Néstor; Bale, Shridhar; Ingale, Jidnyasa; Dubrovskaya, Viktoriya; O'Dell, Sijy; Pramanik, Lotta; Spångberg, Mats; Corcoran, Martin; Loré, Karin; Mascola, John R; Wyatt, Richard T; Karlsson Hedestam, Gunilla B

    2017-05-16

    The development of soluble envelope glycoprotein (Env) mimetics displaying ordered trimeric symmetry has ushered in a new era in HIV-1 vaccination. The recently reported native, flexibly linked (NFL) design allows the generation of native-like trimers from clinical isolates at high yields and homogeneity. As the majority of infections world-wide are of the clade C subtype, we examined responses in non-human primates to well-ordered subtype C 16055 trimers administered in soluble or high-density liposomal formats. We detected superior germinal center formation and enhanced autologous neutralizing antibodies against the neutralization-resistant (tier 2) 16055 virus following inoculation of liposome-arrayed trimers. Epitope mapping of the neutralizing monoclonal antibodies (mAbs) indicated major contacts with the V2 apex, and 3D electron microscopy reconstructions of Fab-trimer complexes revealed a horizontal binding angle to the Env spike. These vaccine-elicited mAbs target the V2 cap, demonstrating a means to accomplish tier 2 virus neutralization by penetrating the dense N-glycan shield. Copyright © 2017 Elsevier Inc. All rights reserved.

  10. Structural delineation of a quaternary, cleavage-dependent epitope at the gp41-gp120 interface on intact HIV-1 Env trimers.

    Science.gov (United States)

    Blattner, Claudia; Lee, Jeong Hyun; Sliepen, Kwinten; Derking, Ronald; Falkowska, Emilia; de la Peña, Alba Torrents; Cupo, Albert; Julien, Jean-Philippe; van Gils, Marit; Lee, Peter S; Peng, Wenjie; Paulson, James C; Poignard, Pascal; Burton, Dennis R; Moore, John P; Sanders, Rogier W; Wilson, Ian A; Ward, Andrew B

    2014-05-15

    All previously characterized broadly neutralizing antibodies to the HIV-1 envelope glycoprotein (Env) target one of four major sites of vulnerability. Here, we define and structurally characterize a unique epitope on Env that is recognized by a recently discovered family of human monoclonal antibodies (PGT151-PGT158). The PGT151 epitope is comprised of residues and glycans at the interface of gp41 and gp120 within a single protomer and glycans from both subunits of a second protomer and represents a neutralizing epitope that is dependent on both gp120 and gp41. Because PGT151 binds only to properly formed, cleaved trimers, this distinctive property, and its ability to stabilize Env trimers, has enabled the successful purification of mature, cleaved Env trimers from the cell surface as a complex with PGT151. Here we compare the structural and functional properties of membrane-extracted Env trimers from several clades with those of the soluble, cleaved SOSIP gp140 trimer. Copyright © 2014 Elsevier Inc. All rights reserved.

  11. Conditional trimerization and lytic activity of HIV-1 gp41 variants containing the membrane-associated segments.

    Science.gov (United States)

    Dai, Zhou; Tao, Yisong; Liu, Nina; Brenowitz, Michael D; Girvin, Mark E; Lai, Jonathan R

    2015-03-03

    Fusion of host and viral membranes is a critical step during infection by membrane-bound viruses. The HIV-1 glycoproteins gp120 (surface subunit) and gp41 (fusion subunit) represent the prototypic system for studying this process; in the prevailing model, the gp41 ectodomain forms a trimeric six-helix bundle that constitutes a critical intermediate and provides the energetic driving force for overcoming barriers associated with membrane fusion. However, most structural studies of gp41 variants have been performed either on ectodomain constructs lacking one or more of the membrane-associated segments (the fusion peptide, FP, the membrane-proximal external region, MPER, and the transmembrane domain, TM) or on variants consisting of these isolated segments alone without the ectodomain. Several recent reports have suggested that the HIV-1 ectodomain, as well as larger construct containing the membrane-bound segments, dissociates from a trimer to a monomer in detergent micelles. Here we compare the properties of a series of gp41 variants to delineate the roles of the ectodomain, FP, and MPER and TM, all in membrane-mimicking environments. We find that these proteins are prone to formation of a monomer in detergent micelles. In one case, we observed exclusive monomer formation at pH 4 but conditional trimerization at pH 7 even at low micromolar (∼5 μM) protein concentrations. Liposome release assays demonstrate that these gp41-related proteins have the capacity to induce content leakage but that this activity is also strongly modulated by pH with much higher activity at pH 4. Circular dichroism, nuclear magnetic resonance, and binding assays with antibodies specific to the MPER provide insight into the structural and functional roles of the FP, MPER, and TM and their effect on structure within the larger context of the fusion subunit.

  12. Human surfactant protein A2 gene mutations impair dimmer/trimer assembly leading to deficiency in protein sialylation and secretion.

    Directory of Open Access Journals (Sweden)

    Yi Song

    Full Text Available Surfactant protein A2 (SP-A2 plays an essential role in surfactant metabolism and lung host defense. SP-A2 mutations in the carbohydrate recognition domain have been related to familial pulmonary fibrosis and can lead to a recombinant protein secretion deficiency in vitro. In this study, we explored the molecular mechanism of protein secretion deficiency and the subsequent biological effects in CHO-K1 cells expressing both wild-type and several different mutant forms of SP-A2. We demonstrate that the SP-A2 G231V and F198S mutants impair the formation of dimmer/trimer SP-A2 which contributes to the protein secretion defect. A deficiency in sialylation, but not N-linked glycosylation, is critical to the observed dimmer/trimer impairment-induced secretion defect. Furthermore, both mutant forms accumulate in the ER and form NP-40-insoluble aggregates. In addition, the soluble mutant SP-A2 could be partially degraded through the proteasome pathway but not the lysosome or autophagy pathway. Intriguingly, 4-phenylbutyrate acid (4-PBA, a chemical chaperone, alleviates aggregate formation and partially rescued the protein secretion of SP-A2 mutants. In conclusion, SP-A2 G231V and F198S mutants impair the dimmer/trimer assembly, which contributes to the protein sialylation and secretion deficiency. The intracellular protein mutants could be partially degraded through the proteasome pathway and also formed aggregates. The treatment of the cells with 4-PBA resulted in reduced aggregation and rescued the secretion of mutant SP-A2.

  13. Pseudovirion particles bearing native HIV envelope trimers facilitate a novel method for generating human neutralizing monoclonal antibodies against HIV.

    Science.gov (United States)

    Hicar, Mark D; Chen, Xuemin; Briney, Bryan; Hammonds, Jason; Wang, Jaang-Jiun; Kalams, Spyros; Spearman, Paul W; Crowe, James E

    2010-07-01

    Monomeric HIV envelope vaccines fail to elicit broadly neutralizing antibodies or to protect against infection. Neutralizing antibodies against HIV bind to native functionally active Env trimers on the virion surface. Gag-Env pseudovirions recapitulate the native trimer and could serve as an effective epitope presentation platform for study of the neutralizing antibody response in HIV-infected individuals. To address if pseudovirions can recapitulate native HIV virion epitope structures, we carefully characterized these particles, concentrating on the antigenic structure of the coreceptor binding site. By blue native gel shift assays, Gag-Env pseudovirions were shown to contain native trimers that were competent for binding to neutralizing monoclonal antibodies. In enzyme-linked immunosorbent assay, pseudovirions exhibited increased binding of known CD4-induced antibodies after addition of CD4. Using flow cytometric analysis, fluorescently labeled pseudovirions specifically identified a subset of antigen-specific B cells in HIV-infected subjects. Interestingly, the sequence of one of these novel human antibodies, identified during cloning of single HIV-specific B cells and designated 2C6, exhibited homology to mAb 47e, a known anti-CD4-induced coreceptor binding site antibody. The secreted monoclonal antibody 2C6 did not bind monomeric gp120, but specifically bound envelope on pseudovirions. A recombinant form of the antibody 2C6 acted as a CD4-induced epitope-specific antibody in neutralization assays, yet did not bind monomeric gp120. These findings imply specificity against a quaternary epitope presented on the pseudovirion envelope spike. These data demonstrate that Gag-Env pseudovirions recapitulate CD4 and coreceptor binding pocket antigenic structures and can facilitate identification of B-cell clones that secrete neutralizing antibodies.

  14. Anisotropic growth of a nickel trimer formed on a highly-stepped TiO2(110) surface

    OpenAIRE

    Uehara, Hiromitsu; Bin Hanaffi, Muhammad Haneef; Koike, Yuichiro; Fujikawa, Keisuke; Suzuki, Shushi; Ariga, Hiroko; Takakusagi, Satoru; Chun, Wang-Jae; Iwasawa, Yasuhiro; Asakura, Kiyotaka

    2013-01-01

    The structures of Ni clusters formed on a highly-stepped TiO2(110) surface were studied by polarization-dependent total reflection fluorescence X-ray absorption fine structure analysis. When 0.8 monolayers of Ni were deposited, three-dimensional Ni clusters with 1-2 nm diameters and heights less than 1 nm were formed. Conversely, when 0.07 monolayers of Ni were deposited, an anisotropic Ni trimer with a Ni-Ni distance of 0.260 nm was created at the [001] step. We revealed that the surface mod...

  15. Changes in Structure and Antigenicity of HIV-1 Env Trimers Resulting from Removal of a Conserved CD4 Binding Site-Proximal Glycan

    Science.gov (United States)

    Liang, Yu; Guttman, Miklos; Williams, James A.; Verkerke, Hans; Alvarado, Daniel

    2016-01-01

    ABSTRACT The envelope glycoprotein (Env) is the major target for HIV-1 broadly neutralizing antibodies (bNAbs). One of the mechanisms that HIV has evolved to escape the host's immune response is to mask conserved epitopes on Env with dense glycosylation. Previous studies have shown that the removal of a particular conserved glycan at N197 increases the neutralization sensitivity of the virus to antibodies targeting the CD4 binding site (CD4bs), making it a site of significant interest from the perspective of vaccine design. At present, the structural consequences that result from the removal of the N197 glycan have not been characterized. Using native-like SOSIP trimers, we examine the effects on antigenicity and local structural dynamics resulting from the removal of this glycan. A large increase in the binding of CD4bs and V3-targeting antibodies is observed for the N197Q mutant in trimeric Env, while no changes are observed with monomeric gp120. While the overall structure and thermostability are not altered, a subtle increase in the flexibility of the variable loops at the trimeric interface of adjacent protomers is evident in the N197Q mutant by hydrogen-deuterium exchange mass spectrometry. Structural modeling of the glycan chains suggests that the spatial occupancy of the N197 glycan leads to steric clashes with CD4bs antibodies in the Env trimer but not monomeric gp120. Our results indicate that the removal of the N197 glycan enhances the exposure of relevant bNAb epitopes on Env with a minimal impact on the overall trimeric structure. These findings present a simple modification for enhancing trimeric Env immunogens in vaccines. IMPORTANCE The HIV-1 Env glycoprotein presents a dense patchwork of host cell-derived N-linked glycans. This so-called glycan shield is considered to be a major protective mechanism against immune recognition. While the positions of many N-linked glycans are isolate specific, some are highly conserved and are believed to play key

  16. Changes in Structure and Antigenicity of HIV-1 Env Trimers Resulting from Removal of a Conserved CD4 Binding Site-Proximal Glycan.

    Science.gov (United States)

    Liang, Yu; Guttman, Miklos; Williams, James A; Verkerke, Hans; Alvarado, Daniel; Hu, Shiu-Lok; Lee, Kelly K

    2016-10-15

    The envelope glycoprotein (Env) is the major target for HIV-1 broadly neutralizing antibodies (bNAbs). One of the mechanisms that HIV has evolved to escape the host's immune response is to mask conserved epitopes on Env with dense glycosylation. Previous studies have shown that the removal of a particular conserved glycan at N197 increases the neutralization sensitivity of the virus to antibodies targeting the CD4 binding site (CD4bs), making it a site of significant interest from the perspective of vaccine design. At present, the structural consequences that result from the removal of the N197 glycan have not been characterized. Using native-like SOSIP trimers, we examine the effects on antigenicity and local structural dynamics resulting from the removal of this glycan. A large increase in the binding of CD4bs and V3-targeting antibodies is observed for the N197Q mutant in trimeric Env, while no changes are observed with monomeric gp120. While the overall structure and thermostability are not altered, a subtle increase in the flexibility of the variable loops at the trimeric interface of adjacent protomers is evident in the N197Q mutant by hydrogen-deuterium exchange mass spectrometry. Structural modeling of the glycan chains suggests that the spatial occupancy of the N197 glycan leads to steric clashes with CD4bs antibodies in the Env trimer but not monomeric gp120. Our results indicate that the removal of the N197 glycan enhances the exposure of relevant bNAb epitopes on Env with a minimal impact on the overall trimeric structure. These findings present a simple modification for enhancing trimeric Env immunogens in vaccines. The HIV-1 Env glycoprotein presents a dense patchwork of host cell-derived N-linked glycans. This so-called glycan shield is considered to be a major protective mechanism against immune recognition. While the positions of many N-linked glycans are isolate specific, some are highly conserved and are believed to play key functional roles. In

  17. Honokiol trimers and dimers via biotransformation catalyzed by Momordica charantia peroxidase: novel and potent α-glucosidase inhibitors.

    Science.gov (United States)

    He, Ye; Wang, Xiao-Bing; Fan, Bo-Yi; Kong, Ling-Yi

    2014-01-15

    Ten honokiol oligomers (1-10), including four novel trimers (1-4) and four novel dimers (5-8), were obtained by means of biotransformation of honokiol catalyzed by Momordica charantia peroxidase (MCP) for the first time. Their structures were established on the basis of spectroscopic methods. The biological results demonstrated that most of the oligomers were capable of inhibiting α-glucosidase with significant abilities, which were one to two orders of magnitude more potent than the substrate, honokiol. In particular, compound 2, the honokiol trimer, displayed the greatest inhibitory activity against α-glucosidase with an IC50 value of 1.38μM. Kinetic and CD studies indicated that 2 inhibited α-glucosidase in a reversible, mixed-type manner and caused conformational changes in the secondary structure of the enzyme protein. These findings suggested that 2 might be exploited as a promising drug candidate for the treatment of diabetes. Copyright © 2013 Elsevier Ltd. All rights reserved.

  18. Modulation of Interparticle Distance in Discrete Gold Nanoparticle Dimers and Trimers by DNA Single-Base Pairing.

    Science.gov (United States)

    Akiyama, Yoshitsugu; Shikagawa, Hiroto; Kanayama, Naoki; Takarada, Tohru; Maeda, Mizuo

    2015-07-01

    Self-assembled structures of metallic nanoparticles with dynamically changeable interparticle distance hold promise for the regulation of collective physical properties. This paper describes gold nanoparticle dimers and trimers that exhibit spontaneous and reversible changes in interparticle distance. To exploit this property, a gold nanoparticle is modified with precisely one long DNA strand and approximately five short DNA strands. The long DNA serves to align the nanoparticles on a template DNA via hybridization, while the short DNAs function to induce the interparticle distance changes. The obtained dimer and trimer are characterized with gel electrophoresis, dynamic light scattering measurements, and transmission electron microscopy (TEM). When the complementary short DNA is added to form the fully matched duplexes on the particle surface in the presence of MgCl2 , spontaneous reduction of the interparticle distance is observed with TEM and cryo-electron microscopy. By contrast, when the terminal-mismatched DNA is added, no structural change occurs under the same conditions. Therefore, the single base pairing/unpairing at the outermost surface of the nanoparticle impacts the interparticle distance. This unique feature could be applied to the regulation of structures and properties of various DNA-functionalized nanoparticle assemblies. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  19. Direct Probing of Germinal Center Responses Reveals Immunological Features and Bottlenecks for Neutralizing Antibody Responses to HIV Env Trimer.

    Science.gov (United States)

    Havenar-Daughton, Colin; Carnathan, Diane G; Torrents de la Peña, Alba; Pauthner, Matthias; Briney, Bryan; Reiss, Samantha M; Wood, Jennifer S; Kaushik, Kirti; van Gils, Marit J; Rosales, Sandy L; van der Woude, Patricia; Locci, Michela; Le, Khoa M; de Taeye, Steven W; Sok, Devin; Mohammed, Ata Ur Rasheed; Huang, Jessica; Gumber, Sanjeev; Garcia, AnaPatricia; Kasturi, Sudhir P; Pulendran, Bali; Moore, John P; Ahmed, Rafi; Seumois, Grégory; Burton, Dennis R; Sanders, Rogier W; Silvestri, Guido; Crotty, Shane

    2016-11-22

    Generating tier 2 HIV-neutralizing antibody (nAb) responses by immunization remains a challenging problem, and the immunological barriers to induction of such responses with Env immunogens remain unclear. Here, some rhesus monkeys developed autologous tier 2 nAbs upon HIV Env trimer immunization (SOSIP.v5.2) whereas others did not. This was not because HIV Env trimers were immunologically silent because all monkeys made similar ELISA-binding antibody responses; the key difference was nAb versus non-nAb responses. We explored the immunological barriers to HIV nAb responses by combining a suite of techniques, including longitudinal lymph node fine needle aspirates. Unexpectedly, nAb development best correlated with booster immunization GC B cell magnitude and Tfh characteristics of the Env-specific CD4 T cells. Notably, these factors distinguished between successful and unsuccessful antibody responses because GC B cell frequencies and stoichiometry to GC Tfh cells correlated with nAb development, but did not correlate with total Env Ab binding titers. Copyright © 2016 The Author(s). Published by Elsevier Inc. All rights reserved.

  20. The Haemophilus ducreyi trimeric autotransporter adhesin DsrA protects against an experimental infection in the swine model of chancroid.

    Science.gov (United States)

    Fusco, William G; Choudhary, Neelima R; Routh, Patty A; Ventevogel, Melissa S; Smith, Valerie A; Koch, Gary G; Almond, Glen W; Orndorff, Paul E; Sempowski, Gregory D; Leduc, Isabelle

    2014-06-24

    Adherence of pathogens to cellular targets is required to initiate most infections. Defining strategies that interfere with adhesion is therefore important for the development of preventative measures against infectious diseases. As an adhesin to host extracellular matrix proteins and human keratinocytes, the trimeric autotransporter adhesin DsrA, a proven virulence factor of the Gram-negative bacterium Haemophilus ducreyi, is a potential target for vaccine development. A recombinant form of the N-terminal passenger domain of DsrA from H. ducreyi class I strain 35000HP, termed rNT-DsrAI, was tested as a vaccine immunogen in the experimental swine model of H. ducreyi infection. Viable homologous H. ducreyi was not recovered from any animal receiving four doses of rNT-DsrAI administered with Freund's adjuvant at two-week intervals. Control pigs receiving adjuvant only were all infected. All animals receiving the rNT-DsrAI vaccine developed antibody endpoint titers between 3.5 and 5 logs. All rNT-DsrAI antisera bound the surface of the two H. ducreyi strains used to challenge immunized pigs. Purified anti-rNT-DsrAI IgG partially blocked binding of fibrinogen at the surface of viable H. ducreyi. Overall, immunization with the passenger domain of the trimeric autotransporter adhesin DsrA accelerated clearance of H. ducreyi in experimental lesions, possibly by interfering with fibrinogen binding. Copyright © 2014 Elsevier Ltd. All rights reserved.

  1. Dimeric and trimeric fusion proteins generated with fimbrial adhesins of uropathogenic Escherichia coli

    Directory of Open Access Journals (Sweden)

    Víctor Manuel Luna-Pineda

    2016-10-01

    Full Text Available Urinary tract infections (UTIs are associated with high rates of morbidity and mortality worldwide, and uropathogenic Escherichia coli (UPEC is the main etiologic agent. Fimbriae assembled on the bacterial surface are essential for adhesion in the urinary tract epithelium. In this study, the FimH, CsgA, and PapG adhesins were fused to generate biomolecules as potential target vaccines against UTIs. The fusion protein design was generated using bioinformatics tools, and template fusion gene sequences were synthesized by GenScript in the following order fimH-csgA-papG-fimH-csgA (fcpfc linked to the nucleotide sequence encoding the EAAAK5 peptide. Monomeric (fimH, csgA, and papG, dimeric (fimH-csgA, and trimeric (fimH-csgA-papG genes were cloned into the pLATE31 expression vector and generated products of 1040, 539, 1139, 1442, and 2444 bp, respectively. Fusion protein expression in BL21 E. coli was induced with 1 mM IPTG, and His-tagged fusion proteins were purified under denaturing conditions and refolded by dialysis using C-buffer. Coomassie blue-stained SDS-PAGE gels and Western blot analysis revealed bands of 29.5, 11.9, 33.9, 44.9, and 82.1 kDa corresponding to FimH, CsgA, PapG, FC, and FCP proteins, respectively. Mass spectrometry analysis by MALDI-TOF/TOF revealed specific peptides that confirmed the fusion protein structures. Dynamic light scattering analysis revealed the polydispersed state of the fusion proteins. The FimH, CsgA, and PapG stimulated the release of 372 to 398 pg/mL IL-6; interestingly, the FC and FCP stimulated the release of 464.79 pg/mL (p ≤ 0.018 and 521.24 pg/mL (p ≤ 0.002 IL-6, respectively. In addition, the FC and FCP stimulated the release of 398.52 pg/mL (p ≤ 0.001 and 450.40 pg/mL (p ≤ 0.002 IL-8, respectively. High levels of IgA and IgG antibodies in human sera reacted against the fusion proteins, and under identical conditions, low levels of IgA and IgG antibodies were detected in human urine. Rabbit

  2. Dimeric and Trimeric Fusion Proteins Generated with Fimbrial Adhesins of UropathogenicEscherichia coli.

    Science.gov (United States)

    Luna-Pineda, Víctor M; Reyes-Grajeda, Juan Pablo; Cruz-Córdova, Ariadnna; Saldaña-Ahuactzi, Zeus; Ochoa, Sara A; Maldonado-Bernal, Carmen; Cázares-Domínguez, Vicenta; Moreno-Fierros, Leticia; Arellano-Galindo, José; Hernández-Castro, Rigoberto; Xicohtencatl-Cortes, Juan

    2016-01-01

    Urinary tract infections (UTIs) are associated with high rates of morbidity and mortality worldwide, and uropathogenic Escherichia coli (UPEC) is the main etiologic agent. Fimbriae assembled on the bacterial surface are essential for adhesion to the urinary tract epithelium. In this study, the FimH, CsgA, and PapG adhesins were fused to generate biomolecules for use as potential target vaccines against UTIs. The fusion protein design was generated using bioinformatics tools, and template fusion gene sequences were synthesized by GenScript in the following order fimH - csgA - papG - fimH - csgA ( fcpfc ) linked to the nucleotide sequence encoding the [EAAAK] 5 peptide. Monomeric ( fimH, csgA , and papG ), dimeric ( fimH-csgA ), and trimeric ( fimH - csgA - papG ) genes were cloned into the pLATE31 expression vector and generated products of 1040, 539, 1139, 1442, and 2444 bp, respectively. Fusion protein expression in BL21 E. coli was induced with 1 mM IPTG, and His-tagged proteins were purified under denaturing conditions and refolded by dialysis using C-buffer. Coomassie blue-stained SDS-PAGE gels and Western blot analysis revealed bands of 29.5, 11.9, 33.9, 44.9, and 82.1 kDa, corresponding to FimH, CsgA, PapG, FC, and FCP proteins, respectively. Mass spectrometry analysis by MALDI-TOF/TOF revealed specific peptides that confirmed the fusion protein structures. Dynamic light scattering analysis revealed the polydispersed state of the fusion proteins. FimH, CsgA, and PapG stimulated the release of 372-398 pg/mL IL-6; interestingly, FC and FCP stimulated the release of 464.79 pg/mL ( p ≤ 0.018) and 521.24 pg/mL ( p ≤ 0.002) IL-6, respectively. In addition, FC and FCP stimulated the release of 398.52 pg/mL ( p ≤ 0.001) and 450.40 pg/mL ( p ≤ 0.002) IL-8, respectively. High levels of IgA and IgG antibodies in human sera reacted against the fusion proteins, and under identical conditions, low levels of IgA and IgG antibodies were detected in human urine

  3. Amyloid beta dimers/trimers potently induce cofilin-actin rods that are inhibited by maintaining cofilin-phosphorylation

    Directory of Open Access Journals (Sweden)

    Podlisny Marcia

    2011-01-01

    Full Text Available Abstract Background Previously we reported 1 μM synthetic human amyloid beta1-42 oligomers induced cofilin dephosphorylation (activation and formation of cofilin-actin rods within rat hippocampal neurons primarily localized to the dentate gyrus. Results Here we demonstrate that a gel filtration fraction of 7PA2 cell-secreted SDS-stable human Aβ dimers and trimers (Aβd/t induces maximal neuronal rod response at ~250 pM. This is 4,000-fold more active than traditionally prepared human Aβ oligomers, which contain SDS-stable trimers and tetramers, but are devoid of dimers. When incubated under tyrosine oxidizing conditions, synthetic human but not rodent Aβ1-42, the latter lacking tyrosine, acquires a marked increase (620 fold for EC50 in rod-inducing activity. Gel filtration of this preparation yielded two fractions containing SDS-stable dimers, trimers and tetramers. One, eluting at a similar volume to 7PA2 Aβd/t, had maximum activity at ~5 nM, whereas the other, eluting at the void volume (high-n state, lacked rod inducing activity at the same concentration. Fractions from 7PA2 medium containing Aβ monomers are not active, suggesting oxidized SDS-stable Aβ1-42 dimers in a low-n state are the most active rod-inducing species. Aβd/t-induced rods are predominantly localized to the dentate gyrus and mossy fiber tract, reach significance over controls within 2 h of treatment, and are reversible, disappearing by 24 h after Aβd/t washout. Overexpression of cofilin phosphatases increase rod formation when expressed alone and exacerbate rod formation when coupled with Aβd/t, whereas overexpression of a cofilin kinase inhibits Aβd/t-induced rod formation. Conclusions Together these data support a mechanism by which Aβd/t alters the actin cytoskeleton via effects on cofilin in neurons critical to learning and memory.

  4. Modeling of the N-terminal Section and the Lumenal Loop of Trimeric Light Harvesting Complex II (LHCII) by Using EPR.

    Science.gov (United States)

    Fehr, Niklas; Dietz, Carsten; Polyhach, Yevhen; von Hagens, Tona; Jeschke, Gunnar; Paulsen, Harald

    2015-10-23

    The major light harvesting complex II (LHCII) of green plants plays a key role in the absorption of sunlight, the regulation of photosynthesis, and in preventing photodamage by excess light. The latter two functions are thought to involve the lumenal loop and the N-terminal domain. Their structure and mobility in an aqueous environment are only partially known. Electron paramagnetic resonance (EPR) has been used to measure the structure of these hydrophilic protein domains in detergent-solubilized LHCII. A new technique is introduced to prepare LHCII trimers in which only one monomer is spin-labeled. These heterogeneous trimers allow to measure intra-molecular distances within one LHCII monomer in the context of a trimer by using double electron-electron resonance (DEER). These data together with data from electron spin echo envelope modulation (ESEEM) allowed to model the N-terminal protein section, which has not been resolved in current crystal structures, and the lumenal loop domain. The N-terminal domain covers only a restricted area above the superhelix in LHCII, which is consistent with the "Velcro" hypothesis to explain thylakoid grana stacking (Standfuss, J., van Terwisscha Scheltinga, A. C., Lamborghini, M., and Kühlbrandt, W. (2005) EMBO J. 24, 919-928). The conformation of the lumenal loop domain is surprisingly different between LHCII monomers and trimers but not between complexes with and without neoxanthin bound. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  5. Modeling of the N-terminal Section and the Lumenal Loop of Trimeric Light Harvesting Complex II (LHCII) by Using EPR*

    Science.gov (United States)

    Fehr, Niklas; Dietz, Carsten; Polyhach, Yevhen; von Hagens, Tona; Jeschke, Gunnar; Paulsen, Harald

    2015-01-01

    The major light harvesting complex II (LHCII) of green plants plays a key role in the absorption of sunlight, the regulation of photosynthesis, and in preventing photodamage by excess light. The latter two functions are thought to involve the lumenal loop and the N-terminal domain. Their structure and mobility in an aqueous environment are only partially known. Electron paramagnetic resonance (EPR) has been used to measure the structure of these hydrophilic protein domains in detergent-solubilized LHCII. A new technique is introduced to prepare LHCII trimers in which only one monomer is spin-labeled. These heterogeneous trimers allow to measure intra-molecular distances within one LHCII monomer in the context of a trimer by using double electron-electron resonance (DEER). These data together with data from electron spin echo envelope modulation (ESEEM) allowed to model the N-terminal protein section, which has not been resolved in current crystal structures, and the lumenal loop domain. The N-terminal domain covers only a restricted area above the superhelix in LHCII, which is consistent with the “Velcro” hypothesis to explain thylakoid grana stacking (Standfuss, J., van Terwisscha Scheltinga, A. C., Lamborghini, M., and Kühlbrandt, W. (2005) EMBO J. 24, 919–928). The conformation of the lumenal loop domain is surprisingly different between LHCII monomers and trimers but not between complexes with and without neoxanthin bound. PMID:26316535

  6. Lateral and vertical manipulations of single atoms on the Ag(1 1 1) surface with the copper single-atom and trimer-apex tips

    International Nuclear Information System (INIS)

    Xie Yiqun; Yang Tianxing; Ye Xiang; Huang Lei

    2011-01-01

    We study the lateral and vertical manipulations of single Ag and Cu atoms on the Ag(1 1 1) surface with the Cu single-atom and trimer-apex tips using molecular statics simulations. The reliability of the lateral manipulation with the Cu single-atom tip is investigated, and compared with that for the Ag tips. We find that overall the manipulation reliability (MR) increases with the decreasing tip height, and in a wide tip-height range the MR is better than those for both the Ag single-atom and trimer-apex tips. This is due to the stronger attractive force of the Cu tip and its better stability against the interactions with the Ag surface. With the Cu trimer-apex tip, the single Ag and Cu adatoms can be picked up from the flat Ag(1 1 1) surface, and moreover a reversible vertical manipulation of single Ag atoms on the stepped Ag(1 1 1) surface is possible, suggesting a method to modify two-dimensional Ag nanostructures on the Ag(1 1 1) surface with the Cu trimer-apex tip.

  7. A Broadly Neutralizing Antibody Targets the Dynamic HIV Envelope Trimer Apex via a Long, Rigidified, and Anionic β-Hairpin Structure.

    Science.gov (United States)

    Lee, Jeong Hyun; Andrabi, Raiees; Su, Ching-Yao; Yasmeen, Anila; Julien, Jean-Philippe; Kong, Leopold; Wu, Nicholas C; McBride, Ryan; Sok, Devin; Pauthner, Matthias; Cottrell, Christopher A; Nieusma, Travis; Blattner, Claudia; Paulson, James C; Klasse, Per Johan; Wilson, Ian A; Burton, Dennis R; Ward, Andrew B

    2017-04-18

    Broadly neutralizing antibodies (bnAbs) to HIV delineate vaccine targets and are prophylactic and therapeutic agents. Some of the most potent bnAbs target a quaternary epitope at the apex of the surface HIV envelope (Env) trimer. Using cryo-electron microscopy, we solved the atomic structure of an apex bnAb, PGT145, in complex with Env. We showed that the long anionic HCDR3 of PGT145 penetrated between glycans at the trimer 3-fold axis, to contact peptide residues from all three Env protomers, and thus explains its highly trimer-specific nature. Somatic hypermutation in the other CDRs of PGT145 were crucially involved in stabilizing the structure of the HCDR3, similar to bovine antibodies, to aid in recognition of a cluster of conserved basic residues hypothesized to facilitate trimer disassembly during viral entry. Overall, the findings exemplify the creative solutions that the human immune system can evolve to recognize a conserved motif buried under a canopy of glycans. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

  8. Characterization of a stable HIV-1 B/C recombinant, soluble, and trimeric envelope glycoprotein (Env) highly resistant to CD4-induced conformational changes.

    Science.gov (United States)

    Kumar, Rajesh; Ozorowski, Gabriel; Kumar, Vivek; Holden, Lauren G; Shrivastava, Tripti; Patil, Shilpa; Deshpande, Suprit; Ward, Andrew B; Bhattacharya, Jayanta

    2017-09-22

    The HIV-1 envelope (Env) is a glycoprotein consisting of a trimer of heterodimers containing gp120 and gp41 subunits that mediates virus entry and is a major target of broadly neutralizing antibodies (bnAbs) developed during infection in some individuals. The engagement of the HIV-1 gp120 glycoprotein to the host CD4 protein triggers conformational changes in gp120 that allow its binding to co-receptors and is necessary for virus entry to establish infection. Native-like HIV-1 Env immunogens representing distinct clades have been proposed to improve immunogenicity. In the present study, we examined the basis of resistance of an HIV-1 B/C recombinant Env (LT5.J4b12C) to non-neutralizing antibodies targeting CD4-induced Env epitopes in the presence of soluble CD4 (sCD4). Using native polyacrylamide gel shift assay and negative-stain EM, we found that the prefusion conformational state of LT5.J4b12C trimeric Env was largely unaffected in the presence of excess sCD4 with most Env trimers appearing to be in a ligand-free state. This resistance to CD4-induced conformational changes was associated with a lower affinity for CD4. Moreover, the LT5.J4b12C trimeric Env preferentially bound to the neutralizing antibodies compared with non-neutralizing antibodies. Taken together, we report on an HIV-1 B/C recombinant, native-like trimeric Env protein that is highly resistant to CD4-induced conformational changes but displays epitopes recognized by a diverse array of bnAbs. Such features make this B/C recombinant trimeric Env a useful addition to the pool of other recently identified native-like HIV-1 Env trimers suitable for use as antigenic bait for bnAb isolation, structural studies, and use as potential immunogens. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  9. Full-length cellular β-secretase has a trimeric subunit stoichiometry, and its sulfur-rich transmembrane interaction site modulates cytosolic copper compartmentalization.

    Science.gov (United States)

    Liebsch, Filip; Aurousseau, Mark R P; Bethge, Tobias; McGuire, Hugo; Scolari, Silvia; Herrmann, Andreas; Blunck, Rikard; Bowie, Derek; Multhaup, Gerd

    2017-08-11

    The β-secretase (BACE1) initiates processing of the amyloid precursor protein (APP) into Aβ peptides, which have been implicated as central players in the pathology of Alzheimer disease. BACE1 has been described as a copper-binding protein and its oligomeric state as being monomeric, dimeric, and/or multimeric, but the native cellular stoichiometry has remained elusive. Here, by using single-molecule fluorescence and in vitro cross-linking experiments with photo-activatable unnatural amino acids, we show that full-length BACE1, independently of its subcellular localization, exists as trimers in human cells. We found that trimerization requires the BACE1 transmembrane sequences (TMSs) and cytoplasmic domains, with residues Ala 463 and Cys 466 buried within the trimer interface of the sulfur-rich core of the TMSs. Our 3D model predicts that the sulfur-rich core of the trimeric BACE1 TMS is accessible to metal ions, but copper ions did not trigger trimerization. The results of functional assays of endogenous BACE1 suggest that it has a role in intracellular copper compartmentalization by transferring cytosolic copper to intracellular compartments, while leaving the overall cellular copper concentration unaltered. Adding to existing physiological models, our results provide novel insight into the atypical interactions between copper and BACE1 and into its non-enzymatic activities. In conclusion, therapeutic Alzheimer disease prevention strategies aimed at decreasing BACE1 protein levels should be regarded with caution, because adverse effects in copper homeostasis may occur. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  10. Emergent spin-1 trimerized valence bond crystal in the spin-1/2 Heisenberg model on the star lattice

    Science.gov (United States)

    Ran, Shi-Ju; Li, Wei; Gong, Shou-Shu; Weichselbaum, Andreas; von Delft, Jan; Su, Gang

    2018-02-01

    We explore the frustrated spin-1/2 Heisenberg model on the star lattice with antiferromagnetic (AF) couplings inside each triangle and ferromagnetic (FM) intertriangle couplings (Jefrustration from the AF interactions inside each triangle, but trigger a fully gapped inversion-symmetry-breaking trimerized valence bond crystal (TVBC) with emergent spin-1 degrees of freedom. We discover that with strengthening Je, the system exhibits a universal scaling behavior either with or without a magnetic field h : the order parameter, the five critical fields that separate the Je-h ground-state phase diagram into six phases, and the excitation gap obtained by low-temperature specific heat, all depend exponentially on Je. Our work implies that the spin-1 VBCs can be stabilized by introducing small FM couplings in the geometrically frustrated spin-1/2 systems.

  11. Functionalization of nanoparticle titanium dioxide with different bifunctional organic molecules and trimers of transition compounds for obtaining new materials

    International Nuclear Information System (INIS)

    Rivera Martinez, Maria Cinthya

    2012-01-01

    Functionalization of titanium dioxide in nanoporous anatase phase is investigated for obtaining new nanomaterials. Functionalizations were performed using two heating methods: the conventional of refluxing heating method and microwave irradiation with bifunctional organic molecules is used to study how to anchor molecules and the change in the wettability of the material. Besides, reactions with organic molecules were performed as the derived from nanoproxene. The growth layer by layer is performed using the bifunctional molecules previous for the immobilization of cobalt trimers. Functionalized molecules were characterized by infrared spectroscopy, X-ray diffraction, contact angle, scanning electron microscopy, x-ray elemental analysis, plasma atomic emission spectroscopy coupled inductively, x-ray photoelectron spectroscopy and thermogravimetric analysis. This type of functionalizations on nanoporous titanium dioxide could potentially improve optical sensitivity and activity of this nanomaterial in the visible region. (author) [es

  12. Crystal structure of tetranectin, a trimeric plasminogen-binding protein with an alpha-helical coiled coil

    DEFF Research Database (Denmark)

    Nielsen, B B; Kastrup, J S; Rasmussen, H

    1997-01-01

    Tetranectin is a plasminogen kringle 4-binding protein. The crystal structure has been determined at 2.8 A resolution using molecular replacement. Human tetranectin is a homotrimer forming a triple alpha-helical coiled coil. Each monomer consists of a carbohydrate recognition domain (CRD) connected...... the third is present only in long-form CRDs. Tetranectin represents the first structure of a long-form CRD with intact calcium-binding sites. In tetranectin, the third disulfide bridge tethers the CRD to the long helix in the coiled coil. The trimerization of tetranectin as well as the fixation of the CRDs...... relative to the helices in the coiled coil indicate a demand for high specificity in the recognition and binding of ligands....

  13. The Arabidopsis Cellulose Synthase Complex: A Proposed Hexamer of CESA Trimers in an Equimolar Stoichiometry

    Energy Technology Data Exchange (ETDEWEB)

    Hill, Joseph L. [Pennsylvania State Univ., University Park, PA (United States); Hammudi, Mustafa B. [Pennsylvania State Univ., University Park, PA (United States); Tien, Ming [Pennsylvania State Univ., University Park, PA (United States)

    2014-12-01

    In this study, we show a 1:1:1 stoichiometry between the three Arabidopsis thaliana secondary cell wall isozymes: CESA4, CESA7, and CESA8. This ratio was determined utilizing a simple but elegant method of quantitative immunoblotting using isoform-specific antibodies and 35S-labeled protein standards for each CESA. Additionally, the observed equimolar stoichiometry was found to be fixed along the axis of the stem, which represents a developmental gradient. Our results complement recent spectroscopic analyses pointing toward an 18-chain cellulose microfibril. Taken together, we propose that the CSC is composed of a hexamer of catalytically active CESA trimers, with each CESA in equimolar amounts. This finding is a crucial advance in understanding how CESAs integrate to form higher order complexes, which is a key determinate of cellulose microfibril and cell wall properties.

  14. A trimeric structural fusion of an antagonistic tumor necrosis factor-α mutant enhances molecular stability and enables facile modification.

    Science.gov (United States)

    Inoue, Masaki; Ando, Daisuke; Kamada, Haruhiko; Taki, Shintaro; Niiyama, Mayumi; Mukai, Yohei; Tadokoro, Takashi; Maenaka, Katsumi; Nakayama, Taisuke; Kado, Yuji; Inoue, Tsuyoshi; Tsutsumi, Yasuo; Tsunoda, Shin-Ichi

    2017-04-21

    Tumor necrosis factor-α (TNF) exerts its biological effect through two types of receptors, p55 TNF receptor (TNFR1) and p75 TNF receptor (TNFR2). An inflammatory response is known to be induced mainly by TNFR1, whereas an anti-inflammatory reaction is thought to be mediated by TNFR2 in some autoimmune diseases. We have been investigating the use of an antagonistic TNF mutant (TNFR1-selective antagonistic TNF mutant (R1antTNF)) to reveal the pharmacological effect of TNFR1-selective inhibition as a new therapeutic modality. Here, we aimed to further improve and optimize the activity and behavior of this mutant protein both in vitro and in vivo Specifically, we examined a trimeric structural fusion of R1antTNF, formed via the introduction of short peptide linkers, as a strategy to enhance bioactivity and molecular stability. By comparative analysis with R1antTNF, the trimeric fusion, referred to as single-chain R1antTNF (scR1antTNF), was found to retain in vitro molecular properties of receptor selectivity and antagonistic activity but displayed a marked increase in thermal stability. The residence time of scR1antTNF in vivo was also significantly prolonged. Furthermore, molecular modification using polyethylene glycol (PEG) was easily controlled by limiting the number of reactive sites. Taken together, our findings show that scR1antTNF displays enhanced molecular stability while maintaining biological activity compared with R1antTNF. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  15. Structural basis for the oligomerization-state switch from a dimer to a trimer of an engineered cortexillin-1 coiled-coil variant.

    Directory of Open Access Journals (Sweden)

    Saša Bjelić

    Full Text Available Coiled coils are well suited to drive subunit oligomerization and are widely used in applications ranging from basic research to medicine. The optimization of these applications requires a detailed understanding of the molecular determinants that control of coiled-coil formation. Although many of these determinants have been identified and characterized in great detail, a puzzling observation is that their presence does not necessarily correlate with the oligomerization state of a given coiled-coil structure. Thus, other determinants must play a key role. To address this issue, we recently investigated the unrelated coiled-coil domains from GCN4, ATF1 and cortexillin-1 as model systems. We found that well-known trimer-specific oligomerization-state determinants, such as the distribution of isoleucine residues at heptad-repeat core positions or the trimerization motif Arg-h-x-x-h-Glu (where h = hydrophobic amino acid; x = any amino acid, switch the peptide's topology from a dimer to a trimer only when inserted into the trigger sequence, a site indispensable for coiled-coil formation. Because high-resolution structural information could not be obtained for the full-length, three-stranded cortexillin-1 coiled coil, we here report the detailed biophysical and structural characterization of a shorter variant spanning the trigger sequence using circular dichroism, anatytical ultracentrifugation and x-ray crystallography. We show that the peptide forms a stable α-helical trimer in solution. We further determined the crystal structure of an optimised variant at a resolution of 1.65 Å, revealing that the peptide folds into a parallel, three-stranded coiled coil. The two complemented R-IxxIE trimerization motifs and the additional hydrophobic core isoleucine residue adopt the conformations seen in other extensively characterized parallel, three-stranded coiled coils. These findings not only confirm the structural basis for the switch in

  16. Targeted N-glycan deletion at the receptor-binding site retains HIV Env NFL trimer integrity and accelerates the elicited antibody response.

    Science.gov (United States)

    Dubrovskaya, Viktoriya; Guenaga, Javier; de Val, Natalia; Wilson, Richard; Feng, Yu; Movsesyan, Arlette; Karlsson Hedestam, Gunilla B; Ward, Andrew B; Wyatt, Richard T

    2017-09-01

    Extensive shielding by N-glycans on the surface of the HIV envelope glycoproteins (Env) restricts B cell recognition of conserved neutralizing determinants. Elicitation of broadly neutralizing antibodies (bNAbs) in selected HIV-infected individuals reveals that Abs capable of penetrating the glycan shield can be generated by the B cell repertoire. Accordingly, we sought to determine if targeted N-glycan deletion might alter antibody responses to Env. We focused on the conserved CD4 binding site (CD4bs) since this is a known neutralizing determinant that is devoid of glycosylation to allow CD4 receptor engagement, but is ringed by surrounding N-glycans. We selectively deleted potential N-glycan sites (PNGS) proximal to the CD4bs on well-ordered clade C 16055 native flexibly linked (NFL) trimers to potentially increase recognition by naïve B cells in vivo. We generated glycan-deleted trimer variants that maintained native-like conformation and stability. Using a panel of CD4bs-directed bNAbs, we demonstrated improved accessibility of the CD4bs on the N-glycan-deleted trimer variants. We showed that pseudoviruses lacking these Env PNGSs were more sensitive to neutralization by CD4bs-specific bNAbs but remained resistant to non-neutralizing mAbs. We performed rabbit immunogenicity experiments using two approaches comparing glycan-deleted to fully glycosylated NFL trimers. The first was to delete 4 PNGS sites and then boost with fully glycosylated Env; the second was to delete 4 sites and gradually re-introduce these N-glycans in subsequent boosts. We demonstrated that the 16055 PNGS-deleted trimers more rapidly elicited serum antibodies that more potently neutralized the CD4bs-proximal-PNGS-deleted viruses in a statistically significant manner and strongly trended towards increased neutralization of fully glycosylated autologous virus. This approach elicited serum IgG capable of cross-neutralizing selected tier 2 viruses lacking N-glycans at residue N276 (natural or

  17. Structural basis for the oligomerization-state switch from a dimer to a trimer of an engineered cortexillin-1 coiled-coil variant.

    Science.gov (United States)

    Bjelić, Saša; Wieser, Mara; Frey, Daniel; Stirnimann, Christian U; Chance, Mark R; Jaussi, Rolf; Steinmetz, Michel O; Kammerer, Richard A

    2013-01-01

    Coiled coils are well suited to drive subunit oligomerization and are widely used in applications ranging from basic research to medicine. The optimization of these applications requires a detailed understanding of the molecular determinants that control of coiled-coil formation. Although many of these determinants have been identified and characterized in great detail, a puzzling observation is that their presence does not necessarily correlate with the oligomerization state of a given coiled-coil structure. Thus, other determinants must play a key role. To address this issue, we recently investigated the unrelated coiled-coil domains from GCN4, ATF1 and cortexillin-1 as model systems. We found that well-known trimer-specific oligomerization-state determinants, such as the distribution of isoleucine residues at heptad-repeat core positions or the trimerization motif Arg-h-x-x-h-Glu (where h = hydrophobic amino acid; x = any amino acid), switch the peptide's topology from a dimer to a trimer only when inserted into the trigger sequence, a site indispensable for coiled-coil formation. Because high-resolution structural information could not be obtained for the full-length, three-stranded cortexillin-1 coiled coil, we here report the detailed biophysical and structural characterization of a shorter variant spanning the trigger sequence using circular dichroism, anatytical ultracentrifugation and x-ray crystallography. We show that the peptide forms a stable α-helical trimer in solution. We further determined the crystal structure of an optimised variant at a resolution of 1.65 Å, revealing that the peptide folds into a parallel, three-stranded coiled coil. The two complemented R-IxxIE trimerization motifs and the additional hydrophobic core isoleucine residue adopt the conformations seen in other extensively characterized parallel, three-stranded coiled coils. These findings not only confirm the structural basis for the switch in oligomerization state from

  18. Modeling Transmembrane Domain Dimers/Trimers of Plexin Receptors: Implications for Mechanisms of Signal Transmission across the Membrane

    Science.gov (United States)

    Zhang, Liqun; Polyansky, Anton; Buck, Matthias

    2015-01-01

    Single-pass transmembrane (TM) receptors transmit signals across lipid bilayers by helix association or by configurational changes within preformed dimers. The structure determination for such TM regions is challenging and has mostly been accomplished by NMR spectroscopy. Recently, the computational prediction of TM dimer structures is becoming recognized for providing models, including alternate conformational states, which are important for receptor regulation. Here we pursued a strategy to predict helix oligomers that is based on packing considerations (using the PREDDIMER webserver) and is followed by a refinement of structures, utilizing microsecond all-atom molecular dynamics simulations. We applied this method to plexin TM receptors, a family of 9 human proteins, involved in the regulation of cell guidance and motility. The predicted models show that, overall, the preferences identified by PREDDIMER are preserved in the unrestrained simulations and that TM structures are likely to be diverse across the plexin family. Plexin-B1 and –B3 TM helices are regular and tend to associate, whereas plexin-A1, -A2, –A3, -A4, -C1 and –D1 contain sequence elements, such as poly-Glycine or aromatic residues that distort helix conformation and association. Plexin-B2 does not form stable dimers due to the presence of TM prolines. No experimental structural information on the TM region is available for these proteins, except for plexin-C1 dimeric and plexin-B1 – trimeric structures inferred from X-ray crystal structures of the intracellular regions. Plexin-B1 TM trimers utilize Ser and Thr sidechains for interhelical contacts. We also modeled the juxta-membrane (JM) region of plexin-C1 and plexin-B1 and show that it synergizes with the TM structures. The structure and dynamics of the JM region and TM-JM junction provide determinants for the distance and distribution of the intracellular domains, and for their binding partners relative to the membrane. The structures

  19. Characterization of Intermolecular Interactions at Play in the 2,2,2-TRIFLUOROETHANOL Trimers Using Cavity and Chirped-Pulse Microwave Spectroscopy

    Science.gov (United States)

    Seifert, Nathan A.; Thomas, Javix; Jäger, Wolfgang; Xu, Yunjie

    2017-06-01

    2,2,2-trifluoroethanol (TFE) is a common aqueous co-solvent in biological chemistry which may induce or destabilize secondary structures of proteins and polypeptides, thanks to its diverse intermolecular linkages originating from the hydrogen bonding potential of both the hydroxyl and perfluoro groups. Theoretically, the TFE monomer is predicted to have two stable gauche (gauche^{+}/gauche^{-}) conformations whereas the trans form is unstable or is supported only by a very shallow potential. Only the gauche conformers have been identified in the gas phase, whereas liquid phase studies suggest a trans:gauche ratio of 2:3. The question at which sample (cluster) size the trans form of TFE would appear was one major motivation for our study. Here, we report the detection of three trimers of TFE using Balle-Flygare cavity and chirped-pulse Fourier transform microwave spectroscopy (CP-FTMW) techniques. The most stable observed trimer features one trans- and two gauche-TFE subunits. The other two trimers, observed using a newly constructed 2-6 GHz CP-FTMW spectrometer, consist of only the two gauche conformers of TFE. Quantum Theory of Atoms in Molecules (QTAIM) and non-covalent interactions (NCI) analyses give detailed insights into which intermolecular interactions are at play to stabilize the trans form of TFE in the most stable trimer. M. Buck, Q. Rev. Biophys. 1998, 31, 297-335. I. Bakó, T. Radnai, M. Claire, B. Funel, J. Chem. Phys. 2004, 121, 12472-12480. R. F. W. Bader, Chem. Rev. 1991, 91, 893-928. E. R. Johnson, S. Keinan, P. Mori-Sánchez, J. Contreras-Garcia, A. J. Cohen, W. Yang, J. Am. Chem. Soc., 2010, 132, 6498-6506.

  20. Microsecond Dynamics and Network Analysis of the HIV-1 SOSIP Env Trimer Reveal Collective Behavior and Conserved Microdomains of the Glycan Shield.

    Science.gov (United States)

    Lemmin, Thomas; Soto, Cinque; Stuckey, Jonathan; Kwong, Peter D

    2017-10-03

    The trimeric HIV-1-envelope (Env) spike is one of the most glycosylated protein complexes known, with roughly half its mass comprising host-derived N-linked glycan. Here we use molecular dynamics to provide insight into its structural dynamics and into how both protomer and glycan movements coordinate to shield the Env protein surface. A 2-μs molecular dynamics simulation of a fully glycosylated atomistic model of the HIV-1 SOSIP Env trimer revealed a spectrum of protomer-scissoring and trimer-opening movements. Network analysis showed that highly conserved glycans combined with protomer scissoring to restrict access to the binding site of the CD4 receptor. The network property of betweenness centrality appeared to identify whether glycans spread to restrict access or cluster to maintain the high-mannose character of the shield. We also observed stable microdomains comprising patches of glycan, with neutralizing antibodies generally binding at the interface between glycan microdomains. Overall, our results provide a microsecond-based understanding of the Env glycan shield. Published by Elsevier Ltd.

  1. A recombinant rabies vaccine expressing the trimeric form of the glycoprotein confers enhanced immunogenicity and protection in outbred mice.

    Science.gov (United States)

    Koraka, Penelope; Bosch, Berend-Jan; Cox, Manon; Chubet, Rick; Amerongen, Geert van; Lövgren-Bengtsson, Karen; Martina, Byron E E; Roose, Jouke; Rottier, Peter J M; Osterhaus, Albert D M E

    2014-08-06

    Rabies is a disease characterized by an invariably lethal encephalitis of viral origin that can be controlled by preventive vaccination programs of wildlife, domestic animals and humans in areas with a high risk of exposure. Currently available vaccines are expensive, cumbersome to produce and require intensive immunization and booster schemes to induce and maintain protective immunity. In the present study, we describe the development of candidate recombinant subunit rabies vaccines based on the glycoprotein G of the prototype rabies virus (RABV-G) expressed either as a monomer (RABV-mG) or in its native trimeric configuration (RABV-tG), with or without Matrix-M™ adjuvant. Immunogenicity and protective efficacy of the respective candidate vaccines were tested in outbred NIH Swiss albino mice. The RABV-tG candidate vaccine proved to be superior to the RABV-mG vaccine candidate both in terms of immunogenicity and efficacy. The relatively poor immunogenicity of the RABV-mG vaccine candidate was greatly improved by the addition of the adjuvant. A single, low dose of RABV-tG in combination with Matrix-M™ induced high levels of high avidity neutralizing antibodies and protected all mice against challenge with a lethal dose of RABV. Consequently RABV-tG used in combination with Matrix-M™ is a promising vaccine candidate that overcomes the limitations of currently used vaccines. Copyright © 2014 Elsevier Ltd. All rights reserved.

  2. Structural Diversity in the Complexes of Trimeric Perfluoro- o -phenylene Mercury with Tetrathia- and Tetramethyltetraselenafulvalene

    Energy Technology Data Exchange (ETDEWEB)

    Castañeda, Raúl [New Mexico Highlands Univ., Las Vegas, NV (United States); Yakovenko, Andrey A. [Argonne National Lab. (ANL), Argonne, IL (United States); Draguta, Sergiu [New Mexico Highlands Univ., Las Vegas, NV (United States); Fonari, Marina S. [New Mexico Highlands Univ., Las Vegas, NV (United States); Academy of Sciences of Moldova, Chisinau (Moldova); Antipin, Mikhail Yu. [New Mexico Highlands Univ., Las Vegas, NV (United States); Russian Academy of Sciences (RAS), Moscow (Russian Federation); Timofeeva, Tatiana V. [New Mexico Highlands Univ., Las Vegas, NV (United States); ITMO Univ., St. Petersburg (Russia)

    2015-03-04

    Five potential charge transfer complexes of trimeric perfluoro-o-phenylene mercury (I) with tetrathiafulvalene (TTF) and tetramethyltetraselenefulvalene (TMTSF) were grown from different solvent mixtures. The adducts (I)2·TTF (1) and I·TTF (2) were grown by slow evaporation from the 1:1 mixture of dichloromethane (CH2Cl2, DCM) and carbon disulfide (CS2). Use of the different 1:1 solvent mixtures of dichloromethane (CH2Cl2, DCM) and dichloroethane (C2H4Cl2, DCE) has led to the crystalline adducts I·TTF (3) and I·TTF·DCE (4). Adduct I.TMTSF (5) was grown by the interface crystallization on the border of two immiscible layers, ethyl acetate, and carbon disulfide. The cocrystals differ by the donor–acceptor ratio, molecular packing, and the solvent inclusion. The components in 1–5 form mixed donor–acceptor stacks. The stacks are stabilized by Hg···S and Hg···C short contacts, while the lateral interactions between stacks include F···F, CH···F, and S/Se···F short contacts.

  3. Trimeric autotransporter adhesins in members of the Burkholderia cepacia complex: a multifunctional family of proteins implicated in virulence

    Directory of Open Access Journals (Sweden)

    Arsénio Mendes Fialho

    2011-12-01

    Full Text Available Trimeric autotransporter adhesins (TAAs are multimeric surface proteins, involved in various biological traits of pathogenic Gram-negative bacteria including adherence, biofilm formation, invasion, survival within eukaryotic cells, serum resistance and cytotoxicity. TAAs have a modular architecture composed by a conserved membrane-anchored C-terminal domain and a variable number of stalk and head domains. In this study, a bioinformatic approach has been used to analyze the distribution and architecture of TAAs among Burkholderia cepacia complex (Bcc genomes. Fifteen genomes were probed revealing a total of 74 encoding sequences. Compared with other bacterial species, the Bcc genomes contain a disproportionately large number of TAAs (two genes to up to 8 genes, such as in B.cenocepacia. Phylogenetic analysis showed that the TAAs grouped into at least eight distinct clusters. TAAs with serine-rich repeats are clearly well separated from others, thereby representing a different evolutionary lineage. Comparative gene mapping across Bcc genomes reveals that TAA genes are inserted within conserved synteny blocks. We further focused our analysis on the epidemic strain B. cenocepacia J2315 in which 7 TAAs were annotated. Among these, 3 TAA-encoding genes (BCAM019, BCAM0223 and BCAM0224 are organized into a cluster and are candidates for multifunctional virulence factors. Here we review the current insights into the functional role of BCAM0224 as a model locus.

  4. Isolation of dimeric, trimeric, tetrameric and pentameric procyanidins from unroasted cocoa beans (Theobroma cacao L.) using countercurrent chromatography.

    Science.gov (United States)

    Esatbeyoglu, Tuba; Wray, Victor; Winterhalter, Peter

    2015-07-15

    The main procyanidins, including dimeric B2 and B5, trimeric C1, tetrameric and pentameric procyanidins, were isolated from unroasted cocoa beans (Theobroma cacao L.) using various techniques of countercurrent chromatography, such as high-speed countercurrent chromatography (HSCCC), low-speed rotary countercurrent chromatography (LSRCCC) and spiral-coil LSRCCC. Furthermore, dimeric procyanidins B1 and B7, which are not present naturally in the analysed cocoa beans, were obtained after semisynthesis of cocoa bean polymers with (+)-catechin as nucleophile and separated by countercurrent chromatography. In this way, the isolation of dimeric procyanidin B1 in considerable amounts (500mg, purity>97%) was possible in a single run. This is the first report concerning the isolation and semisynthesis of dimeric to pentameric procyanidins from T. cacao by countercurrent chromatography. Additionally, the chemical structures of tetrameric (cinnamtannin A2) and pentameric procyanidins (cinnamtannin A3) were elucidated on the basis of (1)H NMR spectroscopy. Interflavanoid linkage was determined by NOE-correlations, for the first time. Copyright © 2015 Elsevier Ltd. All rights reserved.

  5. Strong electroactive biodegradable shape memory polymer networks based on star-shaped polylactide and aniline trimer for bone tissue engineering.

    Science.gov (United States)

    Xie, Meihua; Wang, Ling; Ge, Juan; Guo, Baolin; Ma, Peter X

    2015-04-01

    Preparation of functional shape memory polymer (SMP) for tissue engineering remains a challenge. Here the synthesis of strong electroactive shape memory polymer (ESMP) networks based on star-shaped polylactide (PLA) and aniline trimer (AT) is reported. Six-armed PLAs with various chain lengths were chemically cross-linked to synthesize SMP. After addition of an electroactive AT segment into the SMP, ESMP was obtained. The polymers were characterized by (1)H NMR, GPC, FT-IR, CV, DSC, DMA, tensile test, and degradation test. The SMP and ESMP exhibited strong mechanical properties (modulus higher than GPa) and excellent shape memory performance: short recovery time (several seconds), high recovery ratio (over 94%), and high fixity ratio (almost 100%). Moreover, cyclic voltammetry test confirmed the electroactivity of the ESMP. The ESMP significantly enhanced the proliferation of C2C12 cells compared to SMP and linear PLA (control). In addition, the ESMP greatly improved the osteogenic differentiation of C2C12 myoblast cells compared to PH10 and PLA in terms of ALP enzyme activity, immunofluorescence staining, and relative gene expression by quantitative real-time polymerase chain reaction (qRT-PCR). These intelligent SMPs and electroactive SMP with strong mechanical properties, tunable degradability, good electroactivity, biocompatibility, and enhanced osteogenic differentiation of C2C12 cells show great potential for bone regeneration.

  6. The cyclic ground state structure of the HF trimer revealed by far infrared jet-cooled Fourier transform spectroscopy.

    Science.gov (United States)

    Asselin, P; Soulard, P; Madebène, B; Goubet, M; Huet, T R; Georges, R; Pirali, O; Roy, P

    2014-03-14

    The rovibrationally resolved Fourier transform (FT) far infrared (FIR) spectra of two intermolecular librations of (HF)3, namely the in-plane ν6 and out-of-plane ν4 bending fundamentals centered, respectively, at about 494 cm(-1) and 602 cm(-1), have been recorded for the first time under jet-cooled conditions using the supersonic jet of the Jet-AILES apparatus. The simultaneous rotational analysis of 245 infrared transitions belonging to both bands enabled us to determine the ground state (GS), ν6 and ν4 rotational and centrifugal distortion constants. These results provided definite experimental answers to the structure of such a weakly bound trimer: firstly the vibrationally averaged planarity of cyclic (HF)3, also supported by the very small value of the inertia defect obtained in the GS, secondly the slight weakening of the hydrogen bond in the intermolecular excited states evidenced from the center of mass separations of the HF constituents determined in the ground, ν6 = 1 and ν4 = 1 states of (HF)3 as well as the decrease of the fitted rotational constants upon excitation. Finally, lower bounds of about 2 ns on ν6 and ν4 state lifetimes could be derived from the deconvolution of experimental linewidths. Such long lifetimes highlight the interest in probing low frequency intermolecular motions of molecular complexes to get rid of constraints related to the vibrational dynamics of coupled anharmonic vibrations at higher energy, resulting in loss of rotational information.

  7. Support effects and reaction mechanism of acetylene trimerization over silica-supported Cu4 clusters: A DFT study

    Science.gov (United States)

    Maleki, Farahnaz; Schlexer, Philomena; Pacchioni, Gianfranco

    2018-02-01

    Oxide-supported Cu nanoparticles and clusters catalyze a variety of important reactions, such as CO/CO2 hydrogenation to methanol. Recent studies demonstrate that also sub-nanometer clusters consisting of only a few atoms can actively catalyze chemical reactions. In this study, we investigate the interaction between Cu4 clusters and silica-surfaces, considering the de-hydroxylated and the fully hydroxylated α-quartz surfaces. We also considered various dopants such as Ti- and Nb-ions substitutional to Si, respectively, in order to see if an electronic change of the support has an effect on the reaction of the supported cluster. We find that hydroxyl groups can enhance the adsorption energy of the cluster, whereas the dopants have only little effects on the adsorption mode of the Cu cluster. On the fully hydroxylated surface, the cluster may react with the hydroxyl groups via reverse hydrogen spillover. Finally, we explore the reactivity of the silica-supported Cu4 cluster in terms of acetylene trimerization, for which extended Cu surfaces have shown catalytic activity. We find that this reaction should occur with activation barriers below 0.8 eV; Nb-doping of the support does not seem to produce any direct effect on the reactivity of the Cu tetramer.

  8. HIV-1 adenoviral vector vaccines expressing multi-trimeric BAFF and 4-1BBL enhance T cell mediated anti-viral immunity.

    Directory of Open Access Journals (Sweden)

    Saravana Kanagavelu

    Full Text Available Adenoviral vectored vaccines have shown considerable promise but could be improved by molecular adjuvants. Ligands in the TNF superfamily (TNFSF are potential adjuvants for adenoviral vector (Ad5 vaccines based on their central role in adaptive immunity. Many TNFSF ligands require aggregation beyond the trimeric state (multi-trimerization for optimal biological function. Here we describe Ad5 vaccines for HIV-1 Gag antigen (Ad5-Gag adjuvanted with the TNFSF ligands 4-1BBL, BAFF, GITRL and CD27L constructed as soluble multi-trimeric proteins via fusion to Surfactant Protein D (SP-D as a multimerization scaffold. Mice were vaccinated with Ad5-Gag combined with Ad5 expressing one of the SP-D-TNFSF constructs or single-chain IL-12p70 as adjuvant. To evaluate vaccine-induced protection, mice were challenged with vaccinia virus expressing Gag (vaccinia-Gag which is known to target the female genital tract, a major route of sexually acquired HIV-1 infection. In this system, SP-D-4-1BBL or SP-D-BAFF led to significantly reduced vaccinia-Gag replication when compared to Ad5-Gag alone. In contrast, IL-12p70, SP-D-CD27L and SP-D-GITRL were not protective. Histological examination following vaccinia-Gag challenge showed a dramatic lymphocytic infiltration into the uterus and ovaries of SP-D-4-1BBL and SP-D-BAFF-treated animals. By day 5 post challenge, proinflammatory cytokines in the tissue were reduced, consistent with the enhanced control over viral replication. Splenocytes had no specific immune markers that correlated with protection induced by SP-D-4-1BBL and SP-D-BAFF versus other groups. IL-12p70, despite lack of anti-viral efficacy, increased the total numbers of splenic dextramer positive CD8+ T cells, effector memory T cells, and effector Gag-specific CD8+ T cells, suggesting that these markers are poor predictors of anti-viral immunity in this model. In conclusion, soluble multi-trimeric 4-1BBL and BAFF adjuvants led to strong protection from

  9. Immunization of rabbits with highly purified, soluble, trimeric human immunodeficiency virus type 1 envelope glycoprotein induces a vigorous B cell response and broadly cross-reactive neutralization.

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    Gerald V Quinnan

    Full Text Available Previously we described induction of cross-reactive HIV-1 neutralizing antibody responses in rabbits using a soluble HIV-1 gp140 envelope glycoprotein (Env in an adjuvant containing monophosphoryl lipid A (MPL and QS21 (AS02A. Here, we compared different forms of the same HIV-1 strain R2 Env for antigenic and biophysical characteristics, and in rabbits characterized the extent of B cell induction for specific antibody expression and secretion and neutralizing responses. The forms of this Env that were produced in and purified from stably transformed 293T cells included a primarily dimeric gp140, a trimeric gp140 appended to a GCN4 trimerization domain (gp140-GCN4, gp140-GCN4 with a 15 amino acid flexible linker between the gp120 and gp41 ectodomain (gp140-GCN4-L, also trimeric, and a gp140 with the flexible linker purified from cell culture supernatants as either dimer (gp140-L(D or monomer (gp140-L(M. Multimeric states of the Env proteins were assessed by native gel electrophoresis and analytical ultracentrifugation. The different forms of gp140 bound broadly cross-reactive neutralizing (BCN human monoclonal antibodies (mAbs similarly in ELISA and immunoprecipitation assays. All Envs bound CD4i mAbs in the presence and absence of sCD4, as reported for the R2 Env. Weak neutralization of some strains of HIV-1 was seen after two additional doses in AS02A. Rabbits that were given a seventh dose of gp140-GCN4-L developed BCN responses that were weak to moderate, similar to our previous report. The specificity of these responses did not appear similar to that of any of the known BCN human mAbs. Induction of spleen B cell and plasma cells producing immunoglobulins that bound trimeric gp140-GCN4-L was vigorous, based on ELISpot and flow cytometry analyses. The results demonstrate that highly purified gp140-GCN4-L trimer in adjuvant elicits BCN responses in rabbits accompanied by vigorous B cell induction.

  10. Structure-Based Design of a Soluble Prefusion-Closed HIV-1 Env Trimer with Reduced CD4 Affinity and Improved Immunogenicity

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    Chuang, Gwo-Yu; Geng, Hui; Pancera, Marie; Xu, Kai; Cheng, Cheng; Acharya, Priyamvada; Chambers, Michael; Druz, Aliaksandr; Tsybovsky, Yaroslav; Wanninger, Timothy G.; Yang, Yongping; Doria-Rose, Nicole A.; Georgiev, Ivelin S.; Gorman, Jason; Joyce, M.Gordon; O; Dell, Sijy; Zhou, Tongqing; McDermott, Adrian B.; Mascola, John R.; Kwong, Peter D. (NIH); (FNL)

    2017-03-08

    ABSTRACT

    The HIV-1 envelope (Env) trimer is a target for vaccine design as well as a conformational machine that facilitates virus entry by transitioning between prefusion-closed, CD4-bound, and coreceptor-bound conformations by transitioning into a postfusion state. Vaccine designers have sought to restrict the conformation of the HIV-1 Env trimer to its prefusion-closed state as this state is recognized by most broadly neutralizing, but not nonneutralizing, antibodies. We previously identified a disulfide bond, I201C-A433C (DS), which stabilizes Env in the vaccine-desired prefusion-closed state. When placed into the context of BG505 SOSIP.664, a soluble Env trimer mimic developed by Sanders, Moore, and colleagues, the engineered DS-SOSIP trimer showed reduced conformational triggering by CD4. Here, we further stabilize DS-SOSIP through a combination of structure-based design and 96-well-based expression and antigenic assessment. From 103 designs, we identified one, named DS-SOSIP.4mut, with four additional mutations at the interface of potentially mobile domains of the prefusion-closed structure. We also determined the crystal structures of DS-SOSIP.4mut at 4.1-Å resolution and of an additional DS-SOSIP.6mut variant at 4.3-Å resolution, and these confirmed the formation of engineered disulfide bonds. Notably, DS-SOSIP.4mut elicited a higher ratio of tier 2 autologous titers versus tier 1 V3-sensitive titers than BG505 SOSIP.664. DS-SOSIP.4mut also showed reduced recognition of CD4 and increased thermostability. The improved antigenicity, thermostability, and immunogenicity of DS-SOSIP.4mut suggest utility as an immunogen or a serologic probe; moreover, the specific four alterations identified here, M154, M300, M302, and L320 (4mut), can also be transferred to other HIV-1 Env trimers of interest to improve their properties.

    IMPORTANCEOne approach to elicit broadly neutralizing antibodies against HIV-1 is to stabilize the

  11. Structure-Based Design of a Soluble Prefusion-Closed HIV-1 Env Trimer with Reduced CD4 Affinity and Improved Immunogenicity.

    Science.gov (United States)

    Chuang, Gwo-Yu; Geng, Hui; Pancera, Marie; Xu, Kai; Cheng, Cheng; Acharya, Priyamvada; Chambers, Michael; Druz, Aliaksandr; Tsybovsky, Yaroslav; Wanninger, Timothy G; Yang, Yongping; Doria-Rose, Nicole A; Georgiev, Ivelin S; Gorman, Jason; Joyce, M Gordon; O'Dell, Sijy; Zhou, Tongqing; McDermott, Adrian B; Mascola, John R; Kwong, Peter D

    2017-05-15

    The HIV-1 envelope (Env) trimer is a target for vaccine design as well as a conformational machine that facilitates virus entry by transitioning between prefusion-closed, CD4-bound, and coreceptor-bound conformations by transitioning into a postfusion state. Vaccine designers have sought to restrict the conformation of the HIV-1 Env trimer to its prefusion-closed state as this state is recognized by most broadly neutralizing, but not nonneutralizing, antibodies. We previously identified a disulfide bond, I201C-A433C (DS), which stabilizes Env in the vaccine-desired prefusion-closed state. When placed into the context of BG505 SOSIP.664, a soluble Env trimer mimic developed by Sanders, Moore, and colleagues, the engineered DS-SOSIP trimer showed reduced conformational triggering by CD4. Here, we further stabilize DS-SOSIP through a combination of structure-based design and 96-well-based expression and antigenic assessment. From 103 designs, we identified one, named DS-SOSIP.4mut, with four additional mutations at the interface of potentially mobile domains of the prefusion-closed structure. We also determined the crystal structures of DS-SOSIP.4mut at 4.1-Å resolution and of an additional DS-SOSIP.6mut variant at 4.3-Å resolution, and these confirmed the formation of engineered disulfide bonds. Notably, DS-SOSIP.4mut elicited a higher ratio of tier 2 autologous titers versus tier 1 V3-sensitive titers than BG505 SOSIP.664. DS-SOSIP.4mut also showed reduced recognition of CD4 and increased thermostability. The improved antigenicity, thermostability, and immunogenicity of DS-SOSIP.4mut suggest utility as an immunogen or a serologic probe; moreover, the specific four alterations identified here, M154, M300, M302, and L320 (4mut), can also be transferred to other HIV-1 Env trimers of interest to improve their properties. IMPORTANCE One approach to elicit broadly neutralizing antibodies against HIV-1 is to stabilize the structurally flexible HIV-1 envelope (Env) trimer

  12. Mechanistic studies of olefin and alkyne trimerization with chromium catalysts: deuterium labeling and studies of regiochemistry using a model chromacyclopentane complex.

    Science.gov (United States)

    Agapie, Theodor; Labinger, Jay A; Bercaw, John E

    2007-11-21

    A system for catalytic trimerization of ethylene utilizing chromium(III) precursors supported by diphosphine ligand PNP(O4) = (o-MeO-C6H4)2PN(Me)P(o-MeO-C6H4)2 has been investigated. The mechanism of the olefin trimerization reaction was examined using deuterium labeling and studies of reactions with alpha-olefins and internal olefins. A well-defined chromium precursor utilized in this studies is Cr(PNP(O4))(o,o'-biphenyldiyl)Br. A cationic species, obtained by halide abstraction with NaB[C6H3(CF3)2]4, is required for catalytic turnover to generate 1-hexene from ethylene. The initiation byproduct is vinylbiphenyl; this is formed even without activation by halide abstraction. Trimerization of 2-butyne is accomplished by the same cationic system but not by the neutral species. Catalytic trimerization, with various (PNP(O4))Cr precursors, of a 1:1 mixture of C2D4 and C2H4 gives isotopologs of 1-hexene without H/D scrambling (C6D12, C6D8H4, C6D4H8, and C6H12 in a 1:3:3:1 ratio). The lack of crossover supports a mechanism involving metallacyclic intermediates. Using a SHOP catalyst to perform the oligomerization of a 1:1 mixture of C2D4 and C2H4 leads to the generation of a broader distribution of 1-hexene isotopologs, consistent with a Cossee-type mechanism for 1-hexene formation. The ethylene trimerization reaction was further studied by the reaction of trans-, cis-, and gem-ethylene-d2 upon activation of Cr(PNP(O4))(o,o'-biphenyldiyl)Br with NaB[C6H3(CF3)2]4. The trimerization of cis- and trans-ethylene-d2 generates 1-hexene isotopomers having terminal CDH groups, with an isotope effect of 3.1(1) and 4.1(1), respectively. These results are consistent with reductive elimination of 1-hexene from a putative Cr(H)[(CH2)4CH=CH2] occurring much faster than a hydride 2,1-insertion or with concerted 1-hexene formation from a chromacycloheptane via a 3,7-H shift. The trimerization of gem-ethylene-d2 has an isotope effect of 1.3(1), consistent with irreversible formation of a

  13. Molecular determinants of transport stimulation of EAAT2 are located at interface between the trimerization and substrate transport domains.

    Science.gov (United States)

    Mortensen, Ole V; Liberato, José L; Coutinho-Netto, Joaquim; Dos Santos, Wagner F; Fontana, Andréia C K

    2015-04-01

    Excitatory amino acid transporters (EAATs) regulate glutamatergic signal transmission by clearing extracellular glutamate. Dysfunction of these transporters has been implicated in the pathogenesis of various neurological disorders. Previous studies have shown that venom from the spider Parawixia bistriata and a purified compound (Parawixin1) stimulate EAAT2 activity and protect retinal tissue from ischemic damage. In the present study, the EAAT2 subtype specificity of this compound was explored, employing chimeric proteins between EAAT2 and EAAT3 transporter subtypes and mutants to characterize the structural region targeted by the compound. This identified a critical residue (Histidine-71 in EAAT2 and Serine-45 in EAAT3) in transmembrane domain 2 (TM2) to be important for the selectivity between EAAT2 and EAAT3 and for the activity of the venom. Using the identified residue in TM2 as a structural anchor, several neighboring amino acids within TM5 and TM8 were identified to also be important for the activity of the venom. This structural domain of the transporter lies at the interface of the rigid trimerization domain and the central substrate-binding transport domain. Our studies suggest that the mechanism of glutamate transport enhancement involves an interaction with the transporter that facilitates the movement of the transport domain. We identified a domain (purple star) in the glutamate transporter EAAT2 that is important for transport stimulation through a spider venom, and suggest a mechanism for enhanced transporter function through facilitated substrate translocation (arrow). Because the dysfunction of glutamate transporters is implicated in the pathogenesis of neurological disorders, understanding the mechanisms of enhanced transport could have therapeutic implications. © 2015 International Society for Neurochemistry.

  14. High-resolution X-ray structure of the trimeric Scar/WAVE-complex precursor Brk1.

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    Joern Linkner

    Full Text Available The Scar/WAVE-complex links upstream Rho-GTPase signaling to the activation of the conserved Arp2/3-complex. Scar/WAVE-induced and Arp2/3-complex-mediated actin nucleation is crucial for actin assembly in protruding lamellipodia to drive cell migration. The heteropentameric Scar/WAVE-complex is composed of Scar/WAVE, Abi, Nap, Pir and a small polypeptide Brk1/HSPC300, and recent work suggested that free Brk1 serves as a homooligomeric precursor in the assembly of this complex. Here we characterized the Brk1 trimer from Dictyostelium by analytical ultracentrifugation and gelfiltration. We show for the first time its dissociation at concentrations in the nanomolar range as well as an exchange of subunits within different DdBrk1 containing complexes. Moreover, we determined the three-dimensional structure of DdBrk1 at 1.5 Å resolution by X-ray crystallography. Three chains of DdBrk1 are associated with each other forming a parallel triple coiled-coil bundle. Notably, this structure is highly similar to the heterotrimeric α-helical bundle of HSPC300/WAVE1/Abi2 within the human Scar/WAVE-complex. This finding, together with the fact that Brk1 is collectively sandwiched by the remaining subunits and also constitutes the main subunit connecting the triple-coil domain of the HSPC300/WAVE1/Abi2/ heterotrimer to Sra1(Pir1, implies a critical function of this subunit in the assembly process of the entire Scar/WAVE-complex.

  15. A single immunization with soluble recombinant trimeric hemagglutinin protects chickens against highly pathogenic avian influenza virus H5N1.

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    Lisette A H M Cornelissen

    Full Text Available BACKGROUND: The highly pathogenic avian influenza (HPAI virus H5N1 causes multi-organ disease and death in poultry, resulting in significant economic losses in the poultry industry. In addition, it poses a major public health threat as it can be transmitted directly from infected poultry to humans with very high (60% mortality rate. Effective vaccination against HPAI H5N1 would protect commercial poultry and would thus provide an important control measure by reducing the likelihood of bird-to-bird and bird-to-human transmission. METHODOLOGY/PRINCIPAL FINDINGS: In the present study we evaluated the vaccine potential of recombinant soluble trimeric subtype 5 hemagglutinin (sH5(3 produced in mammalian cells. The secreted, purified sH5(3 was biologically active as demonstrated by its binding to ligands in a sialic acid-dependent manner. It was shown to protect chickens, in a dose-dependent manner, against a lethal challenge with H5N1 after a single vaccination. Protected animals did not shed challenge virus as determined by a quantitative RT-PCR on RNA isolated from trachea and cloaca swabs. Also in mice, vaccination with sH5(3 provided complete protection against challenge with HPAI H5N1. CONCLUSIONS/SIGNIFICANCE: Our results demonstrate that sH5(3 constitutes an attractive vaccine antigen for protection of chickens and mammals against HPAI H5N1. As these recombinant soluble hemagglutinin preparations can be produced with high yields and with relatively short lead time, they enable a rapid response to circulating and potentially pandemic influenza viruses.

  16. THE EFFECT OF THE HYDROGEN ION CONCENTRATION ON THE RATE OF HYDROLYSIS OF GLYCYL GLYCINE, GLYCYL LEUCINE, GLYCYL ALANINE, GLYCYL ASPARAGINE, GLYCYL ASPARTIC ACID, AND BIURET BASE BY EREPSIN

    Science.gov (United States)

    Northrop, John H.; Simms, Henry S.

    1928-01-01

    1. The rate of hydrolysis at different pH values of glycyl glycine, glycyl leucine, glycyl alanine, glycyl asparagine, glycyl aspartic acid and biuret base has been determined. 2. The pH-activity curves obtained in this way differ for the different substrates. 3. The curves can be satisfactorily predicted by the assumption that erepsin is a weak acid or base with a dissociation constant of 10–7.6 and that the reaction takes place between a particular ionic species of the enzyme and of the substrate. There are several possible arrangements which will predict the experimental results. 4. The rate of inactivation of erepsin at various pH values has been determined and found to agree with the assumption used above, that the enzyme is a weak acid or base with a dissociation constant of about 10–7.6. 5. It is pointed out that if the mechanism assumed is correct, the determination of a significant value for the relative rate of hydrolysis of various peptides is a very uncertain procedure. PMID:19872461

  17. THE EFFECT OF THE HYDROGEN ION CONCENTRATION ON THE RATE OF HYDROLYSIS OF GLYCYL GLYCINE, GLYCYL LEUCINE, GLYCYL ALANINE, GLYCYL ASPARAGINE, GLYCYL ASPARTIC ACID, AND BIURET BASE BY EREPSIN.

    Science.gov (United States)

    Northrop, J H; Simms, H S

    1928-11-20

    1. The rate of hydrolysis at different pH values of glycyl glycine, glycyl leucine, glycyl alanine, glycyl asparagine, glycyl aspartic acid and biuret base has been determined. 2. The pH-activity curves obtained in this way differ for the different substrates. 3. The curves can be satisfactorily predicted by the assumption that erepsin is a weak acid or base with a dissociation constant of 10(-7.6) and that the reaction takes place between a particular ionic species of the enzyme and of the substrate. There are several possible arrangements which will predict the experimental results. 4. The rate of inactivation of erepsin at various pH values has been determined and found to agree with the assumption used above, that the enzyme is a weak acid or base with a dissociation constant of about 10(-7.6). 5. It is pointed out that if the mechanism assumed is correct, the determination of a significant value for the relative rate of hydrolysis of various peptides is a very uncertain procedure.

  18. Hidden Lineage Complexity of Glycan-Dependent HIV-1 Broadly Neutralizing Antibodies Uncovered by Digital Panning and Native-Like gp140 Trimer

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    Linling He

    2017-08-01

    Full Text Available Germline precursors and intermediates of broadly neutralizing antibodies (bNAbs are essential to the understanding of humoral response to HIV-1 infection and B-cell lineage vaccine design. Using a native-like gp140 trimer probe, we examined antibody libraries constructed from donor-17, the source of glycan-dependent PGT121-class bNAbs recognizing the N332 supersite on the HIV-1 envelope glycoprotein. To facilitate this analysis, a digital panning method was devised that combines biopanning of phage-displayed antibody libraries, 900 bp long-read next-generation sequencing, and heavy/light (H/L-paired antibodyomics. In addition to single-chain variable fragments resembling the wild-type bNAbs, digital panning identified variants of PGT124 (a member of the PGT121 class with a unique insertion in the heavy chain complementarity-determining region 1, as well as intermediates of PGT124 exhibiting notable affinity for the native-like trimer and broad HIV-1 neutralization. In a competition assay, these bNAb intermediates could effectively compete with mouse sera induced by a scaffolded BG505 gp140.681 trimer for the N332 supersite. Our study thus reveals previously unrecognized lineage complexity of the PGT121-class bNAbs and provides an array of library-derived bNAb intermediates for evaluation of immunogens containing the N332 supersite. Digital panning may prove to be a valuable tool in future studies of bNAb diversity and lineage development.

  19. β-Arrestin interacts with the beta/gamma subunits of trimeric G-proteins and dishevelled in the Wnt/Ca(2+ pathway in xenopus gastrulation.

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    Katharina Seitz

    Full Text Available β-Catenin independent, non-canonical Wnt signaling pathways play a major role in the regulation of morphogenetic movements in vertebrates. The term non-canonical Wnt signaling comprises multiple, intracellularly divergent, Wnt-activated and β-Catenin independent signaling cascades including the Wnt/Planar Cell Polarity and the Wnt/Ca(2+ cascades. Wnt/Planar Cell Polarity and Wnt/Ca(2+ pathways share common effector proteins, including the Wnt ligand, Frizzled receptors and Dishevelled, with each other and with additional branches of Wnt signaling. Along with the aforementioned proteins, β-Arrestin has been identified as an essential effector protein in the Wnt/β-Catenin and the Wnt/Planar Cell Polarity pathway. Our results demonstrate that β-Arrestin is required in the Wnt/Ca(2+ signaling cascade upstream of Protein Kinase C (PKC and Ca(2+/Calmodulin-dependent Protein Kinase II (CamKII. We have further characterized the role of β-Arrestin in this branch of non-canonical Wnt signaling by knock-down and rescue experiments in Xenopus embryo explants and analyzed protein-protein interactions in 293T cells. Functional interaction of β-Arrestin, the β subunit of trimeric G-proteins and Dishevelled is required to induce PKC activation and membrane translocation. In Xenopus gastrulation, β-Arrestin function in Wnt/Ca(2+ signaling is essential for convergent extension movements. We further show that β-Arrestin physically interacts with the β subunit of trimeric G-proteins and Dishevelled, and that the interaction between β-Arrestin and Dishevelled is promoted by the beta/gamma subunits of trimeric G-proteins, indicating the formation of a multiprotein signaling complex.

  20. Bipartite Topology of Treponema pallidum Repeat Proteins C/D and I: OUTER MEMBRANE INSERTION, TRIMERIZATION, AND PORIN FUNCTION REQUIRE A C-TERMINAL β-BARREL DOMAIN.

    Science.gov (United States)

    Anand, Arvind; LeDoyt, Morgan; Karanian, Carson; Luthra, Amit; Koszelak-Rosenblum, Mary; Malkowski, Michael G; Puthenveetil, Robbins; Vinogradova, Olga; Radolf, Justin D

    2015-05-08

    We previously identified Treponema pallidum repeat proteins TprC/D, TprF, and TprI as candidate outer membrane proteins (OMPs) and subsequently demonstrated that TprC is not only a rare OMP but also forms trimers and has porin activity. We also reported that TprC contains N- and C-terminal domains (TprC(N) and TprC(C)) orthologous to regions in the major outer sheath protein (MOSP(N) and MOSP(C)) of Treponema denticola and that TprC(C) is solely responsible for β-barrel formation, trimerization, and porin function by the full-length protein. Herein, we show that TprI also possesses bipartite architecture, trimeric structure, and porin function and that the MOSP(C)-like domains of native TprC and TprI are surface-exposed in T. pallidum, whereas their MOSP(N)-like domains are tethered within the periplasm. TprF, which does not contain a MOSP(C)-like domain, lacks amphiphilicity and porin activity, adopts an extended inflexible structure, and, in T. pallidum, is tightly bound to the protoplasmic cylinder. By thermal denaturation, the MOSP(N) and MOSP(C)-like domains of TprC and TprI are highly thermostable, endowing the full-length proteins with impressive conformational stability. When expressed in Escherichia coli with PelB signal sequences, TprC and TprI localize to the outer membrane, adopting bipartite topologies, whereas TprF is periplasmic. We propose that the MOSP(N)-like domains enhance the structural integrity of the cell envelope by anchoring the β-barrels within the periplasm. In addition to being bona fide T. pallidum rare outer membrane proteins, TprC/D and TprI represent a new class of dual function, bipartite bacterial OMP. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  1. Homo-trimeric Structure of the Type IVb Minor Pilin CofB Suggests Mechanism of CFA/III Pilus Assembly in Human Enterotoxigenic Escherichia coli.

    Science.gov (United States)

    Kawahara, Kazuki; Oki, Hiroya; Fukakusa, Shunsuke; Yoshida, Takuya; Imai, Tomoya; Maruno, Takahiro; Kobayashi, Yuji; Motooka, Daisuke; Iida, Tetsuya; Ohkubo, Tadayasu; Nakamura, Shota

    2016-03-27

    In gram-negative bacteria, the assembly of type IV pilus (T4P) and the evolutionally related pseudopilus of type II secretion system involves specialized structural proteins called pilins and pseudopilins, respectively, and is dynamically regulated to promote bacterial pathogenesis. Previous studies have suggested that a structural "tip"-like hetero-complex formed through the interaction of at least three minor (pseudo) pilins plays an important role in this process, while some members of the pathogenic type IVb subfamily are known to have only one such minor pilin subunit whose function is still unknown. Here, we determined the crystal structure of the type IVb minor pilin CofB of colonization factor antigen/III from human enterotoxigenic Escherichia coli at 1.88-Å resolution. The crystal structure, in conjunction with physicochemical analysis in solution, reveals a symmetrical homo-trimeric arrangement distinct from the hetero-complexes of minor (pseudo) pilins observed in other T4P and type II secretion systems. Each CofB monomer adopts a unique three-domain architecture, in which the C-terminal β-sheet-rich lectin domain can effectively initiate trimer association of its pilin-like N-terminal domain through extensive hydrophobic interactions followed by domain swapping at the central hinge-like domain. Deletion of cofB produces a phenotype with no detectable pili formation on the cell surface, while molecular modeling indicates that the characteristic homo-trimeric structure of CofB is well situated at the pilus tip of colonization factor antigen/III formed by the major pilin CofA, suggesting a role for the minor pilin in the efficient initiation of T4P assembly. Copyright © 2016 Elsevier Ltd. All rights reserved.

  2. Structural basis of HIV-1 neutralization by affinity matured Fabs directed against the internal trimeric coiled-coil of gp41.

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    Elena Gustchina

    2010-11-01

    Full Text Available The conserved internal trimeric coiled-coil of the N-heptad repeat (N-HR of HIV-1 gp41 is transiently exposed during the fusion process by forming a pre-hairpin intermediate, thus representing an attractive target for the design of fusion inhibitors and neutralizing antibodies. In previous studies we reported a series of broadly neutralizing mini-antibodies derived from a synthetic naïve human combinatorial antibody library by panning against a mimetic of the trimeric N-HR coiled coil, followed by affinity maturation using targeted diversification of the CDR-H2 loop. Here we report crystal structures of the N-HR mimetic 5-Helix with two Fabs that represent the extremes of this series: Fab 8066 is broadly neutralizing across a wide panel of B and C type HIV-1 viruses, whereas Fab 8062 is non-neutralizing. The crystal structures reveal important differences in the conformations of the CDR-H2 loops in the complexes that propagate into other regions of the antigen-antibody interface, and suggest that both neutralization properties and affinity for the target can be attributed, at least in part, to the differences in the interactions of the CDR-H2 loops with the antigen. Furthermore, modeling of the complex of an N-HR trimer with three Fabs suggests that the CDR-H2 loop may be involved in close intermolecular contacts between neighboring antibody molecules, and that such contacts may hinder the formation of complexes between the N-HR trimer and more than one antibody molecule depending on the conformation of the bound CDR-H2 loop which is defined by its interactions with antigen. Comparison with the crystal structure of the complex of 5-Helix with another neutralizing monoclonal antibody known as D5, derived using an entirely different antibody library and panning procedure, reveals remarkable convergence in the optimal sequence and conformation of the CDR-H2 loop.

  3. Effects of the I559P gp41 Change on the Conformation and Function of the Human Immunodeficiency Virus (HIV-1) Membrane Envelope Glycoprotein Trimer

    Science.gov (United States)

    Sodroski, Joseph; Finzi, Andrés

    2015-01-01

    The mature human immunodeficiency virus (HIV-1) envelope glycoprotein (Env) trimer is produced by proteolytic cleavage of a precursor and consists of three gp120 exterior and three gp41 transmembrane subunits. The metastable Env complex is induced to undergo conformational changes required for virus entry by the binding of gp120 to the receptors, CD4 and CCR5/CXCR4. An isoleucine-to-proline change (I559P) in the gp41 ectodomain has been used to stabilize soluble forms of HIV-1 Env trimers for structural characterization and for use as immunogens. In the native membrane-anchored HIV-1BG505 Env, the I559P change modestly decreased proteolytic maturation, increased the non-covalent association of gp120 with the Env trimer, and resulted in an Env conformation distinctly different from that of the wild-type HIV-1BG505 Env. Compared with the wild-type Env, the I559P Env was recognized inefficiently by polyclonal sera from HIV-1-infected individuals, by several gp41-directed antibodies, by some antibodies against the CD4-binding site of gp120, and by antibodies that preferentially recognize the CD4-bound Env. Some of the gp120-associated antigenic differences between the wild-type HIV-1BG505 Env and the I559P mutant were compensated by the SOS disulfide bond between gp120 and gp41, which has been used to stabilize cleaved soluble Env trimers. Nonetheless, regardless of the presence of the SOS changes, Envs with proline 559 were recognized less efficiently than Envs with isoleucine 559 by the VRC01 neutralizing antibody, which binds the CD4-binding site of gp120, and the PGT151 neutralizing antibody, which binds a hybrid gp120-gp41 epitope. The I559P change completely eliminated the ability of the HIV-1BG505 Env to mediate cell-cell fusion and virus entry, and abolished the capacity of the SOS Env to support virus infection in the presence of a reducing agent. These results suggest that differences exist between the quaternary structures of functional Env spikes and I559P

  4. Effects of the I559P gp41 change on the conformation and function of the human immunodeficiency virus (HIV-1 membrane envelope glycoprotein trimer.

    Directory of Open Access Journals (Sweden)

    Nirmin Alsahafi

    Full Text Available The mature human immunodeficiency virus (HIV-1 envelope glycoprotein (Env trimer is produced by proteolytic cleavage of a precursor and consists of three gp120 exterior and three gp41 transmembrane subunits. The metastable Env complex is induced to undergo conformational changes required for virus entry by the binding of gp120 to the receptors, CD4 and CCR5/CXCR4. An isoleucine-to-proline change (I559P in the gp41 ectodomain has been used to stabilize soluble forms of HIV-1 Env trimers for structural characterization and for use as immunogens. In the native membrane-anchored HIV-1BG505 Env, the I559P change modestly decreased proteolytic maturation, increased the non-covalent association of gp120 with the Env trimer, and resulted in an Env conformation distinctly different from that of the wild-type HIV-1BG505 Env. Compared with the wild-type Env, the I559P Env was recognized inefficiently by polyclonal sera from HIV-1-infected individuals, by several gp41-directed antibodies, by some antibodies against the CD4-binding site of gp120, and by antibodies that preferentially recognize the CD4-bound Env. Some of the gp120-associated antigenic differences between the wild-type HIV-1BG505 Env and the I559P mutant were compensated by the SOS disulfide bond between gp120 and gp41, which has been used to stabilize cleaved soluble Env trimers. Nonetheless, regardless of the presence of the SOS changes, Envs with proline 559 were recognized less efficiently than Envs with isoleucine 559 by the VRC01 neutralizing antibody, which binds the CD4-binding site of gp120, and the PGT151 neutralizing antibody, which binds a hybrid gp120-gp41 epitope. The I559P change completely eliminated the ability of the HIV-1BG505 Env to mediate cell-cell fusion and virus entry, and abolished the capacity of the SOS Env to support virus infection in the presence of a reducing agent. These results suggest that differences exist between the quaternary structures of functional Env

  5. Complexes of neutralizing and non-neutralizing affinity matured Fabs with a mimetic of the internal trimeric coiled-coil of HIV-1 gp41.

    Directory of Open Access Journals (Sweden)

    Elena Gustchina

    Full Text Available A series of mini-antibodies (monovalent and bivalent Fabs targeting the conserved internal trimeric coiled-coil of the N-heptad repeat (N-HR of HIV-1 gp41 has been previously constructed and reported. Crystal structures of two closely related monovalent Fabs, one (Fab 8066 broadly neutralizing across a wide panel of HIV-1 subtype B and C viruses, and the other (Fab 8062 non-neutralizing, representing the extremes of this series, were previously solved as complexes with 5-Helix, a gp41 pre-hairpin intermediate mimetic. Binding of these Fabs to covalently stabilized chimeric trimers of N-peptides of HIV-1 gp41 (named (CCIZN363 or 3-H has now been investigated using X-ray crystallography, cryo-electron microscopy, and a variety of biophysical methods. Crystal structures of the complexes between 3-H and Fab 8066 and Fab 8062 were determined at 2.8 and 3.0 Å resolution, respectively. Although the structures of the complexes with the neutralizing Fab 8066 and its non-neutralizing counterpart Fab 8062 were generally similar, small differences between them could be correlated with the biological properties of these antibodies. The conformations of the corresponding CDRs of each antibody in the complexes with 3-H and 5-Helix are very similar. The adaptation to a different target upon complex formation is predominantly achieved by changes in the structure of the trimer of N-HR helices, as well as by adjustment of the orientation of the Fab molecule relative to the N-HR in the complex, via rigid-body movement. The structural data presented here indicate that binding of three Fabs 8062 with high affinity requires more significant changes in the structure of the N-HR trimer compared to binding of Fab 8066. A comparative analysis of the structures of Fabs complexed to different gp41 intermediate mimetics allows further evaluation of biological relevance for generation of neutralizing antibodies, as well as provides novel structural insights into immunogen

  6. Modulation of Kingella kingae adherence to human epithelial cells by type IV Pili, capsule, and a novel trimeric autotransporter.

    Science.gov (United States)

    Porsch, Eric A; Kehl-Fie, Thomas E; St Geme, Joseph W

    2012-10-23

    Kingella kingae is an emerging bacterial pathogen that is being recognized increasingly as an important etiology of septic arthritis, osteomyelitis, and bacteremia, especially in young children. Colonization of the posterior pharynx is a key step in the pathogenesis of K. kingae disease. Previous work established that type IV pili are necessary for K. kingae adherence to the respiratory epithelium. In this study, we set out to identify additional factors that influence K. kingae interactions with human epithelial cells. We found that genetic disruption of the gene encoding a predicted trimeric autotransporter protein called Knh (Kingella NhhA homolog) resulted in reduced adherence to human epithelial cells. In addition, we established that K. kingae elaborates a surface-associated polysaccharide capsule that requires a predicted ABC-type transporter export operon called ctrABCD for surface presentation. Furthermore, we discovered that the presence of a surface capsule interferes with Knh-mediated adherence to human epithelial cells by nonpiliated organisms and that maximal adherence in the presence of a capsule requires the predicted type IV pilus retraction machinery, PilT/PilU. On the basis of the data presented here, we propose a novel adherence mechanism that allows K. kingae to adhere efficiently to human epithelial cells while remaining encapsulated and more resistant to immune clearance. Kingella kingae is a Gram-negative bacterium that is being recognized increasingly as a cause of joint and bone infections in young children. The pathogenesis of disease due to K. kingae begins with bacterial colonization of the upper respiratory tract, and previous work established that surface hair-like fibers called type IV pili are necessary for K. kingae adherence to respiratory epithelial cells. In this study, we set out to identify additional factors that influence K. kingae interactions with respiratory epithelial cells. We discovered a novel surface protein called

  7. Reliable lateral manipulation of a single Ag adatom on a Ag(1 1 1) surface with a trimer-apex tip

    International Nuclear Information System (INIS)

    Xie Yiqun; Shi Wangzhou; Du Guoping

    2009-01-01

    We study the reliability of the lateral manipulation of a single Ag adatom on a Ag(1 1 1) surface with the single-atom and trimer-apex tips based on molecular statics simulations using surface embedded-atom-method potential. The dependence of the manipulation reliability on tip height and orientation is investigated. For the single-atom tip the manipulation reliability increases monotonically with decreasing tip height, which is owing to the strengthened lateral tip-adatom interaction as the tip height lowers. For the trimer-apex tip, the manipulation reliability is sensitive to the tip orientation in the lower tip-height range, while in the higher tip-height range the manipulation reliability is independent of the tip orientation and moreover can be greatly improved due to the strong vertical attraction of the tip on the adatom as compared to the single-atom tip. We also compare these results to those for manipulating single Cu adatoms on the Cu(1 1 1) surface, reveal the underlying physics, and propose the method to improve the manipulation reliability for different systems.

  8. Isolation and Structural Characterization of Monomeric and Trimeric Photosystem I Complexes (P700·FA/FB and P700·Fx) from the Cyanobacterium Synechocystis PCC 6803

    NARCIS (Netherlands)

    Kruip, Jochen; Boekema, Egbert J.; Bald, Dirk; Boonstra, Arjen F.; Rögner, Matthias

    1993-01-01

    An isolation procedure was developed for the cyanobacterium Synechocystis 6803 (and 6714) which yields both monomeric and trimeric photosystem I complexes (P700·FA/FB complexes) depleted of the stroma-exposed subunits PsaC, -D, and -E (P700·FX complexes). Analysis by high resolution gel

  9. ISOLATION AND STRUCTURAL CHARACTERIZATION OF MONOMERIC AND TRIMERIC PHOTOSYSTEM-I COMPLEXES (P700-CENTER-DOT-FA/FB AND P700-CENTER-DOT-FX) FROM THE CYANOBACTERIUM SYNECHOCYSTIS PCC-6803

    NARCIS (Netherlands)

    KRUIP, J; BOEKEMA, EJ; BALD, D; BOONSTRA, AF; ROGNER, M

    1993-01-01

    An isolation procedure was developed for the cyanobacterium Synechocystis 6803 (and 6714) which yields both monomeric and trimeric photosystem I complexes (P700.F(A)/F(B) complexes) depleted of the stroma-exposed subunits PsaC, -D, and -E (P700-F(x) complexes). Analysis by high resolution gel

  10. Complete epitopes for vaccine design derived from a crystal structure of the broadly neutralizing antibodies PGT128 and 8ANC195 in complex with an HIV-1 Env trimer.

    Science.gov (United States)

    Kong, Leopold; Torrents de la Peña, Alba; Deller, Marc C; Garces, Fernando; Sliepen, Kwinten; Hua, Yuanzi; Stanfield, Robyn L; Sanders, Rogier W; Wilson, Ian A

    2015-10-01

    The HIV-1 envelope gp160 glycoprotein (Env) is a trimer of gp120 and gp41 heterodimers that mediates cell entry and is the primary target of the humoral immune response. Broadly neutralizing antibodies (bNAbs) to HIV-1 have revealed multiple epitopes or sites of vulnerability, but mapping of most of these sites is incomplete owing to a paucity of structural information on the full epitope in the context of the Env trimer. Here, a crystal structure of the soluble BG505 SOSIP gp140 trimer at 4.6 Å resolution with the bNAbs 8ANC195 and PGT128 reveals additional interactions in comparison to previous antibody-gp120 structures. For 8ANC195, in addition to previously documented interactions with gp120, a substantial interface with gp41 is now elucidated that includes extensive interactions with the N637 glycan. Surprisingly, removal of the N637 glycan did not impact 8ANC195 affinity, suggesting that the antibody has evolved to accommodate this glycan without loss of binding energy. PGT128 indirectly affects the N262 glycan by a domino effect, in which PGT128 binds to the N301 glycan, which in turn interacts with and repositions the N262 glycan, thereby illustrating the important role of neighboring glycans on epitope conformation and stability. Comparisons with other Env trimer and gp120 structures support an induced conformation for glycan N262, suggesting that the glycan shield is allosterically modified upon PGT128 binding. These complete epitopes of two broadly neutralizing antibodies on the Env trimer can now be exploited for HIV-1 vaccine design.

  11. Energy migration in allophycocyanin-B trimer with a linker polypeptide: analysis by the principal multi-component spectral estimation (PMSE) method.

    Science.gov (United States)

    Yamazaki, T; Nishimura, Y; Yamazaki, I; Hirano, M; Matsuura, K; Shimada, K; Mimuro, M

    1994-10-10

    Energy migration processes in allophycocyanin-B trimer with a linker polypeptide were analyzed using the principal multi-component spectral estimation (PMSE) method, which does not require assumption of component number, decay function, or the spectral band shape. We determined the number of spectral components showing independent kinetic behavior by the eigen-value of an auto-correlation matrix, and further the spectra of the components and their rise and decay curves. Two decay components were resolved at 20 degrees C: one corresponded to the decay of one type of beta-84 chromophore, and the other to the decay from the thermally equilibrated state between another type of beta-84 chromophore and the alpha-allophycocyanin B chromophore. An additional slow decay process was resolved at -196 degrees C. We also compared the component spectra obtained using the PMSE method with the decay-associated spectra obtained using the global analysis.

  12. Mixed-Stack Architecture and Solvatomorphism of Trimeric Perfluoro-ortho-Phenylene Mercury complexes with Dithieno[3,2-b:2',3'-d]thiophene

    KAUST Repository

    Castañeda, Raúl

    2015-08-01

    The formation of the mixed-stack donor-acceptor complex of dithieno[3,2-b:2\\',3\\'-d]thiophene (1) and trimeric perfluoro-ortho-phenylene mercury (I) has been investigated under different conditions. Two solvatomorphs – mixed-stack complexes with a 1:1 donor-acceptor ratio and different solvent molecules in the solid state (dichloromethane (2) and dichloroethane (3)) have been obtained and characterized by experimental methods (FT-IR spectroscopy, differential thermogravimetric analysis, and X-ray crystallography) and quantum-chemical calculations at the density functional theory level. The differences in the solid state packing, thermal stability and potential charge-transfer properties of 2 and 3 are discussed.

  13. A novel, live-attenuated vesicular stomatitis virus vector displaying conformationally intact, functional HIV-1 envelope trimers that elicits potent cellular and humoral responses in mice.

    Directory of Open Access Journals (Sweden)

    Svetlana Rabinovich

    Full Text Available Though vaccination with live-attenuated SIV provides the greatest protection from progressive disease caused by SIV challenge in rhesus macaques, attenuated HIV presents safety concerns as a vaccine; therefore, live viral vectors carrying HIV immunogens must be considered. We have designed a replication-competent vesicular stomatitis virus (VSV displaying immunogenic HIV-1 Env trimers and attenuating quantities of the native surface glycoprotein (G. The clade B Env immunogen is an Env-VSV G hybrid (EnvG in which the transmembrane and cytoplasmic tail regions are derived from G. Relocation of the G gene to the 5'terminus of the genome and insertion of EnvG into the natural G position induced a ∼1 log reduction in surface G, significant growth attenuation compared to wild-type, and incorporation of abundant EnvG. Western blot analysis indicated that ∼75% of incorporated EnvG was a mature proteolytically processed form. Flow cytometry showed that surface EnvG bound various conformationally- and trimer-specific antibodies (Abs, and in-vitro growth assays on CD4+CCR5+ cells demonstrated EnvG functionality. Neither intranasal (IN or intramuscular (IM administration in mice induced any observable pathology and all regimens tested generated potent Env-specific ELISA titers of 10(4-10(5, with an IM VSV prime/IN VSV boost regimen eliciting the highest binding and neutralizing Ab titers. Significant quantities of Env-specific CD4+ T cells were also detected, which were augmented as much as 70-fold by priming with IM electroporated plasmids encoding EnvG and IL-12. These data suggest that our novel vector can achieve balanced safety and immunogenicity and should be considered as an HIV vaccine platform.

  14. TprC/D (Tp0117/131), a Trimeric, Pore-Forming Rare Outer Membrane Protein of Treponema pallidum, Has a Bipartite Domain Structure

    Science.gov (United States)

    Anand, Arvind; Luthra, Amit; Dunham-Ems, Star; Caimano, Melissa J.; Karanian, Carson; LeDoyt, Morgan; Cruz, Adriana R.; Salazar, Juan C.

    2012-01-01

    Identification of Treponema pallidum rare outer membrane proteins (OMPs) has been a longstanding objective of syphilis researchers. We recently developed a consensus computational framework that employs a battery of cellular localization and topological prediction tools to generate ranked clusters of candidate rare OMPs (D. L. Cox et al., Infect. Immun. 78:5178–5194, 2010). TP0117/TP0131 (TprC/D), a member of the T. pallidum repeat (Tpr) family, was a highly ranked candidate. Circular dichroism, heat modifiability by SDS-PAGE, Triton X-114 phase partitioning, and liposome incorporation confirmed that full-length, recombinant TprC (TprCFl) forms a β-barrel capable of integrating into lipid bilayers. Moreover, TprCFl increased efflux of terbium-dipicolinic acid complex from large unilamellar vesicles and migrated as a trimer by blue-native PAGE. We found that in T. pallidum, TprC is heat modifiable, trimeric, expressed in low abundance, and, based on proteinase K accessibility and opsonophagocytosis assays, surface exposed. From these collective data, we conclude that TprC is a bona fide rare OMP as well as a functional ortholog of Escherichia coli OmpF. We also discovered that TprC has a bipartite architecture consisting of a soluble N-terminal portion (TprCN), presumably periplasmic and bound directly or indirectly to peptidoglycan, and a C-terminal β-barrel (TprCC). Syphilitic rabbits generate antibodies exclusively against TprCC, while secondary syphilis patients fail to mount a detectable antibody response against either domain. The syphilis spirochete appears to have resolved a fundamental dilemma arising from its extracellular lifestyle, namely, how to enhance OM permeability without increasing its vulnerability to the antibody-mediated defenses of its natural human host. PMID:22389487

  15. Unexpected Potency Differences between B-Cell-Activating Factor (BAFF) Antagonist Antibodies against Various Forms of BAFF: Trimer, 60-Mer, and Membrane-Bound.

    Science.gov (United States)

    Nicoletti, Amy M; Kenny, Cynthia Hess; Khalil, Ashraf M; Pan, Qi; Ralph, Kerry L M; Ritchie, Julie; Venkataramani, Sathyadevi; Presky, David H; DeWire, Scott M; Brodeur, Scott R

    2016-10-01

    Therapeutic agents antagonizing B-cell-activating factor/B-lymphocyte stimulator (BAFF/BLyS) are currently in clinical development for autoimmune diseases; belimumab is the first Food and Drug Administration-approved drug in more than 50 years for the treatment of lupus. As a member of the tumor necrosis factor superfamily, BAFF promotes B-cell survival and homeostasis and is overexpressed in patients with systemic lupus erythematosus and other autoimmune diseases. BAFF exists in three recognized forms: membrane-bound and two secreted, soluble forms of either trimeric or 60-mer oligomeric states. To date, most in vitro pharmacology studies of BAFF neglect one or more of these forms. Here, we report a comprehensive in vitro cell-based analysis of BAFF in assay systems that measure all forms of BAFF-mediated activation. We demonstrate the effects of these BAFF forms in both a primary human B-cell proliferation assay and in nuclear factor κB reporter assay systems in Chinese hamster ovary cells expressing BAFF receptors and transmembrane activator and calcium-modulator and cyclophilin ligand interactor (TACI). In contrast to the mouse system, we find that BAFF trimer activates the human TACI receptor. Further, we profiled the activities of two clinically advanced BAFF antagonist antibodies, belimumab and tabalumab. Unexpectedly, we revealed differences in inhibitory potencies against the various BAFF forms, in particular that belimumab does not potently inhibit BAFF 60-mer. Through this increased understanding of the activity of BAFF antagonists against different forms of BAFF, we hope to influence the discovery of BAFF antagonist antibodies with distinct therapeutic mechanisms for improvement in the treatment of lupus or other related autoimmune pathologies. Copyright © 2016 by The American Society for Pharmacology and Experimental Therapeutics.

  16. Single-Chain Soluble BG505.SOSIP gp140 Trimers as Structural and Antigenic Mimics of Mature Closed HIV-1 Env.

    Science.gov (United States)

    Georgiev, Ivelin S; Joyce, M Gordon; Yang, Yongping; Sastry, Mallika; Zhang, Baoshan; Baxa, Ulrich; Chen, Rita E; Druz, Aliaksandr; Lees, Christopher R; Narpala, Sandeep; Schön, Arne; Van Galen, Joseph; Chuang, Gwo-Yu; Gorman, Jason; Harned, Adam; Pancera, Marie; Stewart-Jones, Guillaume B E; Cheng, Cheng; Freire, Ernesto; McDermott, Adrian B; Mascola, John R; Kwong, Peter D

    2015-05-01

    Similar to other type I fusion machines, the HIV-1 envelope glycoprotein (Env) requires proteolytic activation; specifically, cleavage of a gp160 precursor into gp120 and gp41 subunits creates an N-terminal gp41 fusion peptide and permits folding from an immature uncleaved state to a mature closed state. While the atomic-level consequences of cleavage for HIV-1 Env are still being determined, the uncleaved state is antigenically distinct from the mature closed state, and cleavage has been reported to be essential for mimicry of the mature viral spike by soluble versions of Env. Here we report the redesign of a current state-of-the-art soluble Env mimic, BG505.SOSIP, to make it cleavage independent. Specifically, we replaced the furin cleavage site between gp120 and gp41 with Gly-Ser linkers of various lengths. The resultant linked gp120-gp41 constructs, termed single-chain gp140 (sc-gp140), exhibited different levels of structural and antigenic mimicry of the parent cleaved BG505.SOSIP. When constructs were subjected to negative selection to remove subspecies recognized by poorly neutralizing antibodies, trimers of high antigenic mimicry of BG505.SOSIP could be obtained; negative-stain electron microscopy indicated these to resemble the mature closed state. Higher proportions of BG505.SOSIP-trimer mimicry were observed in sc-gp140s with linkers of 6 or more residues, with a linker length of 15 residues exhibiting especially promising traits. Overall, flexible linkages between gp120 and gp41 in BG505.SOSIP can thus substitute for cleavage, and sc-gp140s that closely mimicked the vaccine-preferred mature closed state of Env could be obtained. The trimeric HIV-1 envelope glycoprotein (Env) is the sole target of virus-directed neutralizing antibody responses and a primary focus of vaccine design. Soluble mimics of Env have proven challenging to obtain and have been thought to require proteolytic cleavage into two-component subunits, gp120 and gp41, to achieve

  17. Comparison of biological properties of 99mTc-labeled cyclic RGD Peptide trimer and dimer useful as SPECT radiotracers for tumor imaging

    International Nuclear Information System (INIS)

    Zhao, Zuo-Quan; Yang, Yong; Fang, Wei; Liu, Shuang

    2016-01-01

    Introduction: This study sought to evaluate a 99m Tc-labeled trimeric cyclic RGD peptide ( 99m Tc-4P-RGD 3 ) as the new radiotracer for tumor imaging. The objective was to compare its biological properties with those of 99m Tc-3P-RGD 2 in the same animal model. Methods: HYNIC-4P-RGD 3 was prepared by reacting 4P-RGD 3 with excess HYNIC-OSu in the presence of diisopropylethylamine. 99m Tc-4P-RGD 3 was prepared using a kit formulation, and evaluated for its tumor-targeting capability and biodistribution properties in the BALB/c nude mice with U87MG human glioma xenografts. Planar and SPECT imaging studies were performed in athymic nude mice with U87MG glioma xenografts. For comparison purpose, 99m Tc-3P-RGD 2 (a α v β 3 -targeted radiotracer currently under clinical evaluation for tumor imaging in cancer patients) was also evaluated in the same animal models. Blocking experiments were used to demonstrate the α v β 3 specificity of 99m Tc-4P-RGD 3 . Results: 99m Tc-4P-RGD 3 was prepared with > 95% RCP and high specific activity (~ 200 GBq/μmol). 99m Tc-4P-RGD 3 and 99m Tc-3P-RGD 2 shared almost identical tumor uptake and similar biodistribution properties. 99m Tc-4P-RGD 3 had higher uptake than 99m Tc-3P-RGD 2 in the intestines and kidneys; but it showed better metabolic stability. The U87MG tumors were clearly visualized by SPECT with excellent contrast with 99m Tc-4P-RGD 3 and 99m Tc-3P-RGD 2 . Conclusion: Increasing peptide multiplicity from 3P-RGD 2 to 4P-RGD 3 offers no advantages with respect to the tumor-targeting capability. 99m Tc-4P-RGD 3 is as good a SPECT radiotracer as 99m Tc-3P-RGD 2 for imaging α v β 3 -positive tumors. -- Graphical abstract: This report presents evaluations of a 99m Tc-labeled cyclic RGD peptide trimer ( 99m Tc-4P-RGD 3 ) as the new SPECT radiotracer for tumor imaging. It was found that 99m Tc-4P-RGD 3 was able to accumulate in the xenografted U87MG tumors with high specificity. Display Omitted

  18. A CO32--containing, dimanganese-substituted silicotungstate trimer, K9[H14{SiW10MnIIMnIIIO38}3(CO3)]·39H2O.

    Science.gov (United States)

    Yang, Ling; Liu, Qisen; Ma, Pengtao; Niu, Jingyang; Wang, Jingping

    2015-08-14

    An unprecedented silicotungstate trimer K(9)[H(14){SiW(10)Mn(II)Mn(III)O(38)}(3)(CO(3))]·39H(2)O (1) has been successfully synthesized, in which the CO(3)(2-) resides inside the three Keggin {SiW(10)Mn(II)Mn(III)O(38)} units and the three O atoms serve as μ(2)-O atoms to connect with three Mn(III). Magnetic investigation indicates that 1 exhibits antiferromagnetic coupling.

  19. Introduction of a point mutation into an HLA class I single-chain trimer induces enhancement of CTL priming and antitumor immunity

    Directory of Open Access Journals (Sweden)

    Masanori Matsui

    2014-01-01

    Full Text Available We previously discovered one particular HLA-A*02:01 mutant that enhanced peptide-specific cytotoxic T lymphocyte (CTL recognition in vitro compared to wild-type HLA-A*02:01. This mutant contains a single amino acid substitution from histidine to leucine at position 74 (H74L that is located in the peptide-binding groove. To investigate the effect of the H74L mutation on the in vivo CTL priming, we took advantage of the technology of the HLA class I single-chain trimer (SCT in which three components involving a peptide, β2 microglobulin and the HLA class I heavy chain are joined together via flexible linkers. We generated recombinant adenovirus expressing SCT comprised influenza A matrix protein (FMP-derived peptide, β2 microglobulin and the H74L heavy chain. HLA-A*02:01 transgenic mice were immunized with the adenovirus, and the induction of peptide-specific CTLs and antitumor immunity was investigated. It was clearly shown that the H74L mutation enabled the HLA-A*02:01 SCT molecule to dramatically enhance both in vivo priming of FMP-specific CTLs and protection against a lethal challenge of tumor cells expressing FMP. These data present the first evidence that a simple point mutation in the HLA class I heavy chain of SCT is beneficial for improving CTL-based immunotherapy and prophylaxis to control tumors.

  20. Maturation of dendritic cells by recombinant human CD40L-trimer leads to a homogeneous cell population with enhanced surface marker expression and increased cytokine production

    DEFF Research Database (Denmark)

    Würtzen, P A; Nissen, Mogens Holst; Claesson, M H

    2001-01-01

    -cell activating capacity of the DC. We studied DC phenotype and cytokine production as well as the T-cell proliferation and cytotoxic T lympocyte (CTL) activation induced by DC generated in vitro. In addition, the effect of exposure to recombinant human CD40L-trimer (huCD40LT) on these parameters was investigated...... enhanced by exposure to huCD40LT even compared to TNF-alpha exposure. Only a moderate cytokine production was observed initially, while TNF-alpha addition or CD40 triggering, especially, induced enhanced production of IL-6 and IL-12 p40. Surprisingly, comparable induction of T-cell proliferation by a DC......Dendritic cells (DC) have been shown to be potent inducers of specific cytotoxic T-cell responses both in vivo and in vitro. Furthermore, exposure to cytokines such as tumour necrosis factor (TNF)-alpha or CD40 triggering changes DC phenotype and cytokine production and may enhance the T...

  1. Membrane-bound SIV envelope trimers are immunogenic in ferrets after intranasal vaccination with a replication-competent canine distemper virus vector.

    Science.gov (United States)

    Zhang, Xinsheng; Wallace, Olivia; Wright, Kevin J; Backer, Martin; Coleman, John W; Koehnke, Rebecca; Frenk, Esther; Domi, Arban; Chiuchiolo, Maria J; DeStefano, Joanne; Narpala, Sandeep; Powell, Rebecca; Morrow, Gavin; Boggiano, Cesar; Zamb, Timothy J; Richter King, C; Parks, Christopher L

    2013-11-01

    We are investigating canine distemper virus (CDV) as a vaccine vector for the delivery of HIV envelope (Env) that closely resembles the native trimeric spike. We selected CDV because it will promote vaccine delivery to lymphoid tissues, and because human exposure is infrequent, reducing potential effects of pre-existing immunity. Using SIV Env as a model, we tested a number of vector and gene insert designs. Vectors containing a gene inserted between the CDV H and L genes, which encoded Env lacking most of its cytoplasmic tail, propagated efficiently in Vero cells, expressed the immunogen on the cell surface, and incorporated the SIV glycoprotein into progeny virus particles. When ferrets were vaccinated intranasally, there were no signs of distress, vector replication was observed in the gut-associated lymphoid tissues, and the animals produced anti-SIV Env antibodies. These data show that live CDV-SIV Env vectors can safely induce anti-Env immune responses following intranasal vaccination. © 2013 Elsevier Inc. All rights reserved.

  2. Crystal structure of β1→6-galactosidase from Bifidobacterium bifidum S17: trimeric architecture, molecular determinants of the enzymatic activity and its inhibition by α-galactose.

    Science.gov (United States)

    Godoy, Andre Schutzer; Camilo, Cesar Moises; Kadowaki, Marco Antonio; Muniz, Heloisa Dos S; Espirito Santo, Melissa; Murakami, Mario Tyago; Nascimento, Alessandro S; Polikarpov, Igor

    2016-11-01

    In a search for better comprehension of β-galactosidase function and specificity, we solved the crystal structures of the GH42 β-galactosidase BbgII from Bifidobacterium bifidum S17, a well-adapted probiotic microorganism from the human digestive tract, and its complex with d-α-galactose. BbgII is a three-domain molecule that forms barrel-shaped trimers in solution. BbgII interactions with d-α-galactose, a competitive inhibitor, showed a number of residues that are involved in the coordination of ligands. A combination of site-directed mutagenesis of these amino acid residues with enzymatic activity measurements confirmed that Glu161 and Glu320 are fundamental for catalysis and their substitution by alanines led to catalytically inactive mutants. Mutation Asn160Ala resulted in a two orders of magnitude decrease of the enzyme k cat without significant modification in its K m , whereas mutations Tyr289Phe and His371Phe simultaneously decreased k cat and increased K m values. Enzymatic activity of Glu368Ala mutant was too low to be detected. Our docking and molecular dynamics simulations showed that the enzyme recognizes and tightly binds substrates with β1→6 and β1→3 bonds, while binding of the substrates with β1→4 linkages is less favorable. Structural data are available in the PDB under the accession numbers 4UZS and 4UCF. © 2016 Federation of European Biochemical Societies.

  3. Rotational spectra of the Xe-(H2O)2 van der Waals trimer: xenon as a probe of electronic structure and dynamics.

    Science.gov (United States)

    Wen, Qing; Jäger, Wolfgang

    2007-03-22

    Rotational spectra of three isotopomers of the Xe-(H2O)2 van der Waals trimer were recorded using a pulsed-nozzle, Fourier transform microwave spectrometer. Nine [nine, four] a-type and twelve [eleven, seven] b-type transitions were measured for the 132Xe-(H2O)2 [129Xe-(H2O)2, 131Xe-(H2O)2] isotopomer. The determined rotational and centrifugal distortion constants were used to extract information about the structure and vibrational motions of the complex. The nuclear quadrupole hyperfine structures due to the 131Xe (nuclear spin quantum number I=3/2) nucleus were also detected. The large value of the off-diagonal nuclear quadrupole coupling constant chiab in particular provides detailed insight into the electronic environment of the xenon atom and the orientations of the water molecules within the complex. An effective structure that best reproduces the experimental 131Xe nuclear quadrupole coupling constants is rationalized by ab initio calculations. An overall goal of this line of work is to determine how the successive solvation of a xenon atom with water molecules affects the xenon electron distribution and its intermolecular interactions. The results may provide molecular level interpretations of 129Xe NMR data from, for example, imaging experiments.

  4. Crystal structure of a trimeric archaeal adenylate kinase from the mesophile Methanococcus maripaludis with an unusually broad functional range and thermal stability.

    Science.gov (United States)

    Davlieva, Milya; Shamoo, Yousif

    2010-02-01

    The structure of the trimeric adenylate kinase from the Archaebacteria Methanococcus mariplaludis (AK(MAR)) has been solved to 2.5-A resolution and the temperature dependent stability and kinetics of the enzyme measured. The K(M) and V(max) of AK(MAR) exhibit only modest temperature dependence from 30 degrees -60 degrees C. Although M. mariplaludis is a mesophile with a maximum growth temperature of 43 degrees C, AK(MAR) has a very broad functional range and stability (T(m) = 74.0 degrees C) that are more consistent with a thermophilic enzyme with high thermostability and exceptional activity over a wide range of temperatures, suggesting that this microbe may have only recently invaded a mesophilic niche and has yet to fully adapt. A comparison of the Local Structural Entropy (LSE) for AK(MAR) to the related adenylate kinases from the mesophile Methanococcus voltae and thermophile Methanococcus thermolithotrophicus show that changes in LSE are able to fully account for the intermediate stability of AK(MAR) and highlights a general mechanism for protein adaptation in this class of enzymes. (c) 2009 Wiley-Liss, Inc.

  5. Structures, Thermochemical Properties, and Bonding of Mixed Alkaline-Earth-Metal Silicon Trimers Si3M(+/0/-) with M = Be, Mg, Ca.

    Science.gov (United States)

    Hang, Tran Dieu; Hung, Huynh Minh; Nguyen, Huyen Thi; Nguyen, Minh Tho

    2015-06-18

    The ground state geometries, electronic structures, and thermochemical properties of binary alkaline-earth-metal silicon clusters Si3M with M = Be, Mg, Ca in neutral, cationic, and anionic states were investigated using quantum chemical computations. Lowest-lying isomers of the clusters were determined on the basis of the composite G4 energies. Along with total atomization energies, thermochemical parameters were determined for the first time by means of the G4 and coupled-cluster theory with complete basis set CCSD(T)/CBS approaches. The most favored equilibrium formation sequences for Si3M clusters emerge as follows: all Si3M(+/0/-) clusters are formed by attaching the M atom into the corresponding cation, neutral and anion silicon trimer Si3(+/0/-), except for the Si3Mg(+) and Si3Ca(+) where the metal cations are bound to the neutral Si3. The resulting mixed tetramers exhibit geometrical and electronic features similar to those of the pure silicon tetramer Si4(+/0/-). Electron localization function (ELF) and ring current analyses point out that the σ-aromatic character of silicon tetramer remains unchanged upon substituting one Si atom by one alkaline-earth-metal atom.

  6. Defect induced intermittency in the transit time dynamics generates 1/f noise in a trimer described by the discrete nonlinear Schroedinger equation

    International Nuclear Information System (INIS)

    Pando L, C.L.; Doedel, E.J.

    2006-08-01

    We investigate the nonlinear dynamics in a trimer, described by the one-dimensional discrete nonlinear Schrodinger equation (DNLSE), with periodic boundary conditions in the presence of a single on-site defect. We make use of numerical continuation to study different families of stationary and periodic solutions, which allows us to consider suitable perturbations. Taking into account a Poincare section, we are able to study the dynamics in both a thin stochastic layer solution and a global stochasticity solution. We find that the time series of the transit times, the time intervals to traverse some suitable sets in phase space, generate 1/f noise for both stochastic solutions. In the case of the thin stochastic layer solution, we find that transport between two almost invariant sets along with intermittency in small and large time scales are relevant features of the dynamics. These results are reflected in the behaviour of the standard map with suitable parameters. In both chaotic solutions, the distribution of transit times has a maximum and a tail with exponential decay in spite of the presence of long-range correlations in the time series. We motivate our study by considering a ring of weakly-coupled Bose-Einstein condensates (BEC) with attractive interactions, where inversion of populations between two spatially symmetric sites and phase locking take place in both chaotic solutions. (author)

  7. Combinatorial control of adhesion of Brucella abortus 2308 to host cells by transcriptional rewiring of the trimeric autotransporter btaE gene.

    Science.gov (United States)

    Sieira, Rodrigo; Bialer, Magalí G; Roset, Mara S; Ruiz-Ranwez, Verónica; Langer, Tomás; Arocena, Gastón M; Mancini, Estefanía; Zorreguieta, Angeles

    2017-02-01

    Regulatory network plasticity is a key attribute underlying changes in bacterial gene expression and a source of phenotypic diversity to interact with the surrounding environment. Here, we sought to study the transcriptional circuit of HutC, a regulator of both metabolic and virulence genes of the facultative intracellular pathogen Brucella. Using in silico and biochemical approaches, we identified a novel functional HutC-binding site upstream of btaE, a trimeric-autotransporter adhesin involved in the attachment of Brucella to host extracellular matrix components. Moreover, we identified two additional regulators, one of which, MdrA, acts in concert with HutC to exert a combinatorial control of both btaE promoter activity and attachment of Brucella to HeLa cells. Analysis of btaE promoter sequences of different species indicated that this HutC-binding site was generated de novo by a single point mutation in a virulent Brucella strain, indicative of a transcriptional rewiring event. In addition to major domain organization differences existing between BtaE proteins within the genus Brucella, our analyses revealed that sequences upstream of btaE display high variability probably associated to intrinsic promoter structural features, which may serve as a substrate for reciprocal selection during co-evolution between this pathogen and its mammalian host. © 2016 John Wiley & Sons Ltd.

  8. Maturation of dendritic cells by recombinant human CD40L-trimer leads to a homogeneous cell population with enhanced surface marker expression and increased cytokine production

    DEFF Research Database (Denmark)

    Würtzen, P A; Nissen, Mogens Holst; Claesson, M H

    2001-01-01

    Dendritic cells (DC) have been shown to be potent inducers of specific cytotoxic T-cell responses both in vivo and in vitro. Furthermore, exposure to cytokines such as tumour necrosis factor (TNF)-alpha or CD40 triggering changes DC phenotype and cytokine production and may enhance the T-cell act...... marker expression and high production of pro-inflammatory cytokines. In addition, the induction of responses to allo or recall antigens presented by huCD40LT maturated DC was comparable to the responses obtained with the DC maturated through TNF-alpha exposure.......Dendritic cells (DC) have been shown to be potent inducers of specific cytotoxic T-cell responses both in vivo and in vitro. Furthermore, exposure to cytokines such as tumour necrosis factor (TNF)-alpha or CD40 triggering changes DC phenotype and cytokine production and may enhance the T......-cell activating capacity of the DC. We studied DC phenotype and cytokine production as well as the T-cell proliferation and cytotoxic T lympocyte (CTL) activation induced by DC generated in vitro. In addition, the effect of exposure to recombinant human CD40L-trimer (huCD40LT) on these parameters was investigated...

  9. The molecular dynamic study of anharmonic effects at Cu(111) and Ag(111) surfaces in the presence of Cu- and Ag-trimer island

    International Nuclear Information System (INIS)

    Shah, Zulfiqar Ali; Hayat, Sardar Sikandar; Rehman, Z.; Bouafia, Farida

    2014-01-01

    The molecular dynamics (MD) technique based on semi-empirical potentials, is used to carry out the diffusion of Cu- and Ag-trimer on Cu- and Ag(111) surface at 300, 500 and 700 K temperatures. The constant energy MD simulation elaborates the anharmonic effects at the surface such as fissures, dislocations and vacancy creation, in the presence of island. The fissures and dislocations formed are in the range of 1.5–4 Å and 1–7 Å, respectively, from the island's position. The Cu and Ag islands both diffuse easily on Cu(111) surface, manipulate that the trend of diffusion is faster on Cu surface as compared to Ag surface. The process of breaking and opening of the island has also been observed. Moreover, a surface atom popped-up at 700 K by creating a vacancy near the Cu island on Ag surface. The rate of diffusion increases with the increase in temperature, both for homo- and hetero-cases.

  10. Structure-function analyses of a pertussis-like toxin from pathogenicEscherichia colireveal a distinct mechanism of inhibition of trimeric G-proteins.

    Science.gov (United States)

    Littler, Dene R; Ang, Sheng Y; Moriel, Danilo G; Kocan, Martina; Kleifeld, Oded; Johnson, Matthew D; Tran, Mai T; Paton, Adrienne W; Paton, James C; Summers, Roger J; Schembri, Mark A; Rossjohn, Jamie; Beddoe, Travis

    2017-09-08

    Pertussis-like toxins are secreted by several bacterial pathogens during infection. They belong to the AB 5 virulence factors, which bind to glycans on host cell membranes for internalization. Host cell recognition and internalization are mediated by toxin B subunits sharing a unique pentameric ring-like assembly. Although the role of pertussis toxin in whooping cough is well-established, pertussis-like toxins produced by other bacteria are less studied, and their mechanisms of action are unclear. Here, we report that some extra-intestinal Escherichia coli pathogens ( i.e. those that reside in the gut but can spread to other bodily locations) encode a pertussis-like toxin that inhibits mammalian cell growth in vitro We found that this protein, Ec Plt, is related to toxins produced by both nontyphoidal and typhoidal Salmonella serovars. Pertussis-like toxins are secreted as disulfide-bonded heterohexamers in which the catalytic ADP-ribosyltransferase subunit is activated when exposed to the reducing environment in mammalian cells. We found here that the reduced Ec Plt exhibits large structural rearrangements associated with its activation. We noted that inhibitory residues tethered within the NAD + -binding site by an intramolecular disulfide in the oxidized state dissociate upon the reduction and enable loop restructuring to form the nucleotide-binding site. Surprisingly, although pertussis toxin targets a cysteine residue within the α subunit of inhibitory trimeric G-proteins, we observed that activated Ec Plt toxin modifies a proximal lysine/asparagine residue instead. In conclusion, our results reveal the molecular mechanism underpinning activation of pertussis-like toxins, and we also identified differences in host target specificity. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  11. Amino acid duplication in the coiled-coil structure of collagen XVII alters its maturation and trimerization causing mild junctional epidermolysis bullosa.

    Science.gov (United States)

    Kroeger, Jasmin K; Hofmann, Silke C; Leppert, Juna; Has, Cristina; Franzke, Claus-Werner

    2017-02-01

    The function and stability of collagens depend on the accurate triple helix formation of three distinct polypeptide chains. Disruption of this triple-helical structure can result in connective-tissue disorders. Triple helix formation is thought to depend on three-stranded coiled-coil oligomerization sites within non-collagenous domains. However, only little is known about the physiological relevance of these coiled-coil structures. Transmembrane collagen XVII, also known as 180 kDa bullous pemphigoid antigen provides mechanical stability through the anchorage of epithelial cells to the basement membrane. Mutations in the collagen XVII gene, COL17A1, cause junctional epidermolysis bullosa (JEB), characterized by chronic trauma-induced skin blistering. Here we exploited a novel naturally occurring COL17A1 mutation, leading to an in-frame lysine duplication within the coiled-coil structure of the juxtamembranous NC16A domain of collagen XVII, which resulted in a mild phenotype of JEB due to reduced membrane-anchored collagen XVII molecules. This mutation causes structural changes in the mutant molecule and interferes with its maturation. The destabilized coiled-coil structure of the mutant collagen XVII unmasks a furin cleavage site that results in excessive and non-physiological ectodomain shedding during its maturation. Furthermore, it decreases its triple-helical stability due to defective coiled-coil oligomerization, which makes it highly susceptible to proteolytic degradation. As a consequence of altered maturation and decreased stability of collagen XVII trimers, reduced collagen XVII is incorporated into the cell membrane, resulting in compromised dermal-epidermal adhesion. Taken together, using this genetic model, we provide the first proof that alteration of the coiled-coil structure destabilizes oligomerization and impairs physiological shedding of collagen XVII in vivo. © The Author 2016. Published by Oxford University Press. All rights reserved. For

  12. Functional characterization of two scFv-Fc antibodies from an HIV controller selected on soluble HIV-1 Env complexes: a neutralizing V3- and a trimer-specific gp41 antibody.

    Science.gov (United States)

    Trott, Maria; Weiβ, Svenja; Antoni, Sascha; Koch, Joachim; von Briesen, Hagen; Hust, Michael; Dietrich, Ursula

    2014-01-01

    HIV neutralizing antibodies (nAbs) represent an important tool in view of prophylactic and therapeutic applications for HIV-1 infection. Patients chronically infected by HIV-1 represent a valuable source for nAbs. HIV controllers, including long-term non-progressors (LTNP) and elite controllers (EC), represent an interesting subgroup in this regard, as here nAbs can develop over time in a rather healthy immune system and in the absence of any therapeutic selection pressure. In this study, we characterized two particular antibodies that were selected as scFv antibody fragments from a phage immune library generated from an LTNP with HIV neutralizing antibodies in his plasma. The phage library was screened on recombinant soluble gp140 envelope (Env) proteins. Sequencing the selected peptide inserts revealed two major classes of antibody sequences. Binding analysis of the corresponding scFv-Fc derivatives to various trimeric and monomeric Env constructs as well as to peptide arrays showed that one class, represented by monoclonal antibody (mAb) A2, specifically recognizes an epitope localized in the pocket binding domain of the C heptad repeat (CHR) in the ectodomain of gp41, but only in the trimeric context. Thus, this antibody represents an interesting tool for trimer identification. MAb A7, representing the second class, binds to structural elements of the third variable loop V3 and neutralizes tier 1 and tier 2 HIV-1 isolates of different subtypes with matching critical amino acids in the linear epitope sequence. In conclusion, HIV controllers are a valuable source for the selection of functionally interesting antibodies that can be selected on soluble gp140 proteins with properties from the native envelope spike.

  13. Functional characterization of two scFv-Fc antibodies from an HIV controller selected on soluble HIV-1 Env complexes: a neutralizing V3- and a trimer-specific gp41 antibody.

    Directory of Open Access Journals (Sweden)

    Maria Trott

    Full Text Available HIV neutralizing antibodies (nAbs represent an important tool in view of prophylactic and therapeutic applications for HIV-1 infection. Patients chronically infected by HIV-1 represent a valuable source for nAbs. HIV controllers, including long-term non-progressors (LTNP and elite controllers (EC, represent an interesting subgroup in this regard, as here nAbs can develop over time in a rather healthy immune system and in the absence of any therapeutic selection pressure. In this study, we characterized two particular antibodies that were selected as scFv antibody fragments from a phage immune library generated from an LTNP with HIV neutralizing antibodies in his plasma. The phage library was screened on recombinant soluble gp140 envelope (Env proteins. Sequencing the selected peptide inserts revealed two major classes of antibody sequences. Binding analysis of the corresponding scFv-Fc derivatives to various trimeric and monomeric Env constructs as well as to peptide arrays showed that one class, represented by monoclonal antibody (mAb A2, specifically recognizes an epitope localized in the pocket binding domain of the C heptad repeat (CHR in the ectodomain of gp41, but only in the trimeric context. Thus, this antibody represents an interesting tool for trimer identification. MAb A7, representing the second class, binds to structural elements of the third variable loop V3 and neutralizes tier 1 and tier 2 HIV-1 isolates of different subtypes with matching critical amino acids in the linear epitope sequence. In conclusion, HIV controllers are a valuable source for the selection of functionally interesting antibodies that can be selected on soluble gp140 proteins with properties from the native envelope spike.

  14. Advantages of a validated UPLC-MS/MS standard addition method for the quantification of A-type dimeric and trimeric proanthocyanidins in cranberry extracts in comparison with well-known quantification methods.

    Science.gov (United States)

    van Dooren, Ines; Foubert, Kenn; Theunis, Mart; Naessens, Tania; Pieters, Luc; Apers, Sandra

    2018-01-30

    The berries of Vaccinium macrocarpon, cranberry, are widely used for the prevention of urinary tract infections. This species contains A-type proanthocyanidins (PACs), which intervene in the initial phase of the development of urinary tract infections by preventing the adherence of Escherichia coli by their P-type fimbriae to uroepithelial cells. Unfortunately, the existing clinical studies used different cranberry preparations, which were poorly standardized. Because of this, the results were hard to compare, which led sometimes to conflicting results. Currently, PACs are quantified using the rather non-specific spectrophotometric 4-dimethylaminocinnamaldehyde (DMAC) method. In addition, a normal phase HPTLC-densitometric method, a HPLC-UV method and three LC-MS/MS methods for quantification of procyanidin A2 were recently published. All these methods contain some shortcomings and errors. Hence, the development and validation of a fast and sensitive standard addition LC-MS/MS method for the simultaneous quantification of A-type dimers and trimers in a cranberry dry extract was carried out. A linear calibration model could be adopted for dimers and, after logaritmic transformation, for trimers. The maximal interday and interconcentration precision was found to be 4.86% and 4.28% for procyanidin A2, and 5.61% and 7.65% for trimeric PACs, which are all acceptable values for an analytical method using LC-MS/MS. In addition, twelve different cranberry extracts were analyzed by means of the newly validated method and other widely used methods. There appeared to be an enormous variation in dimeric and trimeric PAC content. Comparison of these results with LC-MS/MS analysis without standard addition showed the presence of matrix effects for some of the extracts and proved the necessity of standard addition. A comparison of the well-known and widely used DMAC method, the butanol-HCl assay and this newly developed LC-MS/MS method clearly indicated the need for a reliable

  15. Bartonella henselae trimeric autotransporter adhesin BadA expression interferes with effector translocation by the VirB/D4 type IV secretion system.

    Science.gov (United States)

    Lu, Yun-Yueh; Franz, Bettina; Truttmann, Matthias C; Riess, Tanja; Gay-Fraret, Jérémie; Faustmann, Marco; Kempf, Volkhard A J; Dehio, Christoph

    2013-05-01

    The Gram-negative, zoonotic pathogen Bartonella henselae is the aetiological agent of cat scratch disease, bacillary angiomatosis and peliosis hepatis in humans. Two pathogenicity factors of B. henselae - each displaying multiple functions in host cell interaction - have been characterized in greater detail: the trimeric autotransporter Bartonella adhesin A (BadA) and the type IV secretion system VirB/D4 (VirB/D4 T4SS). BadA mediates, e.g. binding to fibronectin (Fn), adherence to endothelial cells (ECs) and secretion of vascular endothelial growth factor (VEGF). VirB/D4 translocates several Bartonella effector proteins (Beps) into the cytoplasm of infected ECs, resulting, e.g. in uptake of bacterial aggregates via the invasome structure, inhibition of apoptosis and activation of a proangiogenic phenotype. Despite this knowledge of the individual activities of BadA or VirB/D4 it is unknown whether these major virulence factors affect each other in their specific activities. In this study, expression and function of BadA and VirB/D4 were analysed in a variety of clinical B. henselae isolates. Data revealed that most isolates have lost expression of either BadA or VirB/D4 during in vitro passages. However, the phenotypic effects of coexpression of both virulence factors was studied in one clinical isolate that was found to stably coexpress BadA and VirB/D4, as well as by ectopic expression of BadA in a strain expressing VirB/D4 but not BadA. BadA, which forms a dense layer on the bacterial surface, negatively affected VirB/D4-dependent Bep translocation and invasome formation by likely preventing close contact between the bacterial cell envelope and the host cell membrane. In contrast, BadA-dependent Fn binding, adhesion to ECs and VEGF secretion were not affected by a functional VirB/D4 T4SS. The obtained data imply that the essential virulence factors BadA and VirB/D4 are likely differentially expressed during different stages of the infection cycle of

  16. Protective efficacy of Newcastle disease virus expressing soluble trimeric hemagglutinin against highly pathogenic H5N1 influenza in chickens and mice.

    Directory of Open Access Journals (Sweden)

    Lisette A H M Cornelissen

    Full Text Available BACKGROUND: Highly pathogenic avian influenza virus (HPAIV causes a highly contagious often fatal disease in poultry, resulting in significant economic losses in the poultry industry. HPAIV H5N1 also poses a major public health threat as it can be transmitted directly from infected poultry to humans. One effective way to combat avian influenza with pandemic potential is through the vaccination of poultry. Several live vaccines based on attenuated Newcastle disease virus (NDV that express influenza hemagglutinin (HA have been developed to protect chickens or mammalian species against HPAIV. However, the zoonotic potential of NDV raises safety concerns regarding the use of live NDV recombinants, as the incorporation of a heterologous attachment protein may result in the generation of NDV with altered tropism and/or pathogenicity. METHODOLOGY/PRINCIPAL FINDINGS: In the present study we generated recombinant NDVs expressing either full length, membrane-anchored HA of the H5 subtype (NDV-H5 or a soluble trimeric form thereof (NDV-sH5(3. A single intramuscular immunization with NDV-sH5(3 or NDV-H5 fully protected chickens against disease after a lethal challenge with H5N1 and reduced levels of virus shedding in tracheal and cloacal swabs. NDV-sH5(3 was less protective than NDV-H5 (50% vs 80% protection when administered via the respiratory tract. The NDV-sH5(3 was ineffective in mice, regardless of whether administered oculonasally or intramuscularly. In this species, NDV-H5 induced protective immunity against HPAIV H5N1, but only after oculonasal administration, despite the poor H5-specific serum antibody response it elicited. CONCLUSIONS/SIGNIFICANCE: Although NDV expressing membrane anchored H5 in general provided better protection than its counterpart expressing soluble H5, chickens could be fully protected against a lethal challenge with H5N1 by using the latter NDV vector. This study thus provides proof of concept for the use of recombinant

  17. Self-assembly of linear [Mn II 2 Mn III ] units with end-on azido bridges: the construction of a ferromagnetic chain using S T = 7 high-spin trimers

    KAUST Repository

    Jiang, Yuan

    2015-01-01

    © The Royal Society of Chemistry 2015. The controlled organization of high-spin complexes into 1D coordination polymers is a challenge in molecular magnetism. In this work, we report a ferromagnetic Mn trimer Mn3(HL)2(CH3OH)6(Br)4·Br·(CH3OH)21 (H2L = 2-[(9H-fluoren-9-yl)amino]propane-1,3-diol) with the ground spin state of ST = 7 that can be assembled into a one-dimensional coordination chain [Mn3(HL)2(CH3OH)2(Br)4(N3)(H2O)·CH3OH]∞2 using azido bridging ligands. Interestingly, the ferromagnetic nature of 1 is well retained in 2. However, due to the negligible magnetic anisotropy in 1, both 1 and 2 do not show slow-relaxation of magnetization, which indicates that during the process of molecular assembly not only the intratrimer magnetic interaction but also the magnetic anisotropy of the trimer can be reserved.

  18. Self-assembly of linear [Mn(II)2Mn(III)] units with end-on azido bridges: the construction of a ferromagnetic chain using ST = 7 high-spin trimers.

    Science.gov (United States)

    Jiang, Yuan; Qin, Lei; Li, Guanghua; Abbas, Ghulam; Cao, Yaqun; Wu, Gang; Han, Tian; Zheng, Yan-Zhen; Qiu, Shilun

    2015-03-21

    The controlled organization of high-spin complexes into 1D coordination polymers is a challenge in molecular magnetism. In this work, we report a ferromagnetic Mn trimer Mn3(HL)2(CH3OH)6(Br)4·Br·(CH3OH)2 1 (H2L = 2-[(9H-fluoren-9-yl)amino]propane-1,3-diol) with the ground spin state of ST = 7 that can be assembled into a one-dimensional coordination chain [Mn3(HL)2(CH3OH)2(Br)4(N3)(H2O)·CH3OH]∞ 2 using azido bridging ligands. Interestingly, the ferromagnetic nature of 1 is well retained in 2. However, due to the negligible magnetic anisotropy in 1, both 1 and 2 do not show slow-relaxation of magnetization, which indicates that during the process of molecular assembly not only the intratrimer magnetic interaction but also the magnetic anisotropy of the trimer can be reserved.

  19. Zinc chloride - biuret - water and cadmium chloride - biuret - water systems at 30 deg C

    International Nuclear Information System (INIS)

    Shyjtyeva, N.; Davranov, M.; Sulajmankulov, K.

    1983-01-01

    By the solubility method the systems ZnCl 2 -C 2 H 5 N 3 O 2 -H 2 O and CdCl 2 -C 2 H 5 N 3 O 2 -H 2 O are studied at 30 deg. It has been found in the systems a formation of incongruently soluble compounds: ZnCl 2 x2C 2 H 5 N 3 O 2 , ZnCl 2 xC 2 H 5 N 3 O 2 , 2ZnCl 2 xC 2 H 5 N 3 O 2 , CdCl 2 x1.5C 2 H 5 N 3 O 2 and congruently soluble compound CdCl 2 x2C 2 H 5 N 3 O 2

  20. Zinc chloride - biuret - water and cadmium chloride - biuret - water systems at 30 deg C

    Energy Technology Data Exchange (ETDEWEB)

    Shyjtyeva, N.; Davranov, M.; Sulajmankulov, K.

    1983-01-01

    By the solubility method the systems ZnCl/sub 2/-C/sub 2/H/sub 5/N/sub 3/O/sub 2/-H/sub 2/O and CdCl/sub 2/-C/sub 2/H/sub 5/N/sub 3/O/sub 2/-H/sub 2/O are studied at 30 deg. It has been found in the systems a formation of incongruently soluble compounds: ZnCl/sub 2/x2C/sub 2/H/sub 5/N/sub 3/O/sub 2/, ZnCl/sub 2/xC/sub 2/H/sub 5/N/sub 3/O/sub 2/, 2ZnCl/sub 2/xC/sub 2/H/sub 5/N/sub 3/O/sub 2/, CdCl/sub 2/x1.5C/sub 2/H/sub 5/N/sub 3/O/sub 2/ and congruently soluble compound CdCl/sub 2/x2C/sub 2/H/sub 5/N/sub 3/O/sub 2/.

  1. Single-ion anisotropy and exchange interactions in the cyano-bridged trimers MnIII2MIII(CN)6 (MIII = Co, Cr, Fe) species incorporating [Mn(5-Brsalen)]+ units: an inelastic neutron scattering and magnetic susceptibility study

    DEFF Research Database (Denmark)

    Tregenna-Piggott, Philip L W; Sheptyakov, Denis; Keller, Lukas

    2009-01-01

    expectations based on the unquenched orbital angular momentum of the [Fe(CN)(6)](3-) anion, giving rise to an M(s) approximately +/-9/2 ground state, isolated by approximately 11.5 cm(-1) from the higher-lying levels. The reported INS and magnetic data should now serve as a benchmark against which theoretical...... interactions that define the low-lying states of the Mn-M(III)-Mn trimeric units. Despite the presence of an antiferromagnetic intertrimer interaction, the experimental evidence supports the classification of both the Cr(III) and Fe(III) compounds as single-molecule magnets. The value of 17(2) cm(-1...

  2. Magnetism of the spin-trimer compound CaNi 3(P 2O 7)2: Microscopic insight from combined 31P NMR and first-principles studies

    Science.gov (United States)

    Majumder, M.; Kanungo, S.; Ghoshray, A.; Ghosh, M.; Ghoshray, K.

    2015-03-01

    Magnetization, 31P nuclear magnetic resonance study, and first-principles electronic structure calculations have been performed in the spin-1 trimer chain compound CaNi3(P2O7 )2. Two separate spectra arising from magnetically and crystallographically inequivalent P sites are observed. In the ordered state, the resonance lines for both the P sites (P1 and P2) are found to be split into two, which is clear microscopic evidence of the development of two-sublattice AFM order below TM. A nonnegligible contribution of ferromagnetic hyperfine field and dipolar field have also been seen in the ordered state. The first-principles calculations show that the intratrimer (J1) and intertrimer interactions (J2) are of weak ferromagnetic type with the values 2.85 and 1.49 meV, respectively, whereas the interchain interaction (J3) is of strong antiferromagnetic type with a value of 5.63 meV. The anisotropy of the imaginary part of dynamical spin susceptibility around TM along with the exponential decrement of 1 /T1 below TM indicate the probable participation of the Ni -3 d electron's orbital degrees of freedom in the ferrimagnetic transition. The dominance of orbital fluctuations over the spin fluctuations seems to be responsible for showing low value of the binding energy u of the local spin configuration (estimated from local spin models) and an unusually weak exponent in the power-law behavior of 1 /T1 below 50 K, in the paramagnetic state. Electronic structure calculations also reveal the importance of orbital degrees of freedom of Ni -3 d moments, which is consistent with our NMR data analysis.

  3. Combined Cytolytic Effects of a Vaccinia Virus Encoding a Single Chain Trimer of MHC-I with a Tax-Epitope and Tax-Specific CTLs on HTLV-I-Infected Cells in a Rat Model

    Directory of Open Access Journals (Sweden)

    Takashi Ohashi

    2014-01-01

    Full Text Available Adult T cell leukemia (ATL is a malignant lymphoproliferative disease caused by human T cell leukemia virus type I (HTLV-I. To develop an effective therapy against the disease, we have examined the oncolytic ability of an attenuated vaccinia virus (VV, LC16m8Δ (m8Δ, and an HTLV-I Tax-specific cytotoxic T lymphocyte (CTL line, 4O1/C8, against an HTLV-I-infected rat T cell line, FPM1. Our results demonstrated that m8Δ was able to replicate in and lyse tumorigenic FPM1 cells but was incompetent to injure 4O1/C8 cells, suggesting the preferential cytolytic activity toward tumor cells. To further enhance the cytolysis of HTLV-I-infected cells, we modified m8Δ and obtained m8Δ/RT1AlSCTax180L, which can express a single chain trimer (SCT of rat major histocompatibility complex class I with a Tax-epitope. Combined treatment with m8Δ/RT1AlSCTax180L and 4O1/C8 increased the cytolysis of FPM1V.EFGFP/8R cells, a CTL-resistant subclone of FPM1, compared with that using 4O1/C8 and m8Δ presenting an unrelated peptide, suggesting that the activation of 4O1/C8 by m8Δ/RT1AlSCTax180L further enhanced the killing of the tumorigenic HTLV-I-infected cells. Our results indicate that combined therapy of oncolytic VVs with SCTs and HTLV-I-specific CTLs may be effective for eradication of HTLV-I-infected cells, which evade from CTL lysis and potentially develop ATL.

  4. Guidelines for standard and biuretic renogram in children

    International Nuclear Information System (INIS)

    Gordon, I.; Piepsz, A.; Colarinha, P.; Hahn, K.; Fischer, S.; Porn, U.; Sixt, R.; Velzen, J. van

    2000-01-01

    The guidelines are intended to help nuclear medical teams in their daily routine. The information given relates to aspects such as data acquisition, evaluation and interpretation, and indications for pediatric renal functional scintigraphy. The guidelines have been elaborated in response to a request of EANM and the American Society of Nuclear Medicine, who expressed the need for guidelines on recommended procedures for most of the standard nuclear medical examinations. The guidelines express the opinion of the Paediatric Committee of the EANM, and should be seen in the context of generally accepted basic principles in nuclear medicine, as well as local and national regulatory standards in radiation protection. (orig./CB) [de

  5. A monoclinic polymorph of 1-benzoyl-4-thio-biuret.

    Science.gov (United States)

    Kim, Namhun; Kim, Haneol; Kang, Sung Kwon

    2013-11-13

    The title compound, C9H9N3O2S, is a monoclinic (C2/c) polymorph of the previously reported triclinic structure [Kang (2013 ▶). Acta Cryst. E69, o1327]. The mol-ecule is almost planar with an r.m.s. deviation of 0.069 Å from the mean plane of all non-H atoms. The benzoyl and terminal thio-urea fragments adopt a transoid conformation with respect to the central carbonyl O atom. Two intra-molecular N-H⋯O hydrogen bonds are present. In the crystal, N-H⋯O and N-H⋯S inter-actions link the mol-ecules into zigzag chains extending along the c-axis direction.

  6. Determination of biuret in agroindustrial urea by spectrophotometry

    OpenAIRE

    Ferreira, R. B.; Franzini, V. P.; Gomes Neto, J. A.

    2007-01-01

    Foi desenvolvido um método espectrofotométrico para a determinação de biureto em fertilizantes à base de uréia empregando Cu(OH)2(s) como reagente analítico. O método se baseia na reação entre biureto e Cu(OH)2(s) e formação do complexo Cu(C2N3O2H5)2 de coloração violeta cujo espectro apresenta máximo de absorção em 510 nm. Após o estabelecimento das melhores condições analíticas, o método proposto foi aplicado na determinação de biureto em duas amostras comerciais de uréia, uma para pecuária...

  7. Trimerization process of vegetables oils and fats and its application in additives for synthetic fluids used in the perforation of petroleum well bore; Processo de trimerizacao de oleos vegetais e acidos graxos e sua aplicacao em aditivos para fluidos sinteticos utilizados na perfuracao de pocos de petroleo

    Energy Technology Data Exchange (ETDEWEB)

    Moreira, Viviane X.; Souza, Roberta C.R.; Coelho, Anderson S.; Lopes, Grazielle; Yoshida, Renan O.; Goncalves, Gizelle de Fatima G.D.V.; Silva, Sandra Regina da [Centro Tecnico Aeroespacial (CTA), Sao Jose dos Campos, SP (Brazil). Centro de Tecnologia; Poland Quimica Ltda., Macae, RJ (Brazil)

    2007-07-01

    Derivatives of vegetable oils and fatty acids such epoxides, polyols and fatty acid dimmers are industrially used for a variety of polymer applications, such as additives for plastics and composites and in filtration control aid in in perforation fluids as well as in the synthesis of polyamides and polyurethanes. In this article a vegetable oil was chemically modified by trimerization and used as filtration control and in a paraffin based perforation fluid. The rheology and filtration parameters were studied and the results were compared to desired specifications to maintenance of stability of well bore. (author)

  8. Contact allergy to aliphatic polyisocyanates based on hexamethylene-1,6-diisocyanate (HDI).

    Science.gov (United States)

    Aalto-Korte, Kristiina; Pesonen, Maria; Kuuliala, Outi; Alanko, Kristiina; Jolanki, Riitta

    2010-12-01

    Aliphatic polyisocyanates based on hexamethylene 1,6-diisocyanate (HDI) are components of lacquers, coatings, and spray paints. They are mainly composed of HDI trimers, but also contain larger oligomers, and minute amounts of HDI monomers (paints. The patients were examined at the Finnish Institute of Occupational Health in 2000-2009. The first two patients were diagnosed by testing with their own polyurethane hardeners and ingredients of the hardeners. In 2002, HDI isocyanurate (HDI-IC) trimer was added to our isocyanate series, and the last two patients were screened with the series. Patient no. 1 was exposed and sensitized to HDI biuret trimer, patient no. 2 to HDI-IC trimer, and patient no. 4 to HDI asymmetrical trimer. Patient no. 3 was positive with HDI-IC trimer. He had been handling several paint hardeners containing HDI-based polyisocyanates, but the subtypes of the trimers remained unidentified. All 4 patients were negative with HDI monomer. HDI trimers are novel contact allergens in workers who handle polyurethane paints. The allergic reactions cannot be explained by sensitization to HDI monomer. © 2010 John Wiley & Sons A/S.

  9. Monomeric, trimeric, and tetrameric transition metal complexes (Mn, Fe, Co) containing N,N-bis(2-pyridylmethyl)-2-aminoethanol/-ate: preparation, crystal structure, molecular magnetism and oxidation catalysis.

    Science.gov (United States)

    Shin, Jong Won; Rowthu, Sankara Rao; Hyun, Min Young; Song, Young Joo; Kim, Cheal; Kim, Bong Gon; Min, Kil Sik

    2011-06-07

    The reaction of N,N-bis(2-pyridylmethyl)-2-aminoethanol (bpaeOH), NaSCN/NaN(3), and metal (M) ions [M = Mn(II), Fe(II/III), Co(II)] in MeOH, leads to the isolation of a series of monomeric, trimeric, and tetrameric metal complexes, namely [Mn(bpaeOH)(NCS)(2)] (1), [Mn(bpaeO)(N(3))(2)] (2), [Fe(bpaeOH)(NCS)(2)] (3), [Fe(4)(bpaeO)(2)(CH(3)O)(2)(N(3))(8)] (4), [Co(bpaeOH)(NCS)(2)] (5), and [Co(3)(bpaeO)(2)(NO(3))(N(3))(4)](NO(3)) (6). These compounds have been investigated by single crystal X-ray diffractometry and magnetochemistry. In complex 1 the Mn(II) is bonded to one bpaeOH and two thiocyanate ions, while in complex 2 it is coordinated to a deprotonated bpaeO(-) and two azide ions. The oxidation states of manganese ions are 2+ for 1 and 3+ for 2, respectively, indicating that the different oxidation states depend on the type of binding anions. The structures of monomeric iron(II) and cobalt(II) complexes 3 and 5 with two thiocyanate ions are isomorphous to that of 1. Compounds 1, 2, 3, and 5 exhibit high-spin states in the temperature range 5 to 300 K. 4 contains two different iron(III) ions in an asymmetric unit, one is coordinated to a deprotonated bpaeO(-), an azide ion, and a methoxy group, and the other is bonded to three azide ions and two oxygens from bpaeO(-) and a methoxy group. Two independent iron(III) ions in 4 form a tetranuclear complex by symmetry. 4 displays both ferromagnetic and antiferromagnetic couplings (J = 9.8 and -14.3 cm(-1)) between the iron(III) ions. 6 is a mixed-valence trinuclear cobalt complex, which is formulated as Co(III)(S = 0)-Co(II)(S = 3/2)-Co(III)(S = 0). The effective magnetic moment at room temperature corresponds to the high-spin cobalt(II) ion (∼4.27 μ(B)). Interestingly, 6 showed efficient catalytic activities toward various olefins and alcohols with modest to excellent yields, and it has been proposed that a high-valent Co(V)-oxo species might be responsible for oxygen atom transfer in the olefin epoxidation and

  10. Dimensional Effects on the Momentum distribution of Bosonic Trimer States

    DEFF Research Database (Denmark)

    F. Bellotti, F.; Frederico, T.; T. Yamashita, M.

    2013-01-01

    The momentum distribution is a powerful probe of strongly-interacting systems that are expected to display universal behavior. This is contained in the contact parameters which relate few- and many-body properties. Here we consider a Bose gas in two dimensions and explicitly show that the two......-body contact parameter is universal and then demonstrate that the momentum distribution at next-to-leading order has a logarithmic dependence on momentum which is vastly different from the three-dimensional case. Based on this, we propose a scheme for measuring the effective dimensionality of a quantum many......-body system by exploiting the functional form of the momentum distribution....

  11. Homoclinic chaos in the discrete self-trapping trimer

    DEFF Research Database (Denmark)

    Hennig, D.; Gabriel, H.; Jørgensen, Michael Finn

    1995-01-01

    We study the discrete self-trapping (DST) equation with three degrees of freedom. By taking the DST dimer as the underlying unperturbed system we treat the coupling to the additional oscillator as a small perturbation. Using the generalized Melnikov method we prove the existence of homoclinic chaos...

  12. Communication: Isotopic effects on tunneling motions in the water trimer

    Energy Technology Data Exchange (ETDEWEB)

    Videla, Pablo E. [Departamento de Química Inorgánica Analítica y Química-Física e INQUIMAE, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Ciudad Universitaria, Pabellón II, 1428 Buenos Aires (Argentina); Rossky, Peter J. [Department of Chemistry, Rice University, Houston, Texas 77251-1892 (United States); Laria, D., E-mail: dhlaria@cnea.gov.ar [Departamento de Química Inorgánica Analítica y Química-Física e INQUIMAE, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Ciudad Universitaria, Pabellón II, 1428 Buenos Aires (Argentina); Departamento de Física de la Materia Condensada, Comisión Nacional de Energía Atómica, Avenida Libertador 8250, 1429 Buenos Aires (Argentina)

    2016-02-14

    We present results of ring polymer molecular dynamics simulations that shed light on the effects of nuclear quantum fluctuations on tunneling motions in cyclic [H{sub 2}O]{sub 3} and [D{sub 2}O]{sub 3}, at the representative temperature of T = 75 K. In particular, we focus attention on free energies associated with two key isomerization processes: The first one corresponds to flipping transitions of dangling OH bonds, between up and down positions with respect to the O–O–O plane of the cluster; the second involves the interchange between connecting and dangling hydrogen bond character of the H-atoms in a tagged water molecule. Zero point energy and tunneling effects lead to sensible reductions of the free energy barriers. Due to the lighter nature of the H nuclei, these modifications are more marked in [H{sub 2}O]{sub 3} than in [D{sub 2}O]{sub 3}. Estimates of the characteristic time scales describing the flipping transitions are consistent with those predicted based on standard transition-state-approximation arguments.

  13. Viral aggregating and opsonizing activity in collectin trimers

    DEFF Research Database (Denmark)

    Hartshorn, Kevan L; White, Mitchell R; Tecle, Tesfaldet

    2010-01-01

    of bovine serum collectin CL-46 induces aggregation of influenza A virus (IAV) and potently increases IAV uptake by neutrophils. CL-46-NCRD showed calcium-dependent and sugar-sensitive binding to both neutrophils and IAV. Replacement of specific residues of the CRD of human SP-D with those found in bovine...... serum collectins conferred opsonizing activity. The most effective substitution involved replacement of arginine 343 with valine (hSP-D-NCRD/R343V). hSP-D-NCRD/R343V greatly increased viral uptake by neutrophils and monocytes and also potentiated neutrophil respiratory burst responses. These effects...... for IAV even in the absence of the collagen domain or higher order multimerization. This may involve increased affinity of individual CRDs for glycoconjugates displayed on host cells or the viral envelope. Key words: influenza virus, neutrophil, CL-46....

  14. Purification and Characterization of TrzF: Biuret Hydrolysis by Allophanate Hydrolase Supports Growth

    OpenAIRE

    Shapir, Nir; Cheng, Gang; Sadowsky, Michael J.; Wackett, Lawrence P.

    2006-01-01

    TrzF, the allophanate hydrolase from Enterobacter cloacae strain 99, was cloned, overexpressed in the presence of a chaperone protein, and purified to homogeneity. Native TrzF had a subunit molecular weight of 65,401 and a subunit stoichiometry of α2 and did not contain significant levels of metals. TrzF showed time-dependent inhibition by phenyl phosphorodiamidate and is a member of the amidase signature protein family. TrzF was highly active in the hydrolysis of allophanate but was not acti...

  15. Catalytic signal amplification using [Fe(III)(biuret-amide)]-mesoporous silica nanoparticles: visual cyanide detection.

    Science.gov (United States)

    Panda, Chakadola; Dhar, Basab B; Malvi, Bharmana; Bhattacharjee, Yudhajit; Gupta, Sayam Sen

    2013-03-18

    Catalytic signal amplification was used for the colorimetric detection of CN(-) in aqueous media by using the enzyme catalase in tandem with mesoporous silica nanoparticle based synthetic HRP enzyme mimic Fe-MSNs. Signal amplification up to a maximum of eight fold was observed for the reporter "oxidized TMB" with respect to the added CN(-) ion.

  16. Study of interaction of biuret with praseodymium sulfate and samarium bromide in aqueous solutions

    International Nuclear Information System (INIS)

    Yunusov, Z.Yu.; Davranov, M.D.; Balbaev, M.K.; Sulajmankulov, K.S.

    1985-01-01

    Using the solubility method, ternary systems Pr(SO 4 ) 2 - NH(CONH 2 ) 2 - H 2 O and SmBr 3 - NH 4 (CONH 2 ) 2 - H 2 O at 30 deg are studied. In the systems the formation of crystal compounds Pr 2 (SO 4 ) 3 x3NH(CONH 2 ) 2 and SmBr 3 x3NH(CONH 2 ) 2 is detected

  17. Guidelines for standard and biuretic renogram in children; Empfehlungen zur Durchfuehrung der Nierenfunktionsszintigraphie mit und ohne Furosemidbelastung

    Energy Technology Data Exchange (ETDEWEB)

    Gordon, I. [Great Ormond Street Hospital for Children, London (United Kingdom); Piepsz, A. [CHU St. Pierre, Brussels (Belgium); Colarinha, P. [Instituto Portugues de Oncologia, Lisbon (Portugal); Hahn, K.; Fischer, S.; Porn, U. [LMU Muenchen (Germany). Klinik fuer Nuklearmedizin; Olivier, P. [CHU Brabois, Nancy (France); Sixt, R. [The Queen Silvia Children' s Hospital, Goeteborg (Sweden); Velzen, J. van

    2000-11-01

    The guidelines are intended to help nuclear medical teams in their daily routine. The information given relates to aspects such as data acquisition, evaluation and interpretation, and indications for pediatric renal functional scintigraphy. The guidelines have been elaborated in response to a request of EANM and the American Society of Nuclear Medicine, who expressed the need for guidelines on recommended procedures for most of the standard nuclear medical examinations. The guidelines express the opinion of the Paediatric Committee of the EANM, and should be seen in the context of generally accepted basic principles in nuclear medicine, as well as local and national regulatory standards in radiation protection. (orig./CB) [German] Zweck dieser Leitlinie ist es, dem nuklearmedizinischen Team Hilfestellung fuer die taegliche Routinepraxis zu geben. Diese Leitlinie enthaelt Informationen ueber die Datenakquisition, Auswertung, Interpretation sowie die Indikationen der Nierenfunktionsszintigraphie bei Kindern. Das vorliegende Dokument wurde durch den Wunsch der EANM und der American Society of Nuclear Medicine nach Leitlinien fuer die meisten nuklearmedizinischen Untersuchungsverfahren initiiert. Die Leitlinie gibt die Ansicht des Paediatric Committee der EANM wieder. Sie sollte immer in Verbindung gesehen werden mit den allgemeingueltigen Grundlagen der Nuklearmedizin sowie den lokalen und nationalen Regelungen des Strahlenschutzes. (orig./MG)

  18. Optimization of the conditions for biuret complex formation for the determination of peptides by capillary electrophoresis with ultraviolet detection.

    Science.gov (United States)

    Gawron, A J; Lunte, S M

    2000-06-01

    Capillary electrophoresis with UV detection was utilized to optimize copper complexation conditions for the analysis of neuropeptides. Complexation was confirmed by monitoring the response at a visible wavelength. Four complexation strategies were used to compare the UV response of native peptides and their respective copper complexes. All four strategies resulted in complete complexation, but on-capillary complexation provided significant advantages over precapillary and pre-/on-capillary. An increase in UV absorbance along with peak stacking resulted in a significantly greater response using the on-capillary technique. Also, on-capillary complexation does not require dilution of the sample. The effects of temperature and copper concentration were also investigated. The utility of this method for the separation of an enkephalin peptide mixture is presented.

  19. Complexes of biuret with Cr(III), Ag(II), Cu(III) mixed ligand and UO2(II)

    International Nuclear Information System (INIS)

    Sanyal, R.M.; Ansari, B.J.; Srivastava, P.C.; Banerjee, B.K.; Chakraburtty

    1979-01-01

    The methods of preparation of some new complexes of Cr(III), Ag(III), Cu(II) mixed ligand and UO 2 (II) and their characterisation from their electronic, infrared and ESR spectra as well as from their magnetic susceptibility values are described. Crystal field parameters of some of the complexes have been calculated. (auth.)

  20. Surface Induced Dissociations of Protonated Ethanol Monomer, Dimer and Trimer Ions: Trimer Break-down Graph from the Collision Energy Dependence of Projectile Fragmentation

    Czech Academy of Sciences Publication Activity Database

    Mair, C.; Lezius, M.; Herman, Zdeněk; Märk, T. D.

    2003-01-01

    Roč. 118, č. 15 (2003), s. 7090-7096 ISSN 0021-9606 R&D Projects: GA ČR GA203/00/0632 Grant - others:Cooperation Program in Science and Technology(AT) Kontakt 1999-2000 Institutional research plan: CEZ:AV0Z4040901 Keywords : clusters ions * molecule reaction * polyatomic ions Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 2.950, year: 2003

  1. A convenient method for rapid determination of proteins in rices and rice products

    OpenAIRE

    光永, 俊郎; 安藤, ひとみ

    1990-01-01

    [Author abstract]In the biuret determination of protein in rices and rice products, starch and lipids extractable with an alkaline solution such as biuret reagent were found to interfere with the biuret method. Comparative evaluation of their effects revealed that starch, a major component of rice, has the most significant effect on the biuret method. In the presence of starch, the correlation between Kjeldahl protein and the optical density of biuret was poor. It was found that several perha...

  2. Thermoresponsive polymer nanoparticles co-deliver RSV F trimers with a TLR-7/8 adjuvant

    Czech Academy of Sciences Publication Activity Database

    Francica, J. R.; Lynn, G. M.; Laga, Richard; Joyce, M. G.; Ruckwardt, T. J.; Morabito, K. M.; Chen, M.; Chaudhuri, R.; Zhang, B.; Sastry, M.; Druz, A.; Ko, K.; Choe, M.; Pechar, Michal; Georgiev, I. S.; Kueltzo, L. A.; Seymour, L. W.; Mascola, J. R.; Kwong, P. D.; Graham, B. S.; Seder, R. A.

    2016-01-01

    Roč. 27, č. 10 (2016), s. 2372-2385 ISSN 1043-1802 R&D Projects: GA ČR(CZ) GJ16-14957Y; GA MŠk(CZ) LQ1604 Institutional support: RVO:61389013 Keywords : thermoresponsive polymers * polymer vaccines * toll-like receptor agonists Subject RIV: CD - Macromolecular Chemistry Impact factor: 4.818, year: 2016

  3. MicroProtein-mediated recruitment of CONSTANS into a TOPLESS trimeric complex represses flowering in Arabidopsis

    DEFF Research Database (Denmark)

    Graeff, Moritz; Straub, Daniel; Eguen, Tenai E.

    2016-01-01

    Arabidopsis thaliana microProteins, miP1a and miP1b, physically interact with CONSTANS (CO) a potent regulator of flowering time. The miP1a/b-type microProteins evolved in dicotyledonous plants and have an additional carboxy-terminal PF(V/L)FL motif. This motif enables miP1a/b microProteins to interact......MicroProteins are short, single domain proteins that act by sequestering larger, multi-domain proteins into non-functional complexes. MicroProteins have been identified in plants and animals, where they are mostly involved in the regulation of developmental processes. Here we show that two...... with TOPLESS/TOPLESS-RELATED (TPL/TPR) proteins. Interaction of CO with miP1a/b/TPL causes late flowering due to a failure in the induction of FLOWERING LOCUS T (FT) expression under inductive long day conditions. Both miP1a and miP1b are expressed in vascular tissue, where CO and FT are active. Genetically...

  4. Theranostic Value of Multimers: Lessons Learned from Trimerization of Neurotensin Receptor Ligands and Other Targeting Vectors

    Directory of Open Access Journals (Sweden)

    Simone Maschauer

    2017-03-01

    Full Text Available Neurotensin receptor 1 (NTS1 is overexpressed on a variety of cancer entities; for example, prostate cancer, ductal pancreatic adenocarcinoma, and breast cancer. Therefore, it represents an interesting target for the diagnosis of these cancers types by positron emission tomography (PET [...

  5. Versatile coordination of cyclopentadienyl-arene ligands and its role in titanium-catalyzed ethylene trimerization

    NARCIS (Netherlands)

    Otten, Edwin; Batinas, Aurora A.; Meetsma, Auke; Hessen, Bart

    2009-01-01

    Cationic titanium(IV) complexes with ansa-(eta(5)-cyclopentadienyl,eta(6)-arene) ligands were synthesized and characterized by X-ray crystallography. The strength of the metal-arene interaction in these systems was studied by variable-temperature NMR spectroscopy. Complexes with a C, bridge between

  6. Targeting HIV-1 Envelope Glycoprotein Trimers to B Cells by Using APRIL Improves Antibody Responses

    NARCIS (Netherlands)

    Melchers, Mark; Bontjer, Ilja; Tong, Tommy; Chung, Nancy P. Y.; Klasse, Per Johan; Eggink, Dirk; Montefiori, David C.; Gentile, Maurizio; Cerutti, Andrea; Olson, William C.; Berkhout, Ben; Binley, James M.; Moore, John P.; Sanders, Rogier W.

    2012-01-01

    An HIV-1 vaccine remains elusive, in part because various factors limit the quantity and quality of the antibodies raised against the viral envelope glycoprotein complex (Env). We hypothesized that targeting Env vaccines directly to B cells, by fusing them to molecules that bind and activate these

  7. Postionization fragmentation of rare-gas trimers revisited with new theoretical approaches

    Czech Academy of Sciences Publication Activity Database

    Janeček, I.; Cintavá, S.; Hrivňák, D.; Kalus, R.; Fárník, Michal; Gadea, F. X.

    2009-01-01

    Roč. 131, č. 11 (2009), s. 114306 ISSN 0021-9606 R&D Projects: GA AV ČR IAA401870702 Institutional research plan: CEZ:AV0Z40400503 Keywords : molecular dynamics * electron-impact * physical properties Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 3.093, year: 2009

  8. Gold nanoparticles conjugating recombinant influenza hemagglutinin trimers and flagellin enhanced mucosal cellular immunity.

    Science.gov (United States)

    Wang, Chao; Zhu, Wandi; Luo, Yuan; Wang, Bao-Zhong

    2018-04-09

    The immunogenicity of subunit vaccines can be augmented by formulating them into nanoparticles. We conjugated recombinant trimetric influenza A/Aichi/2/68(H3N2) hemagglutinin (HA) onto functionalized gold nanoparticles (AuNPs) surfaces in a repetitive, oriented configuration. To further improve the immunogenicity, we generated Toll-like receptor 5 (TLR5) agonist flagellin (FliC)-coupled AuNPs as particulate adjuvants. Intranasal immunizations with an AuNP-HA and AuNP-FliC particle mixture elicited strong mucosal and systemic immune responses that protected hosts against lethal influenza challenges. Compared with the AuNP-HA alone group, the addition of AuNP-FliC improved mucosal B cell responses as characterized by elevated influenza specific IgA and IgG levels in nasal, tracheal, and lung washes. AuNP-HA/AuNP-FliC also stimulated antigen-specific interferon-γ (IFN-γ)-secreting CD4 + cell proliferation and induced strong effector CD8 + T cell activation. Our results indicate that intranasal co-delivery of antigen and adjuvant-displaying AuNPs enhanced vaccine efficacy by inducing potent cellular immune responses. Copyright © 2018. Published by Elsevier Inc.

  9. Efimov Trimers near the Zero-crossing of a Feshbach Resonance

    DEFF Research Database (Denmark)

    Zinner, Nikolaj Thomas

    2012-01-01

    Near a Feshbach resonance, the two-body scattering length can assume any value. When it approaches zero, the next-order term given by the effective range is known to diverge. We consider the question of whether this divergence (and the vanishing of the scattering length) is accompanied by an anom...... by an anomalous solution of the three-boson Schr\\"odinger equation similar to the one found at infinite scattering length by Efimov. Within a simple zero-range model, we find no such solutions, and conclude that higher-order terms do not support Efimov physics....

  10. Energetics and structures of charged helium clusters: comparing stabilities of dimer and trimer cationic cores.

    Science.gov (United States)

    Marinetti, Fabio; Bodo, Enrico; Gianturco, Franco A; Yurtsever, Ersin

    2008-12-01

    We present accurate ab initio calculations of the most stable structures of He(n)(+) clusters in order to determine the more likely ionic core arrangements existing after reaching structural equilibrium of the clusters. Two potential energy surfaces are presented: one for the He(2)(+) and the other with the He(3)(+) linear ion, both interacting with one He atom. The two computed potentials are in turn employed within a classical structure optimization where the overall interaction forces are obtained within the sum-of-potentials approximation described in the main text. Because of the presence of many-body effects within the ionic core, we find that the arrangements with He(3)(+) as a core turn out to be energetically preferred, leading to the formation of He(3)(+)(He)(n-3) stable aggregates. Nanoscopic considerations about the relative stability of clusters with the two different cores are shown to give us new information on the dynamical processes observed in the impact ionization experiments of pure helium clusters and the importance of pre-equilibrium evaporation of the ionic dimers in the ionized clusters.

  11. Symmetrical trimeric star-shaped mesogens based on 1,3,5 ...

    Indian Academy of Sciences (India)

    joined to the 1,3,5-position of phloroglucinol to yield the symmetrical three-armed star-shaped molecules. The acronyms used to refer the two series of star-shaped mesogens in this article are PSB-X-n and PAZ-X-n, in which PSB-X-n is referred to the star-shaped mesogen incorporating Schiff base moieties while PAZ-X-n is.

  12. Symmetrical trimeric star-shaped mesogens based on 1, 3, 5 ...

    Indian Academy of Sciences (India)

    These star-shaped mesogens exhibit only nematic and smectic phases. The difference between the two series lies on the structure of linking group in the peripheral units (-CH=N- for series PSB-X- and -N=N- for series PAZ-X-). Therefore, a comparison study of the mesomorphic properties between these two series of ...

  13. The role of quantum coherence in dimer and trimer excitation energy transfer

    Science.gov (United States)

    Bengtson, Charlotta; Sjöqvist, Erik

    2017-11-01

    Recent progress in resource theory of quantum coherence has resulted in measures to quantify coherence in quantum systems. Especially, the l 1-norm and relative entropy of coherence have been shown to be proper quantifiers of coherence and have been used to investigate coherence properties in different operational tasks. Since long-lasting quantum coherence has been experimentally confirmed in a number of photosynthetic complexes, it has been debated if and how coherence is connected to the known efficiency of population transfer in such systems. In this study, we investigate quantitatively the relationship between coherence, as quantified by l 1-norm and relative entropy of coherence, and efficiency, as quantified by fidelity, for population transfer between end-sites in a network of two-level quantum systems. In particular, we use the coherence averaged over the duration of the population transfer in order to carry out a quantitative comparison between coherence and fidelity. Our results show that although coherence is a necessary requirement for population transfer, there is no unique relation between coherence and the efficiency of the transfer process.

  14. Bacillus cereus iron uptake protein fishes out an unstable ferric citrate trimer.

    Science.gov (United States)

    Fukushima, Tatsuya; Sia, Allyson K; Allred, Benjamin E; Nichiporuk, Rita; Zhou, Zhongrui; Andersen, Ulla N; Raymond, Kenneth N

    2012-10-16

    Citrate is a common biomolecule that chelates Fe(III). Many bacteria and plants use ferric citrate to fulfill their nutritional requirement for iron. Only the Escherichia coli ferric citrate outer-membrane transport protein FecA has been characterized; little is known about other ferric citrate-binding proteins. Here we report a unique siderophore-binding protein from the gram-positive pathogenic bacterium Bacillus cereus that binds multinuclear ferric citrate complexes. We have demonstrated that B. cereus ATCC 14579 takes up (55)Fe radiolabeled ferric citrate and that a protein, BC_3466 [renamed FctC (ferric citrate-binding protein C)], binds ferric citrate. The dissociation constant (K(d)) of FctC at pH 7.4 with ferric citrate (molar ratio 1:50) is 2.6 nM. This is the tightest binding observed of any B. cereus siderophore-binding protein. Nano electrospray ionization-mass spectrometry (nano ESI-MS) analysis of FctC and ferric citrate complexes or citrate alone show that FctC binds diferric di-citrate, and triferric tricitrate, but does not bind ferric di-citrate, ferric monocitrate, or citrate alone. Significantly, the protein selectively binds triferric tricitrate even though this species is naturally present at very low equilibrium concentrations.

  15. Murine Antibody Responses to Cleaved Soluble HIV-1 Envelope Trimers Are Highly Restricted in Specificity

    NARCIS (Netherlands)

    Hu, Joyce K.; Crampton, Jordan C.; Cupo, Albert; Ketas, Thomas; van Gils, Marit J.; Sliepen, Kwinten; de Taeye, Steven W.; Sok, Devin; Ozorowski, Gabriel; Deresa, Isaiah; Stanfield, Robyn; Ward, Andrew B.; Burton, Dennis R.; Klasse, Per Johan; Sanders, Rogier W.; Moore, John P.; Crotty, Shane

    2015-01-01

    Generating neutralizing antibodies (nAbs) is a major goal of many current HIV-1 vaccine efforts. To be of practical value, these nAbs must be both potent and cross-reactive in order to be capable of preventing the transmission of the highly diverse and generally neutralization resistant (Tier-2)

  16. Effects of analogs of indole-3-carbinol cyclic trimerization product in human breast cancer cells.

    Science.gov (United States)

    Xue, Ling; Schaldach, Charlene M; Janosik, Tomasz; Bergman, Jan; Bjeldanes, Leonard F

    2005-04-15

    5,6,11,12,17,18-Hexahydrocyclonona[1,2-b:4,5-b*:7,8-b**]triindole (CTr) is a major digestive product of indole-3-carbinol (I3C) from Brassica vegetables and exhibits strong estrogenic activities. CTr increases proliferation of estrogen-dependent breast tumor cells, binds with strong affinity for the estrogen receptor-alpha (ERalpha), and activates expression of estrogen (E(2))-dependent genes. To begin to examine the structural features that determine the biological activity of CTr, we prepared and studied the effects of two analogs, 9,18-dihydro-12H-[1,2,5]trithionino[3,4-b:6,7-b*:9,8-b**]triindole (S(3)CTr) and 5,6,11,12,17,18-hexahydro-5,11,17-trimethylcyclonona[1,2-b:4,5-b*:7,8-b**]triindole (Me(3)CTr). N-Methylation of CTr completely ablated the estrogenic activities of CTr. In the dose range in which CTr was clearly estrogenic, Me(3)CTr exhibited no detectable effect on cell growth, ERalpha binding to E(2), or ERalpha-responsive gene expression. S(3)CTr showed mixed ERalpha agonist activities. It bound to the ERalpha and activated receptor binding with DNA, weakly activated expression of transfected E(2)-responsive reporter gene constructs, and strongly inhibited the E(2)-induced activation of these reporter constructs. S(3)CTr activated aryl hydrocarbon receptor (AhR)-mediated pathways, consistent with the moderately strong binding affinity of S(3)CTr for the AhR. Comparisons of the conformational characteristics among CTr and its two analogs indicated that the estrogenic effects of CTr are highly sensitive to apparently minor structural modifications, and further supported the hypothesis for a central role of hydrogen bonding around the nitrogen atom in CTr binding to the ligand binding site of ERalpha.

  17. Designing a Soluble Near Full-Length HIV-1 GP41 Trimer

    Science.gov (United States)

    2012-11-26

    required SDS, a strong ionic detergent , for solubilization. Even with SDS, only partial solubilization was achieved (Fig. 2D, upper panel, lane 2...This work was partly supported by a grant from The National Institute of Allergy and Infectious Diseases (R01AI02725). 1To whom correspondence may be

  18. Helical H-type aggregation of trimeric p-phenylene vinylene with ...

    Indian Academy of Sciences (India)

    on a UV-3000 spectrophotometer. Photoluminescence measurements were carried out by using a Hitachi F-4500 flourescence spectrophotometer. CD spectra was recorded on Bio-kine32 V4·51-[Acquire spectra (MOS-450)] spec- trophotometer. The specimen for TEM observation were prepared using carbon support films.

  19. Therapeutic TNF Inhibitors can Differentially Stabilize Trimeric TNF by Inhibiting Monomer Exchange

    NARCIS (Netherlands)

    van Schie, Karin A.; Ooijevaar-de Heer, Pleuni; Dijk, Lisanne; Kruithof, Simone; Wolbink, Gertjan; Rispens, Theo

    2016-01-01

    Tumor necrosis factor (TNF) is a homotrimeric cytokine that is a key mediator of inflammation. It is unstable at physiological concentrations and slowly converts into an inactive form. Here, we investigated the mechanism of this process by using a Forster resonance energy transfer (FRET) assay that

  20. Trimerization of apolipoprotein A-I retards plasma clearance and preserves antiatherosclerotic properties

    DEFF Research Database (Denmark)

    Graversen, Jonas Heilskov; Laurberg, Jacob Marsvin; Andersen, Mikkel Holmen

    2008-01-01

    An increased plasma level of the major high-density lipoprotein (HDL) component, apolipoprotein A-I (apoA-I) is the aim of several therapeutic strategies for combating atherosclerotic disease. HDL therapy by direct intravenous administration of apoA-I is a plausible way; however, a fast renal...

  1. Subcellular shifts of trimeric G-proteins following activation of baker's yeast by glucose

    Czech Academy of Sciences Publication Activity Database

    Kotyk, Arnošt; Ihnatovych, Ivanna; Lapathitis, G.; Lamash, N.; Svoboda, Petr

    2001-01-01

    Roč. 46, č. 5 (2001), s. 391-396 ISSN 0015-5632 R&D Projects: GA ČR GA204/98/0474; GA MŠk VS97099 Institutional research plan: CEZ:AV0Z5011922 Keywords : G-proteins * baker ´s yeast * glucose activation Subject RIV: CE - Biochemistry Impact factor: 0.776, year: 2001

  2. Computational characterization of parallel dimeric and trimeric coiled-coils using effective amino acid indices.

    Science.gov (United States)

    Li, Chen; Wang, Xiao-Feng; Chen, Zhen; Zhang, Ziding; Song, Jiangning

    2015-02-01

    The coiled-coil, which consists of two or more α-helices winding around each other, is a ubiquitous and the most frequently observed protein-protein interaction motif in nature. The coiled-coil is known for its straightforward heptad repeat pattern and can be readily recognized based on protein primary sequences, exhibiting a variety of oligomer states and topologies. Due to the stable interaction formed between their α-helices, coiled-coils have been under close scrutiny to design novel protein structures for potential applications in the fields of material science, synthetic biology and medicine. However, their broader application requires an in-depth and systematic analysis of the sequence-to-structure relationship of coiled-coil folding and oligomeric formation. In this article, we propose a new oligomerization state predictor, termed as RFCoil, which exploits the most useful and non-redundant amino acid indices combined with the machine learning algorithm - random forest (RF) - to predict the oligomeric states of coiled-coil regions. Benchmarking experiments show that RFCoil achieves an AUC (area under the ROC curve) of 0.849 on the 10-fold cross-validation test using the training dataset and 0.855 on the independent test using the validation dataset, respectively. Performance comparison results indicate that RFCoil outperforms the four existing predictors LOGICOIL, PrOCoil, SCORER 2.0 and Multicoil2. Furthermore, we extract a number of predominant rules from the trained RF model that underlie the oligomeric formation. We also present two case studies to illustrate the applicability of the extracted rules to the prediction of coiled-coil oligomerization state. The RFCoil web server, source codes and datasets are freely available for academic users at http://protein.cau.edu.cn/RFCoil/.

  3. Delta-Opioid receptors exhibit high efficiency when activating trimeric G proteins in membrane domains

    Czech Academy of Sciences Publication Activity Database

    Bouřová, Lenka; Koštrnová, Alexandra; Hejnová, Lucie; Moravcová, Zuzana; Moon, H. E.; Novotný, Jiří; Milligan, G.; Svoboda, Petr

    2003-01-01

    Roč. 85, č. 1 (2003), s. 34-49 ISSN 0022-3042 R&D Projects: GA MŠk LN00A026; GA ČR GA309/01/0255 Grant - others:Welcome Trust(GB) xx Institutional research plan: CEZ:AV0Z5011922 Keywords : membrane domains * GTPase activity * G protein coupling Subject RIV: CE - Biochemistry Impact factor: 4.825, year: 2003

  4. A mixed Ni (II) ionic complex containing V-shaped water trimer ...

    Indian Academy of Sciences (India)

    Author Affiliations. Nallasamy Palanisami1 Kabali Senthilkumar2 Mohan Gopalakrishnan2 Il-Shik Moon1. Department of Chemical Engineering, Sunchon National University, 315 Maegok Dong, Suncheon 540-742, Chonnam, South Korea; Materials Chemistry Division, School of Advanced Sciences, VIT University, Vellore ...

  5. Penentuan Kadar Nitrogen Pada Pupuk Urea Di PT. Pupuk Iskandar Muda

    OpenAIRE

    Tampubolon, Arfandy

    2011-01-01

    In order to know the content of nitrogen in urea fertilizer teh contant oh H2O must be determined (% H2O) wellas the percentape of biuret (% Biuret). Og biuret content the determination of nitrogen content is conducted using the calculation % N = ( 100 - % H2O - % Biuret) x 0.4665 + (% Biuret x 0.4077 ). From result of analysis. the calculation, the content of H2O is 0.233 %, biuret content 0.4966 % and the nitrogen content is 46 %.these results show that the nitrogen content obtained from u...

  6. Evaluation of four commercial biuret reagent kits of serum total protein by the American Association for Clinical Chemistry reference measurement procedure.

    Science.gov (United States)

    He, Meilin; Zhang, Jie

    2011-06-01

    In China, the traceability of clinical chemistry methods is still immature. Therefore, it is necessary to establish a reference measurement procedure and evaluate commercial reagent kits using such established procedures. We reproduced the reference measurement procedure for serum total protein, as recommended by the American Association for Clinical Chemistry (AACC). We evaluated the performance by Clinical Laboratory Standard Institute (CLSI) guidelines EP15-A and EP6-A. Subsequently, four commercial reagent kits were evaluated by the reproduced reference procedure following CLSI guideline EP9-A2. The performance of the reproduced reference procedure was as follows: CVs ranged from 0.47% to 0.85% at medical decision levels (X(c)) of 45 g/L, 60 g/L and 80 g/L. Linearity was Y=1.0022X-0.2121 (r=0.9999), and recovery ranged from 100.2% to 102.4%. The External Quality Assessment Scheme for Reference Laboratories in Laboratory Medicine (RELA) was applied, and the result was within the limit of equivalence. The linear relationships of four commercial reagent kits, Merit Choice, KHB, Leadman, and Olympus, were, respectively: Y=0.9922X+0.5776 (r=0.9961); Y=0.9936X+0.4316 (r=0.9992); Y=0.9949X+0.9129 (r=0.9987) and Y=0.9923X+0.8876 (r=0.9989). KHB showed slight negative bias, and the mean±SD was -0.03±0.60 g/L. Merit Choice, Leadman, and Olympus all showed positive bias, and the mean±SDs were 0.02±0.63 g/L, 0.55±0.77 g/L and 0.34±0.71 g/L, respectively. The correlation and bias of four commercial reagent kits for serum total protein were found to be acceptable. Thus, these reagent kits can be used reliably in China.

  7. Systems LiBO2(NaBO2, RBO2)-(H2NCO)2NY-H2O at 25 deg C

    International Nuclear Information System (INIS)

    Skvortsov, V.G.; Fedorov, Yu.A.; Vanyulin, A.N.; Tsekhanskij, R.S.

    1993-01-01

    Lithium (sodium, potassium) monoborate-biuret-water systems at 25 deg C are studied using solubility, diffraction and pH-metry techniques. Monoborates affect biuret positively with lyotropic effect, while buiret affects borates negatively

  8. A recombinant rabies vaccine expressing the trimeric form of the glycoprotein confers enhanced immunogenicity and protection in outbred mice.

    NARCIS (Netherlands)

    Koraka, Penelope; Bosch, Berend-Jan; Cox, Manon; Chubet, Rick; Amerongen, Geert van; Lövgren-Bengtsson, Karen; Martina, Byron E E; Roose, Jouke; Rottier, Peter J M; Osterhaus, Albert D M E

    2014-01-01

    Rabies is a disease characterized by an invariably lethal encephalitis of viral origin that can be controlled by preventive vaccination programs of wildlife, domestic animals and humans in areas with a high risk of exposure. Currently available vaccines are expensive, cumbersome to produce and

  9. Functional discrepancies between tumor necrosis factor and lymphotoxin alpha explained by trimer stability and distinct receptor interactions

    DEFF Research Database (Denmark)

    Schuchmann, M; Hess, S; Bufler, P

    1995-01-01

    interaction with the human p55TNFR. This was demonstrated in NIH 3T3 cells transfected with the human p55TNFR, where cytotoxicity is mediated exclusively by the transfected receptor. Although the p55ATNFR had virtually identical affinities for TNF and LT alpha, as defined by Scatchard analysis......Tumor necrosis factor (TNF) and lymphotoxin alpha (LT alpha) are closely related cytokines which bind with nearly identical affinities to the same pair of cell surface receptors, p55 and p75TNFR. Therefore it is assumed that TNF and LT alpha are redundant cytokines. This study, however...

  10. Phase I randomised clinical trial of an HIV-1(CN54, clade C, trimeric envelope vaccine candidate delivered vaginally.

    Directory of Open Access Journals (Sweden)

    David J Lewis

    Full Text Available We conducted a phase 1 double-blind randomised controlled trial (RCT of a HIV-1 envelope protein (CN54 gp140 candidate vaccine delivered vaginally to assess immunogenicity and safety. It was hypothesised that repeated delivery of gp140 may facilitate antigen uptake and presentation at this mucosal surface. Twenty two healthy female volunteers aged 18-45 years were entered into the trial, the first receiving open-label active product. Subsequently, 16 women were randomised to receive 9 doses of 100 µg of gp140 in 3 ml of a Carbopol 974P based gel, 5 were randomised to placebo solution in the same gel, delivered vaginally via an applicator. Participants delivered the vaccine three times a week over three weeks during one menstrual cycle, and were followed up for two further months. There were no serious adverse events, and the vaccine was well tolerated. No sustained systemic or local IgG, IgA, or T cell responses to the gp140 were detected following vaginal immunisations. Repeated vaginal immunisation with a HIV-1 envelope protein alone formulated in Carbopol gel was safe, but did not induce local or systemic immune responses in healthy women.ClinicalTrials.gov NCT00637962.

  11. Mono-, di- and trimeric PS I reaction center complexes isolated from the thermophilic cyanobacterium Synechococcus sp. Size, shape and activity

    NARCIS (Netherlands)

    Rögner, M.; Mühlenhoff, U.; Boekema, E.J.; Witt, H.T.

    1990-01-01

    Photosystem I preparations from the cyanobacterium Synechococcus sp. were treated with high concentrations of Tris and octyl glucoside at alkaline pH and elevated temperature. A sucrose density gradient yielded three pigment-protein complexes; these were further purified on a HPLC anion-exchange

  12. A Single Immunization with Soluble Recombinant Trimeric Hemagglutinin Protects Chickens against Highly Pathogenic Avian Influenza Virus H5N1

    NARCIS (Netherlands)

    Cornelissen, A.H.M.; Vries, de R.P.; Boer-Luijtze, de E.A.; Rigter, A.; Rottier, P.J.M.; Haan, de C.A.M.

    2010-01-01

    Background: The highly pathogenic avian influenza (HPAI) virus H5N1 causes multi-organ disease and death in poultry, resulting in significant economic losses in the poultry industry. In addition, it poses a major public health threat as it can be transmitted directly from infected poultry to humans

  13. Enhancement of antiviral activity of collectin trimers through cross-linking and mutagenesis of the carbohydrate recognition domain

    DEFF Research Database (Denmark)

    White, Mitchell R; Boland, Patrick; Tecle, Tesfaldet

    2010-01-01

    . We have sought to find ways to increase the antiviral activity of collectin NCRDs. Cross-linking of the SP-D NCRD with nonblocking monoclonal antibodies (mAbs) markedly potentiates antiviral activity. In the present report, we demonstrate that F(ab')2 [but not F(ab')1] fragments of a cross-linking mAb...... collectins, we have constructed mutant versions of the human SP-D NCRD that have increased antiviral activity. These mutant NCRDs also had potentiated activity after cross-linking with F(ab')2 fragments or S protein complexes. Hence, the antiviral activity of NCRDs can be increased by 2 distinct...... have similar effects. Hence, cross-linking activity, but not the Fc domain of the mAb, is needed for increased antiviral activity. In contrast, the Fc domain of the mAb was important for increasing viral uptake or respiratory burst responses of human neutrophils. Our NCRD constructs contain an S...

  14. The three isoforms of the light-harvesting complex II Spectroscopic features, trimer formation, and functional roles

    CERN Document Server

    Standfuss, Jorg

    2004-01-01

    The major light-harvesting complex (LHC-II) of higher plants plays a crucial role in capturing light energy for photosynthesis and in regulating the flow of energy within the photosynthetic apparatus. Native LHC-II isolated from plant tissue consists of three isoforms, Lhcb1, Lhcb2, and Lhcb3, which form homo- and heterotrimers. All three isoforms are highly conserved among different species, suggesting distinct functional roles. We produced the three LHC-II isoforms by heterologous expression of the polypeptide in Escherichia coli and in vitro refolding with purified pigments. Although Lhcb1 and Lhcb2 are very similar in polypeptide sequence and pigment content, Lhcb3 is clearly different because it lacks an N-terminal phosphorylation site and has a higher chlorophyll a/b ratio, suggesting the absence of one chlorophyll b. Low temperature absorption and fluorescence emission spectra of the pure isoforms revealed small but significant differences in pigment organization. The oligomeric state of the pure isofo...

  15. The host-binding domain of the P2 phage tail spike reveals a trimeric iron-binding structure

    International Nuclear Information System (INIS)

    Yamashita, Eiki; Nakagawa, Atsushi; Takahashi, Junichi; Tsunoda, Kin-ichi; Yamada, Seiko; Takeda, Shigeki

    2011-01-01

    The C-terminal domain of a bacteriophage P2 tail-spike protein, gpV, was crystallized and its structure was solved at 1.27 Å resolution. The refined model showed a triple β-helix structure and the presence of iron, calcium and chloride ions. The adsorption and infection of bacteriophage P2 is mediated by tail fibres and tail spikes. The tail spikes on the tail baseplate are used to irreversibly adsorb to the host cells. Recently, a P2 phage tail-spike protein, gpV, was purified and it was shown that a C-terminal domain, Ser87–Leu211, is sufficient for the binding of gpV to host Escherichia coli membranes [Kageyama et al. (2009 ▶), Biochemistry, 48, 10129–10135]. In this paper, the crystal structure of the C-terminal domain of P2 gpV is reported. The structure is a triangular pyramid and looks like a spearhead composed of an intertwined β-sheet, a triple β-helix and a metal-binding region containing iron, calcium and chloride ions

  16. Protein-Nanocellulose Interactions in Paper Filters for Advanced Separation Applications.

    Science.gov (United States)

    Gustafsson, Simon; Manukyan, Levon; Mihranyan, Albert

    2017-05-16

    Protein-based pharmaceutics are widely explored for healthcare applications, and 6 out of 10 best-selling drugs today are biologicals. The goal of this work was to evaluate the protein nanocellulose interactions in paper filter for advanced separation applications such as virus removal filtration and bioprocessing. The protein recovery was measured for bovine serum albumin (BSA), γ-globulin, and lysozyme using biuret total protein reagent and polyacrylamide gel electrophoresis (PAGE), and the throughput was characterized in terms of flux values from fixed volume filtrations at various protein concentrations and under worst-case experimental conditions. The affinity of cellulose to bind various proteins, such as BSA, lysozyme, γ-globulin, and human IgG was quantified using a quartz crystal microbalance (QCMB) by developing a new method of fixing the cellulose fibers to the electrode surface without cellulose dissolution-precipitation. It was shown that the mille-feuille filter exhibits high protein recovery, that is, ∼99% for both BSA and lysozyme. However, γ-globulin does not pass through the membrane due to its large size (i.e., >180 kDa). The PAGE data show no substantial change in the amount of dimers and trimers before and after filtration. QCMB analysis suggests a low affinity between the nanocellulose surface and proteins. The nanocellulose-based filter exhibits desirable inertness as a filtering material intended for protein purification.

  17. Formation of a Trimeric Xpo1-Ran[GTP]-Ded1 Exportin Complex Modulates ATPase and Helicase Activities of Ded1.

    Directory of Open Access Journals (Sweden)

    Glenn Hauk

    Full Text Available The DEAD-box RNA helicase Ded1, which is essential in yeast and known as DDX3 in humans, shuttles between the nucleus and cytoplasm and takes part in several basic processes including RNA processing and translation. A key interacting partner of Ded1 is the exportin Xpo1, which together with the GTP-bound state of the small GTPase Ran, facilitates unidirectional transport of Ded1 out of the nucleus. Here we demonstrate that Xpo1 and Ran[GTP] together reduce the RNA-stimulated ATPase and helicase activities of Ded1. Binding and inhibition of Ded1 by Xpo1 depend on the affinity of the Ded1 nuclear export sequence (NES for Xpo1 and the presence of Ran[GTP]. Association with Xpo1/Ran[GTP] reduces RNA-stimulated ATPase activity of Ded1 by increasing the apparent KM for the RNA substrate. Despite the increased KM, the Ded1:Xpo1:Ran[GTP] ternary complex retains the ability to bind single stranded RNA, suggesting that Xpo1/Ran[GTP] may modulate the substrate specificity of Ded1. These results demonstrate that, in addition to transport, exportins such as Xpo1 also have the capability to alter enzymatic activities of their cargo.

  18. Biochemical characterization of CK2alpha and alpha' paralogues and their derived holoenzymes: evidence for the existence of a heterotrimeric CK2alpha'-holoenzyme forming trimeric complexes

    DEFF Research Database (Denmark)

    Olsen, Birgitte; Rasmussen, Tine; Niefind, Karsten

    2008-01-01

    Altogether 2 holoenzymes and 4 catalytic CK2 constructs were expressed and characterized i.e. CK2alpha (2) (1-335) beta(2); CK2alpha'-derived holoenzyme; CK2alpha(1-335); MBP-CK2alpha'; His-tagged CK2alpha and His-tagged CK2alpha'. The two His-tagged catalytic subunits were expressed in insect...... cells, all others in Escherichia coli. IC(50) studies involving the established CK2 inhibitors DMAT, TBBt, TBBz, apigenin and emodin were carried out and the K(i) values calculated. Although the differences in the K(i) values found were modest, there was a general tendency showing that the CK2...... holoenzymes were more sensitive towards the inhibitors than the free catalytic subunits. Thermal inactivation experiments involving the individual catalytic subunits showed an almost complete loss of activity after only 2 min at 45 degrees C. In the case of the two holoenzymes, the CK2alpha...

  19. beta-Arrestin Interacts with the Beta/Gamma Subunits of Trimeric G-Proteins and Dishevelled in the Wnt/Ca2+ Pathway in Xenopus Gastrulation

    Czech Academy of Sciences Publication Activity Database

    Seitz, K.; Dursch, V.; Harnoš, J.; Bryja, Vítězslav; Gentzel, M.; Schambony, A.

    2014-01-01

    Roč. 9, č. 1 (2014) E-ISSN 1932-6203 R&D Projects: GA ČR(CZ) GC204/09/J030 Grant - others:GA ČR(CZ) GA204/09/0498 Institutional support: RVO:68081707 Keywords : CONVERGENT EXTENSION MOVEMENTS * WNT SIGNALING PATHWAYS * WNT /BETA-CATENIN Subject RIV: BO - Biophysics Impact factor: 3.234, year: 2014

  20. Alkyne Hydroamination and Trimerization with Titanium Bis(phenolate)pyridine Complexes: Evidence for Low-Valent Titanium Intermediates and Synthesis of an Ethylene Adduct of Titanium(II)

    KAUST Repository

    Tonks, Ian A.

    2013-06-24

    A class of titanium precatalysts of the type (ONO)TiX2 (ONO = pyridine-2,6-bis(4,6-di-tert-butylphenolate); X = Bn, NMe2) has been synthesized and crystallographically characterized. The (ONO)TiX2 (X = Bn, NMe2, X2 = NPh) complexes are highly active precatalysts for the hydroamination of internal alkynes with primary arylamines and some alkylamines. A class of titanium imido/ligand adducts, (ONO)Ti(L)(NR) (L = HNMe2, py; R = Ph, tBu), have also been synthesized and characterized and provide structural analogues to intermediates on the purported catalytic cycle. Furthermore, these complexes exhibit unusual redox behavior. (ONO)TiBn2 (1) promotes the cyclotrimerization of electron-rich alkynes, likely via a catalytically active TiII species that is generated in situ from 1. Depending on reaction conditions, these TiII species are proposed to be generated through Ti benzylidene or imido intermediates. A formally TiII complex, (ONO)Ti II(η2-C2H4)(HNMe2) (7), has been prepared and structurally characterized. © 2013 American Chemical Society.

  1. High-performance liquid chromatographic purification of oligomeric procyanidins, trimers up to nonamers, derived from the bark of Jatoba (Hymenaea courbaril).

    Science.gov (United States)

    Sasaki, Katsunori; Matsukura, Yasuko; Shijima, Kumiko; Miyake, Mika; Fujiwara, Daisuke; Konishi, Yutaka

    2009-06-01

    Procyanidin oligomers with different degrees of polymerization (up to nonamers) were efficiently purified from the bark of Jatoba (Hymenaea courbaril) by using a recently developed chromatographic separation method. Purification relied on a hydrogen bonding interaction between phenolic hydroxyl groups of the procyanidins and polyethylene glycol (PEG)-coated resin in a packed column. The individual procyanidins were identified by using electrospray ionization mass spectrometry (ESI-MS) and verified by a thiolytic degradation analysis. Our results demonstrate that Jatoba bark contained a large amount of procyanidins from monomer to nonamers or higher polymers composed of only B-type linked units (flavan-3-ol units linked through C-4 to C-8 (or C-6)) of epicatechin (EC) without gallate esters.

  2. Theoretical study of the structures and electron affinities of the dimers and trimers of the group IB metals (Cu, Ag, and Au)

    Science.gov (United States)

    Bauschlicher, Charles W., Jr.; Langhoff, Stephen R.; Partridge, Harry

    1989-01-01

    The molecular structure of both the neutral and negatively charged diatomic and triatomic systems containing the Cu, Ag, and Au metals are determined from ab initio calculations. For the neutral triatomic systems, the lowest energy structure is found to be triangular. The relative stability of the 2A1 and 2B2 structures can be predicted simply by knowing the constituent diatomic bond distances and atomic electron affinities (EAs). The lowest energy structure is linear for all of the negative ions. For anionic clusters containing Au, the Au atom(s) preferentially occupy the terminal position(s). The EAs of the heteronuclear systems can be predicted relatively accurately from a weighted average of the corresponding homonuclear systems. Although the theoretical EAs are systematically too small, accurate predictions for the EAs of the triatomics are obtained by uniformly scaling the ab initio results using the accurate experimental EA values available for the atoms and homonuclear diatomics.

  3. Protective Efficacy of Newcastle Disease Virus Expressing Soluble Trimeric Hemagglutinin against Highly Pathogenic H5N1 Influenza in Chickens and Mice

    NARCIS (Netherlands)

    Cornelissen, A.H.M.; Leeuw, de O.S.; Tacken, M.G.J.; Klos, H.C.; Vries, de R.P.; Boer-Luijtze, de E.A.; Zoelen-Bos, van D.J.; Rigter, A.; Rottier, P.J.M.; Moormann, R.J.M.; Haan, de C.A.M.

    2012-01-01

    Background: Highly pathogenic avian influenza virus (HPAIV) causes a highly contagious often fatal disease in poultry, resulting in significant economic losses in the poultry industry. HPAIV H5N1 also poses a major public health threat as it can be transmitted directly from infected poultry to

  4. Sequence analysis and molecular characterization of Clonorchis sinensis hexokinase, an unusual trimeric 50-kDa glucose-6-phosphate-sensitive allosteric enzyme.

    Directory of Open Access Journals (Sweden)

    Tingjin Chen

    Full Text Available Clonorchiasis, which is induced by the infection of Clonorchis sinensis (C. sinensis, is highly associated with cholangiocarcinoma. Because the available examination, treatment and interrupting transmission provide limited opportunities to prevent infection, it is urgent to develop integrated strategies to prevent and control clonorchiasis. Glycolytic enzymes are crucial molecules for trematode survival and have been targeted for drug development. Hexokinase of C. sinensis (CsHK, the first key regulatory enzyme of the glycolytic pathway, was characterized in this study. The calculated molecular mass (Mr of CsHK was 50.0 kDa. The obtained recombinant CsHK (rCsHK was a homotrimer with an Mr of approximately 164 kDa, as determined using native PAGE and gel filtration. The highest activity was obtained with 50 mM glycine-NaOH at pH 10 and 100 mM Tris-HCl at pH 8.5 and 10. The kinetics of rCsHK has a moderate thermal stability. Compared to that of the corresponding negative control, the enzymatic activity was significantly inhibited by praziquantel (PZQ and anti-rCsHK serum. rCsHK was homotropically and allosterically activated by its substrates, including glucose, mannose, fructose, and ATP. ADP exhibited mixed allosteric effect on rCsHK with respect to ATP, while inorganic pyrophosphate (PPi displayed net allosteric activation with various allosteric systems. Fructose behaved as a dose-dependent V activator with the substrate glucose. Glucose-6-phosphate (G6P displayed net allosteric inhibition on rCsHK with respect to ATP or glucose with various allosteric systems in a dose-independent manner. There were differences in both mRNA and protein levels of CsHK among the life stages of adult worm, metacercaria, excysted metacercaria and egg of C. sinensis, suggesting different energy requirements during different development stages. Our study furthers the understanding of the biological functions of CsHK and supports the need to screen for small molecule inhibitors of CsHK to interfere with glycolysis in C. sinensis.

  5. Theoretical study of weakly bound vibrational states of the sodium trimer. Numerical methods; prospects for the formation of Na3 in an ultracold gas

    International Nuclear Information System (INIS)

    Willner, K.

    2006-01-01

    A Mapped Fourier Grid method for solving the radial Schroedinger equation is improved. It is observed that a discrete sine and cosine transform algorithm allows to compute a Hamiltonian matrix the spectrum of which is free of spurious eigenvalues. - The energies of the highest, least bound vibrational states of the Na - Na 2 van der Waals complex are computed using a hyperspherical diabatic-by-sector method. The computed levels are analyzed using quantum defect theory. (orig.)

  6. Synthesis and Low Temperature Spectroscopic Observation of 1,3,5-Trioxane-2,4,6-Trione: The Cyclic Trimer of Carbon Dioxide

    Science.gov (United States)

    2016-08-19

    ozonolysis was then explored.27−29 The Criegee mechanism30 for this chemistry as applied to compound 2a is depicted in Scheme 4. Ozone and the olefin...Unsubstituted Triolefin- Trioxane Scheme 4. Criegee Ozonolysis Mechanism Applied to Compound 2aa aFor clarity, chemistry is represented on one of the three...carbon− carbon double bonds. The Journal of Organic Chemistry Article DOI: 10.1021/acs.joc.6b00647 J. Org. Chem. XXXX, XXX, XXX−XXX B the resultant

  7. Benchmark RI-MP2 database of nucleic acid base trimers: performance of different density functional models for prediction of structures and binding energies

    Czech Academy of Sciences Publication Activity Database

    Kabeláč, Martin; Valdes, Haydee; Sherer, E. C.; Cramer, Ch. J.; Hobza, Pavel

    2007-01-01

    Roč. 9, č. 36 (2007), s. 5000-5008 ISSN 1463-9076 R&D Projects: GA MŠk LC512; GA ČR GA203/05/0009 Grant - others:NSF(US) CHE-0615401 Institutional research plan: CEZ:AV0Z40550506 Keywords : IR spectrum * guanine * deoxyguanosine Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 3.343, year: 2007

  8. The dependency of twist-bend nematic liquid crystals on molecular structure: a progression from dimers to trimers, oligomers and polymers.

    Science.gov (United States)

    Mandle, Richard J

    2016-09-28

    This article gives an overview on recent developments concerning the twist-bend nematic phase. The twist-bend nematic phase has been discussed as the missing link between the uniaxial nematic mesophase (N) and the helical chiral nematic phase (N*). After an introduction discussing the key physical properties of the N TB phase and the methods used to identify the twist-bend nematic mesophase this review focuses on structure property relationships and molecular features that govern the incidence of this phase.

  9. Broadly neutralizing HIV antibodies define a glycan-dependent epitope on the prefusion conformation of gp41 on cleaved envelope trimers

    NARCIS (Netherlands)

    Falkowska, Emilia; Le, Khoa M.; Ramos, Alejandra; Doores, Katie J.; Lee, Jeong Hyun; Blattner, Claudia; Ramirez, Alejandro; Derking, Ronald; van Gils, Marit J.; Liang, Chi-Hui; McBride, Ryan; von Bredow, Benjamin; Shivatare, Sachin S.; Wu, Chung-Yi; Chan-Hui, Po-Ying; Liu, Yan; Feizi, Ten; Zwick, Michael B.; Koff, Wayne C.; Seaman, Michael S.; Swiderek, Kristine; Moore, John P.; Evans, David; Paulson, James C.; Wong, Chi-Huey; Ward, Andrew B.; Wilson, Ian A.; Sanders, Rogier W.; Poignard, Pascal; Burton, Dennis R.

    2014-01-01

    Broadly neutralizing HIV antibodies are much sought after (a) to guide vaccine design, both as templates and as indicators of the authenticity of vaccine candidates, (b) to assist in structural studies, and (c) to serve as potential therapeutics. However, the number of targets on the viral envelope

  10. In vitro kinetics of P700+reduction of Thermosynechococcus elongatus trimeric Photosystem I complexes by recombinant cytochrome c6using a Joliot-type LED spectrophotometer.

    Science.gov (United States)

    Nguyen, Khoa; Vaughn, Michael; Frymier, Paul; Bruce, Barry D

    2017-01-01

    The reduction rate of photo-oxidized Photosystem I (PSI) with various natural and artificial electron donors have been well studied by transient absorption spectroscopy. The electron transfer rate from various donors to P 700 + has been measured for a wide range of photosynthetic organisms encompassing cyanobacteria, algae, and plants. PSI can be a limiting component due to tedious extraction and purification methods required for this membrane protein. In this report, we have determined the in vivo, intracellular cytochrome c 6 (cyt c 6 )/PSI ratio in Thermosynechococcus elongatus (T.e.) using quantitative Western blot analysis. This information permitted the determination of P 700 + reduction kinetics via recombinant cyt c 6 in a physiologically relevant ratio (cyt c 6 : PSI) with a Joliot-type, LED-driven, pump-probe spectrophotometer. Dilute PSI samples were tested under varying cyt c 6 concentration, temperature, pH, and ionic strength, each of which shows similar trends to the reported literature utilizing much higher PSI concentrations with laser-based spectrophotometer. Our results do however indicate kinetic differences between actinic light sources (laser vs. LED), and we have attempted to resolve these effects by varying our LED light intensity and duration. The standardized configuration of this spectrophotometer will also allow a more uniform kinetic analysis of samples in different laboratories. We can conclude that our findings from the LED-based system display an added total protein concentration effect due to multiple turnover events of P 700 + reduction by cyt c 6 during the longer illumination regime.

  11. Towards the mechanism of trimeric purine nucleoside phosphorylases: Stopped-flow studies of binding of multisubstrate analogue inhibitor - 2-amino-9-[2-(phosphonomethoxy)ethyl]-6-sulfanylpurine

    Czech Academy of Sciences Publication Activity Database

    Wielgus-Kutrowska, B.; Antosiewicz, J. M.; Dlugosz, M.; Holý, Antonín; Bzowska, A.

    2007-01-01

    Roč. 125, 2/3 (2007), s. 260-268 ISSN 0301-4622 R&D Projects: GA AV ČR IBS4055109 Grant - others:Descartes Prize(XE) HPAW-CT-2002-9001 Institutional research plan: CEZ:AV0Z40550506 Keywords : PNP * PME-6-thio-Gua * fluorescence * stopped-flow Subject RIV: CC - Organic Chemistry Impact factor: 1.913, year: 2007

  12. Kinetics of binding of multisubstrate analogue inhibitor (2-amino-9-[2-(phosphonomethoxy)ethyl]-6-sulfanylpurine) with trimeric purine nucleoside phosphorylase

    Czech Academy of Sciences Publication Activity Database

    Antosiewicz, J.; Wielgus-Kutrowska, B.; Dlugosz, M.; Holý, Antonín; Bzowska, A.

    2007-01-01

    Roč. 26, 8/9 (2007), s. 969-974 ISSN 1525-7770 Institutional research plan: CEZ:AV0Z40550506 Keywords : binding * PME-6-thio-Gua * multisubstrate analogue inhibitor Subject RIV: CC - Organic Chemistry Impact factor: 0.723, year: 2007

  13. Circular trimers of gelatinase B/matrix metalloproteinase-9 constitute a distinct population of functional enzyme molecules differentially regulated by tissue inhibitor of metalloproteinases-1

    DEFF Research Database (Denmark)

    Vandooren, Jennifer; Born, Benjamin; Solomonov, Inna

    2015-01-01

    Gelatinase B/matrix metalloproteinase-9 (MMP-9) (EC 3.4.24.35) cleaves many substrates and is produced by most cell types as a zymogen, proMMP-9, in complex with the tissue inhibitor of metalloproteinases-1 (TIMP-1). Natural proMMP-9 occurs as monomers, homomultimers and heterocomplexes, but our...... knowledge about the overall structure of proMMP-9 monomers and multimers is limited. We investigated biochemical, biophysical and functional characteristics of zymogen and activated forms of MMP-9 monomers and multimers. In contrast with a conventional notion of a dimeric nature of MMP-9 homomultimers, we...

  14. Critical role for cross-linking of trimeric lectin domains of surfactant protein D in antiviral activity against influenza A virus

    DEFF Research Database (Denmark)

    Tecle, Tesfaldet; White, Mitchell R; Sørensen, Grith Lykke

    2008-01-01

    and antiviral activity of NCRDs as assessed by haemagglutination and neuraminidase inhibition and by viral neutralization. mAb-mediated cross-linking also enabled NCRDs to induce viral aggregation and to increase viral uptake by neutrophils and virus-induced respiratory burst responses by these cells...

  15. Role of the metal oxidation state in the SNS-Cr catalyst for ethylene trimerization: isolation of di- and trivalent cationic intermediates.

    Science.gov (United States)

    Jabri, Amir; Temple, Claire; Crewdson, Patrick; Gambarotta, Sandro; Korobkov, Ilia; Duchateau, Robbert

    2006-07-19

    The reaction of the highly selective [CySCH2CH2N(H)CH2CH2SCy]CrCl3 catalyst precursor with alkyl aluminum activators was examined with the aim of isolating reactive intermediates. Reaction with Me3Al afforded a cationic trivalent chromium alkyl species {[CySCH2CH2N(H)CH2CH2SCy]CrMe(mu-Cl)}2{(AlMe3)2(m-Cl}2.(C7H8)2 (1a). Although it was not possible to obtain crystalline samples of sufficient quality from the reaction with MAO (the most preferred activator), the near-to-identical EPR spectra indicated a very close structural similarity with 1a. Ethylene oligomerization tests clearly revealed that 1 and other cationic trivalent dimeric complexes {[CySCH2CH2N(H)CH2CH2SCy] CrCl(mu-Cl)}2{AlCl4}2.(C7H8)1.5 (2), monomeric [(CySCH2CH2N(H)CH2CH2SCy)CrCl2 (THF)][AlCl4] (3), and {[CySCH2CH2N(H)CH2CH2SCy]Cr(eta2-AlCl4)}{Al2Cl7} (4) adducts display the same catalyst selectivity as the [CySCH2CH2N(H)CH2CH2SCy]CrCl3 complex and, therefore, are probably all precursors to the same catalytically active species. 2, 3, and 4 were obtained upon treatment of [CySCH2CH2N(H)CH2CH2SCy] CrCl3 with different stoichiometric ratios of AlCl3.. When i-BAO activator was used, reduction of the metal center occurred readily, affording {([CySCH2CH2N(H)CH2CH2S Cy]Cr)(mu-Cl)]2}{(i-Bu)2AlCl2}2 (5). 5 is also a selective catalyst, thus indicating that trivalent species are most probably precursors to a divalent catalytically active complex. Reaction of CrCl2(THF)2 with the ligand afforded the labile divalent adduct [CySCH2CH2N(H)CH2CH2SCy]CrCl2(THF) (6), also catalytically active and selective. Instead, deprotonation of the ligand with n-BuLi followed by reaction with CrCl2(THF)2 gave the dinuclear complex [(mu-CySCH2CH2NCH2CH2SCy)CrCl]2 (7), which did not produce oligomers.

  16. The effect of detergents on trimeric G-protein activity in isolated plasma membranes from rat brain cortex: Correlation with studies of DPH and Laurdan fluorescence

    Czech Academy of Sciences Publication Activity Database

    Sýkora, Jan; Bouřová, Lenka; Hof, Martin; Svoboda, Petr

    2009-01-01

    Roč. 1788, č. 2 (2009), s. 324-332 ISSN 0005-2736 R&D Projects: GA MŠk(CZ) LC06063; GA MŠk(CZ) LC554; GA ČR(CZ) GA309/06/0121; GA AV ČR(CZ) IAA500110606 Institutional research plan: CEZ:AV0Z40400503; CEZ:AV0Z50110509 Keywords : detergent * fluorescence * stready-state * time-resolved Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 3.998, year: 2009

  17. Thermal instability of the trimeric structure of the N-terminal propeptide of human procollagen type I in relation to assay technology

    DEFF Research Database (Denmark)

    Brandt, J; Krogh, T N; Jensen, Charlotte Harken

    1999-01-01

    to measure the low-molecular weight form. Thermal transition of the high-molecular weight form caused a change in its elution volume but did not change its migration in sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Mass spectrometry revealed identical results for both forms. We reached...

  18. Plasma Membrane Density of GABA(B)-R1a, GABA(B)-R1b, GABA-R2 and Trimeric G-proteins in the Course of Postnatal Development of Rat Brain Cortex

    Czech Academy of Sciences Publication Activity Database

    Dlouhá, Kateřina; Kagan, Dmytro; Roubalová, Lenka; Ujčíková, Hana; Svoboda, Petr

    2013-01-01

    Roč. 62, č. 5 (2013), s. 547-559 ISSN 0862-8408 R&D Projects: GA ČR(CZ) GAP207/12/0919; GA ČR(CZ) GBP304/12/G069 Institutional support: RVO:67985823 Keywords : GABAB-receptors * postnatal development * rat brain cortex * G-proteins * Na, K- ATPase Subject RIV: CE - Biochemistry Impact factor: 1.487, year: 2013

  19. [(≢SiO)TaVCl2Me2]: A well-defined silica-supported tantalum(V) surface complex as catalyst precursor for the selective cocatalyst-free trimerization of ethylene

    KAUST Repository

    Chen, Yin

    2012-10-22

    On the surface of it: In the absence of co-catalyst, a well-defined silica-supported surface organometallic complex [(≢SiO)Ta VCl2Me2] selectively catalyzes the oligomerization of ethylene. The use of surface organometallic species allows three different pathways to be determined for the reduction of TaV to TaIII species under pressure of ethylene. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  20. Unusual Coexistence of Nickel(II) and Nickel(IV) in the Quadruple Perovskite Ba4Ni2Ir2O12 Containing Ir2NiO12 Mixed-Metal-Cation Trimers.

    Science.gov (United States)

    Ferreira, Timothy; Heald, Steve M; Smith, Mark D; Zur Loye, Hans-Conrad

    2018-03-19

    The crystal chemistry and magnetic properties of two hexagonal nickel(IV)-containing perovskites, Ba 4 Ni 1.94 Ir 2.06 O 12 and BaNiO 3 , are reported. The 12R perovskite, Ba 4 Ni 1.94 Ir 2.06 O 12 , possesses an unexpected coexistence of nickel(II) and nickel(IV). This quadruple perovskite structure contains Ir 2 NiO 12 mixed-metal-cation units in which direct metal-metal bonding between nickel(IV) and iridium(V) is inferred. X-ray absorption near-edge spectroscopy and X-ray photoelectron spectroscopy measurements were conducted to confirm the simultaneous presence of nickel(II) and nickel(IV).

  1. Long-Term Acclimation of the Cyanobacterium Synechocystis sp PCC 6803 to High Light Is Accompanied by an Enhanced Production of Chlorophyll That Is Preferentially Channeled to Trimeric Photosystem I

    Czech Academy of Sciences Publication Activity Database

    Kopečná, Jana; Komenda, Josef; Bučinská, Lenka; Sobotka, Roman

    2012-01-01

    Roč. 160, č. 4 (2012), s. 2239-2250 ISSN 0032-0889 R&D Projects: GA MŠk(CZ) ED2.1.00/03.0110; GA ČR GAP501/10/1000; GA ČR GBP501/12/G055 Institutional support: RVO:61388971 Keywords : CAB-LIKE-PROTEINS * SP PCC-6803 * PHOTOSYNTHETIC APPARATUS Subject RIV: EE - Microbiology, Virology Impact factor: 6.555, year: 2012

  2. The influence of monovalent cations on trimeric G protein Gi1alfa activity in HEK293 cells stably expressing DOR-Gi1alfa (Cys351-Ile351) fusion protein

    Czech Academy of Sciences Publication Activity Database

    Vošahlíková, Miroslava; Svoboda, Petr

    2011-01-01

    Roč. 60, č. 3 (2011), s. 541-547 ISSN 0862-8408 R&D Projects: GA AV ČR(CZ) IAA500110606; GA MŠk(CZ) LC554; GA ČR(CZ) GD305/08/H037 Institutional research plan: CEZ:AV0Z50110509 Keywords : delta-opioid receptor (DOR) * monovalent ions * G(i)1alfa protein Subject RIV: CE - Biochemistry Impact factor: 1.555, year: 2011

  3. Contribution to Casein Determination by UV Spectrophotometry

    Directory of Open Access Journals (Sweden)

    Stefanescu Raluca

    2017-12-01

    Full Text Available In the present paper, the interaction between copper ions and proteins is presented, in order to elaborate a simple and rapid spectrophotometric assay of casein in milk. Under alkaline conditions, copper ions form the biuret complex with the proteins, which can be used in protein determination. Although very specific, the biuret method is less sensitive. Using insoluble copper phosphate, casein is able to extract copper ions, with which it forms the biuret complex, while either the complex or copper ions could be determined in the ultraviolet range. Indeed, an increased absorbance of biuret complex at 215 nm was found. Nevertheless, copper ions can be determined in UV as well, their concentration being proportional to that of casein. When used tetraglycine instead casein, mass spectrometric measurements at pH higher than 11 revealed the formation of complexes with many copper ions bound to each peptide bond-containing molecule. Nevertheless, on diluting the biuret solution the complex may dissociate leading to very complex UV spectra that should be further studied.

  4. Global rescue of defects in HIV-1 envelope glycoprotein incorporation: implications for matrix structure.

    Directory of Open Access Journals (Sweden)

    Philip R Tedbury

    Full Text Available The matrix (MA domain of HIV-1 Gag plays key roles in membrane targeting of Gag, and envelope (Env glycoprotein incorporation into virions. Although a trimeric MA structure has been available since 1996, evidence for functional MA trimers has been elusive. The mechanism of HIV-1 Env recruitment into virions likewise remains unclear. Here, we identify a point mutation in MA that rescues the Env incorporation defects imposed by an extensive panel of MA and Env mutations. Mapping the mutations onto the putative MA trimer reveals that the incorporation-defective mutations cluster at the tips of the trimer, around the perimeter of a putative gap in the MA lattice into which the cytoplasmic tail of gp41 could insert. By contrast, the rescue mutation is located at the trimer interface, suggesting that it may confer rescue of Env incorporation via modification of MA trimer interactions, a hypothesis consistent with additional mutational analysis. These data strongly support the existence of MA trimers in the immature Gag lattice and demonstrate that rescue of Env incorporation defects is mediated by modified interactions at the MA trimer interface. The data support the hypothesis that mutations in MA that block Env incorporation do so by imposing a steric clash with the gp41 cytoplasmic tail, rather than by disrupting a specific MA-gp41 interaction. The importance of the trimer interface in rescuing Env incorporation suggests that the trimeric arrangement of MA may be a critical factor in permitting incorporation of Env into the Gag lattice.

  5. Formation of Sodium cyclo-Triphosphate by the Thermal Reaction of Sodium Dihydrogenphosphate-Organic Nitrogen Compound Mixtures; Rinsan nisuiso natoriumu-yuki chisso kagobutsu kongokei no kanetsu ni yoru cyclo-sanrinsan natoriumu no seisei

    Energy Technology Data Exchange (ETDEWEB)

    Takenaka, Atsushi.; Kikui, Satoshi. [Yonago National College of Technology, Tottori (Japan). Department of Materials Science; Motooka, Itaru.; Nariai, Hiroyuki. [Kobe University, Hyogo (Japan). Department of Chemical Science and Engineering

    1998-11-10

    Mixtures of sodium dihydrogenphosphate (NaH{sub 2}PO{sub 4}) with biuret, cyanuric acid, or melamine were heated in a stream of nitrogen gas. Crystalline sodium polyphosphate of high-temperature type of Maddrell`s salt was produced at above 300 degree C in the NaH{sub 2}PO{sub 4}-cyanuric acid or NaH{sub 2}PO{sub 4}-melamine mixtures. Sodium cyclo-trihosphate was produced in the NaH{sub 2}PO{sub 3}-biuret mixture in a similar manner to the thermal reaction of NaH{sub 2}PO{sub 4}-urea mixture reported previously, which was considered to be responsible for the enhanced water-vapor pressure in samples caused by the the formation of a kind of (shell.) An increase in amount of biuret added was available for the formation of cyclo-triphosphate. (author)

  6. Formation of Sodium cyclo-Triphosphate by the Thermal Reaction of Sodium Dihydrogenphosphate-Organic Nitrogen Compound Mixtures. Rinsan nisuiso natoriumu-yuki chisso kagobutsu kongokei no kanetsu ni yoru cyclo-sanrinsan natoriumu no seisei

    Energy Technology Data Exchange (ETDEWEB)

    Takenaka, Atsushi.; Kikui, Satoshi. (Yonago National College of Technology, Tottori (Japan). Department of Materials Science); Motooka, Itaru.; Nariai, Hiroyuki. (Kobe University, Hyogo (Japan). Department of Chemical Science and Engineering)

    1998-11-10

    Mixtures of sodium dihydrogenphosphate (NaH[sub 2]PO[sub 4]) with biuret, cyanuric acid, or melamine were heated in a stream of nitrogen gas. Crystalline sodium polyphosphate of high-temperature type of Maddrell's salt was produced at above 300 degree C in the NaH[sub 2]PO[sub 4]-cyanuric acid or NaH[sub 2]PO[sub 4]-melamine mixtures. Sodium cyclo-trihosphate was produced in the NaH[sub 2]PO[sub 3]-biuret mixture in a similar manner to the thermal reaction of NaH[sub 2]PO[sub 4]-urea mixture reported previously, which was considered to be responsible for the enhanced water-vapor pressure in samples caused by the the formation of a kind of (shell.) An increase in amount of biuret added was available for the formation of cyclo-triphosphate. (author)

  7. 3-(2,3,5,6,7,8-Hexahydro-1H-cyclopenta[b]quinolin-9-yl-1,5-bis(4-methoxyphenylbiuret

    Directory of Open Access Journals (Sweden)

    Koichiro Nishibe

    2010-03-01

    Full Text Available Ipidacrine (2,3,5,6,7,8-hexahydro-1H-cyclopenta[b]quinolin-9-amine was reacted with 4-methoxyphenyl isocyanate to give the title compound, C28H30N4O4. An intramolecular N—H...O hydrogen bond results in an essentially planar [r.m.s. deviation from the mean plane is 0.126 (1 Å] conformation for the biuret unit. The central ring of the quinoline unit is twisted by 78.2 (1° with respect to the biuret mean plane, whereas the two 4-methoxybenzene rings are twisted out of this plane by 24.3 (1° and 48.5 (1°, resulting in an overall propeller-like structure. An intermolecular N—H...N hydrogen bond between the biuret NH atom and the quinoline ring nitrogen defines the crystal packing.

  8. Thermolysis of urea complexes of uranyl nitrate

    International Nuclear Information System (INIS)

    Kostyuk, N. N.

    2005-01-01

    Quantitative parameters of thermolysis of uranyl nitrate urea complexes, [UO 2 (NO 3 ) 2 {(NH 2 ) 2 CO} 2 ], [UO 2 (H 2 O){(NH 2 ) 2 CO} 4 ](NO 3 ) 2 , and [UO 2 (H 2 O){(NH 2 ) 2 CO} 5 ](NO 3 ) 2 at 175, 200, and 225 deg C were measured. Thermolysis of [UO 2 (NO 3 ) 2 {(NH 2 ) 2 CO} 2 ] at 200 deg C affords the biuret complex of uranyl nitrate in a 90% yield. The urea ligands in the hydrated complexes completely transform into biuret at 175 deg C. Thermolysis of [UO 2 (H 2 O){(NH 2 ) 2 CO} 5 ](NO 3 ) 2 yields the biuret-cyanurate complexes of uranyl nitrate. The features of thermolysis of the uranyl nitrate complexes originate from the chemical transformations of urea at elevated temperatures [ru

  9. Clinical performance evaluation of total protein measurement by digital refractometry and characterization of non-protein solute interferences

    Directory of Open Access Journals (Sweden)

    Joshua J.H. Hunsaker

    2016-12-01

    Full Text Available Objectives: Refractometric methods to measure total protein (TP in serum and plasma specimens have been replaced by automated biuret methods in virtually all routine clinical testing. A subset of laboratories, however, still report using refractometry to measure TP in conjunction with serum protein electrophoresis. The objective of this study was therefore to conduct a modern performance evaluation of a digital refractometer for TP measurement. Design and methods: Performance evaluation of a MISCO Palm Abbe™ digital refractometer was conducted through device familiarization, carryover, precision, accuracy, linearity, analytical sensitivity, analytical specificity, and reference interval verification. Comparison assays included a manual refractometer and an automated biuret assay. Results: Carryover risk was eliminated using a demineralized distilled water (ddH2O wash step. Precision studies demonstrated overall imprecision of 2.2% CV (low TP pool and 0.5% CV (high TP pool. Accuracy studies demonstrated correlation to both manual refractometry and the biuret method. An overall positive bias (+5.0% was observed versus the biuret method. On average, outlier specimens had an increased triglyceride concentration. Linearity was verified using mixed dilutions of: a low and high concentration patient pools, or b albumin-spiked ddH2O and high concentration patient pool. Decreased recovery was observed using ddH2O dilutions at low TP concentrations. Significant interference was detected at high concentrations of glucose (>267 mg/dL and triglycerides (>580 mg/dL. Current laboratory reference intervals for TP were verified. Conclusions: Performance characteristics of this digital refractometer were validated in a clinical laboratory setting. Biuret method remains the preferred assay for TP measurement in routine clinical analyses. Keywords: Refractometry, Digital refractometry, Total protein, Biuret, Serum protein electrophoresis, Monoclonal

  10. 尿素の加熱によるトリウレットの合成とその熱分解

    OpenAIRE

    島崎, 長一郎; 橋本, 重治

    1982-01-01

    The formation of triuret on heating urea prepared by three different conditions, such as untreated, in 36% HCl atmosphere (A series: chlorine content 1--2%, standing time 1 hr.; B series: 8--12%, 3 hr,; C series: 25--30%, 20 hr.), and the urea-biuret mixtures were studied and the obtained triuret was examined by utilizing the DTA, TG, and IR techniques in order to follow the course of thermal decomposition.It was found that in untreated urea, the formation of biuret was attached to the maximu...

  11. Dynamical Negative Differential Resistance in Antiferromagnetically Coupled Few-Atom Spin Chains

    Science.gov (United States)

    Rolf-Pissarczyk, Steffen; Yan, Shichao; Malavolti, Luigi; Burgess, Jacob A. J.; McMurtrie, Gregory; Loth, Sebastian

    2017-11-01

    We present the appearance of negative differential resistance (NDR) in spin-dependent electron transport through a few-atom spin chain. A chain of three antiferromagnetically coupled Fe atoms (Fe trimer) was positioned on a Cu2 N /Cu (100 ) surface and contacted with the spin-polarized tip of a scanning tunneling microscope, thus coupling the Fe trimer to one nonmagnetic and one magnetic lead. Pronounced NDR appears at the low bias of 7 mV, where inelastic electron tunneling dynamically locks the atomic spin in a long-lived excited state. This causes a rapid increase of the magnetoresistance between the spin-polarized tip and Fe trimer and quenches elastic tunneling. By varying the coupling strength between the tip and Fe trimer, we find that in this transport regime the dynamic locking of the Fe trimer competes with magnetic exchange interaction, which statically forces the Fe trimer into its high-magnetoresistance state and removes the NDR.

  12. Catalytic activity of bovine glutamate dehydrogenase requires a hexamer structure.

    OpenAIRE

    Bell, E T; Bell, J E

    1984-01-01

    Previous workers have shown that the hexamers of glutamate dehydrogenase are dissociated first into trimers and subsequently into monomers by increasing guanidinium chloride concentrations. In renaturation experiments it is shown that trimers of glutamate dehydrogenase can be reassociated to give the hexamer form of the enzyme, with full regain of activity. Monomeric subunits produced at high guanidinium chloride concentrations cannot be renatured. The trimer form of the enzyme is shown to ha...

  13. Download this PDF file

    African Journals Online (AJOL)

    Serumtotal proteins and albumin were assayed by molecular absorption photometry with an. Eppendorf spectrophotometer, model 1101 M or, for some samples, with a Cobas apparatus (MIRA. Plus ABX Diagnostics). The assay of total proteins was based on the principle of the Biuret reaction while the determination of ...

  14. Liver function of Streptozotocin- Induced Diabetic Rats Orally ...

    African Journals Online (AJOL)

    Klinische Chemie (Rec. GSCC DGKC) (1972) and the. Szasz (1969) methods respectively. Serum and liver protein concentrations were measured by Biuret method (Gornall et al., 1949), serum albumins concentration was measured by Doumas and Biggs. (1972) method while the amount of globulins was calculated as a ...

  15. Biosynthesis and Characterization of Glucose Syrup Derived from ...

    African Journals Online (AJOL)

    In order to characterize the prepared glucose syrups, Fehling test, Biuret test, glucose concentration (GC), dextrose equivalent (DE), relative density (RD), pH, viscosity and surface tension (ST) were determined. Granular size range of 7 – 15 μm and moisture content of 12 % were determined for the untreated starch sample.

  16. Determinação de proteínas totais via espectrofometria: vantagens e desvantagens dos métodos existentes Determination of total protein by spectrophotometry: advantages and disadvantages of proposed methods

    Directory of Open Access Journals (Sweden)

    Dimas A. M. Zaia

    1998-11-01

    Full Text Available Spectrophotometric determination of total protein is used in several areas such as clinical analysis, food science and technology, biochemistry, protein chemistry, physiology. Five spectrophotometric methods are mostly used: biuret, Lowry, Bradford, Smith and UV absorption. In this review a general overview of these methods is presented (interferences, applications; other methodologies are also discussed.

  17. MorOgoro, Tanzania ! / ". . . "

    African Journals Online (AJOL)

    'ctive per-. '1 formance,. . -. , ~: .... 9r nerdsma.n, ~j milk~ng.feeding .and cleaning kinetic);· Total prot~in by Biuret: Albu~in bv ti"m~s.did;oestru~d(;!tection, ... chemical composition (crude protein (CP)i1nd ash on DM basis) of the major feeds for ...

  18. Determinação de proteínas totais via espectrofometria: vantagens e desvantagens dos métodos existentes

    OpenAIRE

    Zaia,Dimas A. M.; Zaia,Cássia Thaïs B. V.; Lichtig,Jaim

    1998-01-01

    Spectrophotometric determination of total protein is used in several areas such as clinical analysis, food science and technology, biochemistry, protein chemistry, physiology. Five spectrophotometric methods are mostly used: biuret, Lowry, Bradford, Smith and UV absorption. In this review a general overview of these methods is presented (interferences, applications); other methodologies are also discussed.

  19. THE EXTRACTION OF THE CELL CONTENT OF MICRO-ORGANISMS BY PERCUSSION OF THE FROZEN CELLS

    Science.gov (United States)

    Johlin, J. M.; Avery, Roy C.

    1930-01-01

    A method is described for rapidly obtaining fresh extracts of microorganisms by percussion of the frozen cells. Filtrates giving the biuret reaction and yielding heavy precipitates upon the addition of acetic acid were obtained from the washed cells of cultures of yeast, hemolytic streptococcus, pneumococcus, B. coli, B. xerosis, B. dyphtheriae and the bacillus of bovine tuberculosis. PMID:19869774

  20. Download this PDF file

    African Journals Online (AJOL)

    olayemitoyin

    Kingsley, G.R. (1942) The direct Biuret method for the determination of proteins as applied to photoelectric and visual colorimetry. Journal of. Laboratory Clinical Medicine. 27:840-845. Kushner, I., (1993): Semantics, Inflammation. Cytokines and Common Sense, Cytokine Growth. Factor; Prospect Biomedical 9:196-9.

  1. Gamma irradiation of cytosine in aerated aqueous solution. Identification of two new radiolytic products

    International Nuclear Information System (INIS)

    Polverelli, M.

    1975-01-01

    After gamma irradiation of cytosine in aerated aqueous solutions, two new radiolytic products were isolated and identified: N-formyl-biuret and carbamoyl-l-hydroxy-5-hydantoin. The position and importance of those products in the radiolytic degradation of cytosine were examined [fr

  2. Can serum concentration of C-reactive protein, albumin and body ...

    African Journals Online (AJOL)

    The C-reactive protein was estimated using semi-quantitative method, albumin level was estimated using bromocresol green method, and biuret method was used for total protein. The CD4+T-cell count of the subjects was determined using CyFlow Analyzer while their weights were measured using high precision weighing ...

  3. South African Journal of Animal Science - Vol 6, No 1 (1976)

    African Journals Online (AJOL)

    Invloed van die byvoeging van ureum, kalk of biuret tot mieliekuilvoer gesny op twee groeistadia · EMAIL FREE FULL TEXT EMAIL FREE FULL TEXT DOWNLOAD FULL TEXT DOWNLOAD FULL TEXT. J.W. Cilliers, A.P. van Schalkwyk, 29-33 ...

  4. Measurement of some blood proteins of children as an index of ...

    African Journals Online (AJOL)

    Serum total proteins and albumin of 83 Ghanaian children whose ages ranged from less than one year to thirteen years were measured using the biuret and bromocresol green dye binding methods respectively. The globulin levels were obtained by difference. The range obtained for total protein was 54.7-87.9 g/l, that of ...

  5. Experimental Work in Biology: Book 1, Food Tests; Book 2, Enzymes; Book 3, Soil.

    Science.gov (United States)

    Mackean, D. G.

    Laboratory experiments are presented in these first three manuals of a six-volume series for use at the 12- to 16-year-old, or British CSE, level. On the subject of food tests, 17 exercises are prepared in connection with Biuret reactions, starch and emulsion tests, Millon's and Benedict's reagents, reagent sensitivity, and calorific values and…

  6. Prevalence of HBsAg Among Prospective Blood Donors and ...

    African Journals Online (AJOL)

    methods for the measurement of catalytic concentration of enzymes. Part 5.IFCC method for alkaline phosphatase (orthophosphoric-monoester phosphohydrolase, alkaline optimum, EC 3.1.3.1). Journal of Clinical Chemistry and Clinical. Biochemistry, 21: 731-48. Weichselbaum, T.E. (1946). Biuret method of serum total.

  7. Gamma radiolysis of aerated aqueous solution of cytosine. Pt. 3

    International Nuclear Information System (INIS)

    Polverelli, M.; Teoule, R.

    1976-01-01

    In gamma-irradiation of cytosine in aerated aqueous solution, two new radiolysis products were isolated and identified: 1-carbamoyl-5-hydroxyhydantoin and N-formyl-biuret. This formation involves ring cleavage between C4 and C5 and belongs to the nondeamination pathway. (orig.) [de

  8. Some liver function indices and blood parameters in T. brucei ...

    African Journals Online (AJOL)

    JTEkanem

    (RBC), white blood cell (WBC), and platelet count were determined using the automated haematologic analyzer SYSMEX KX21,a product of SYSMEX corporation, Japan employing the method described by Dacie and. Lewis17. Protein concentrations were determined using biuret method18 as described by Plummer19.

  9. Author Details

    African Journals Online (AJOL)

    Mieliemeelbyvoeding aan jong osse op somerveld en verskillende winter voedingspeile. Abstract PDF · Vol 6, No 1 (1976) - Articles Invloed van die byvoeging van ureum, kalk of biuret tot mieliekuilvoer gesny op twee groeistadia. Abstract PDF · Vol 5, No 3 (1975) - Articles Verskillende verhoudings van kuilvoer en kragvoer ...

  10. Exam Question Exchange.

    Science.gov (United States)

    Alexander, John J., Ed.

    1988-01-01

    Describes three examination questions which could be used in college chemistry courses. Discusses each problem and gives acceptable solutions. Problems include: "A Multi-Topic Problem for General Chemistry"; "Consumption of Air by Biuret Reagent--a Question Involving Experimental Design"; and "An Instructive Problem in Heterogeneous Equilibrium."…

  11. Effect of thermo-oxidized soybean oil on growth rate and skeletal ...

    African Journals Online (AJOL)

    Dr. J. T. Ekanem

    2004-07-15

    Jul 15, 2004 ... mortar and pestle. The homogenate was then frozen to be analyzed for protein and enzymes. Protein concentration determination. The skeletal muscle was homogenized with. 0.25M sucrose buffer to make 30 times dilution. The protein concentration in the tissue was then determined using Biuret method4.

  12. The effect of urea on refractometric total protein measurement in dogs and cats with azotemia.

    Science.gov (United States)

    Legendre, Kelsey P; Leissinger, Mary; Le Donne, Viviana; Grasperge, Britton J; Gaunt, Stephen D

    2017-03-01

    While protein is the predominant solute measured in plasma or serum by a refractometer, nonprotein substances also contribute to the angle of refraction. There is debate in the current literature regarding which nonprotein substances cause factitiously high refractometric total protein measurements, as compared to the biuret assay. The purpose of the study was to determine if the blood of azotemic animals, specifically with increased blood urea concentration, will have significantly higher refractometric total protein concentrations compared to the total protein concentrations measured by biuret assay. A prospective case series was conducted by collecting data from azotemic (n = 26) and nonazotemic (n = 34) dogs and cats. In addition, an in vitro study was performed where urea was added to an enhanced electrolyte solution at increasing concentrations, and total protein was assessed by both the refractometer and spectrophotometer. Statistical analysis was performed to determine the effect of urea. The refractometric total protein measurement showed a positive bias when compared to the biuret protein measurement in both groups, but the bias was higher in the azotemic group vs the nonazotemic group. The mean difference in total protein measurements of the nonazotemic group (0.59 g/dL) was significantly less (P biuret assay, both in vivo and in vitro. © 2017 American Society for Veterinary Clinical Pathology.

  13. Temperature dependent Raman spectroscopy of melamine and structural analogs in milk powder

    Science.gov (United States)

    Hyperspectral Raman imaging has the potential for rapid screening of solid-phase samples for potential adulterants. We found that the Raman spectra of melamine analogs changed dramatically and uniquely as a function of elevated temperature. Raman spectra were acquired for urea, biuret, cyanuric acid...

  14. Modelling of the Blood Plasma Species of Biguanide Derivatives ...

    African Journals Online (AJOL)

    99mTc-DMSA (DMSA=dimercaptosuccinic acid), the gold standard for static renal imaging, has a long uptake time, which is a limiting factor in diagnostic procedures and also leads to a relatively high radiation dose to patients. The ligands dimethyl biguanide (DMBG), biuret (BIU), 2-imino-4-thiobiuret (ITB) and ...

  15. Prevalence of Persistent Proteinuria using Urine Protein/Creatinine ...

    African Journals Online (AJOL)

    Those with persistent proteinuria were quantitated using the modified Biuret method while urine creatinine was measured with the modified Jaffe Kinetic method (done with the Biolabo creatinine kit). Urine protein:creatinine ratios > 0.20 were confirmed as persistent proteinuria. Results: Out of the 1,600 pupil, 761(47.6%) ...

  16. ON THE DIALYSABILITY OF THE GROWTH-ACTIVATING PRINCIPLE CONTAINED IN EXTRACTS OF EMBRYONIC TISSUES

    Science.gov (United States)

    Wright, G. Payling

    1926-01-01

    These experiments appear to indicate that whatever substance in embryonic tissue extracts excites mitosis, and therefore would appear to be the growth-stimulating substance, is capable of passing through a collodion membrane which is impermeable to proteins, at all events in such concentration as would give rise to any biuret reaction. The substance is therefore diffusible. PMID:19869146

  17. [Abnormal Serum Total Protein Measurement by Lipoprotein-X in an Infant with Biliary Atresia].

    Science.gov (United States)

    Futatsugi, Akiko; Hidaka, Eiko; Kubota, Noriko; Nishijima, Fumie; Yoshizawa, Katsumi; Ishimine, Nau; Sugano, Mitsutoshi; Hori, Atsushi; Hidaka, Hiroya

    2015-11-01

    Lipoprotein-X (LP-X) in cholestatic jaundice causes abnormal reaction in assays for low-density lipoprotein-cholesterol, but the effects on other test items are unknown. Here, we report an infant with biliary atresia showing abnormal reaction in total serum protein assay using the biuret method, and lipoprotein-X (LP-X) was then detected. In this 11-month-old female infant, jaundice was observed at 2 months old, and a diagnosis of biliary atresia was made. On biochemical tests at 12 months old, the total serum protein concentrations detected by the biuret method were very high, and the response curve and linearity of dilution were abnormal. LP-X was detected by agar electrophoresis. In addition and recovery experiments with normal serum fractionation of the patient's LP-X-rich lipoprotein fraction prepared by ultracentrifugation, normal γ-globulin fractionation showed an abnormal reaction by the biuret method. In infants with biliary atresia, we showed that the total serum protein assay by the biuret method was influenced by LP-X-rich lipoprotein, which may be caused by abnormal reaction of LP-X and γ-globulin. [Case Report].

  18. Spectrophotometric determination of proteins associated with ...

    African Journals Online (AJOL)

    Twenty six Aeromonas isolates from fishes, poultry and humans in Zaria were quantified for total soluble proteins (enzymes) profiles in January, 2007 by spectrophotometric (Biuret) method. Isolates were grown on Brain Heart Infusion (BHI) broth, they were incubated at 370C and centrifuged at 1,000 g/dl using harous ...

  19. Comparison of effects of age and sex on serum protein ...

    African Journals Online (AJOL)

    The aim of this study was to evaluate the influence of age and sex on the concentration of total serum protein measured by the biuret method and protein fractions determined using cellulose acetate electrophoresis in apparently healthy camels (Camelus dromedarius). Blood samples were collected from 21 camels (12 ...

  20. Clinical performance evaluation of total protein measurement by digital refractometry and characterization of non-protein solute interferences.

    Science.gov (United States)

    Hunsaker, Joshua J H; Wyness, Sara P; Snow, Taylor M; Genzen, Jonathan R

    2016-12-01

    Refractometric methods to measure total protein (TP) in serum and plasma specimens have been replaced by automated biuret methods in virtually all routine clinical testing. A subset of laboratories, however, still report using refractometry to measure TP in conjunction with serum protein electrophoresis. The objective of this study was therefore to conduct a modern performance evaluation of a digital refractometer for TP measurement. Performance evaluation of a MISCO Palm Abbe™ digital refractometer was conducted through device familiarization, carryover, precision, accuracy, linearity, analytical sensitivity, analytical specificity, and reference interval verification. Comparison assays included a manual refractometer and an automated biuret assay. Carryover risk was eliminated using a demineralized distilled water (ddH 2 O) wash step. Precision studies demonstrated overall imprecision of 2.2% CV (low TP pool) and 0.5% CV (high TP pool). Accuracy studies demonstrated correlation to both manual refractometry and the biuret method. An overall positive bias (+5.0%) was observed versus the biuret method. On average, outlier specimens had an increased triglyceride concentration. Linearity was verified using mixed dilutions of: a) low and high concentration patient pools, or b) albumin-spiked ddH 2 O and high concentration patient pool. Decreased recovery was observed using ddH 2 O dilutions at low TP concentrations. Significant interference was detected at high concentrations of glucose (>267 mg/dL) and triglycerides (>580 mg/dL). Current laboratory reference intervals for TP were verified. Performance characteristics of this digital refractometer were validated in a clinical laboratory setting. Biuret method remains the preferred assay for TP measurement in routine clinical analyses.

  1. An Asymmetric Deuterium Labeling Strategy to Identify Interprotomer and Intraprotomer NOEs in Oligomeric Proteins

    International Nuclear Information System (INIS)

    Jasanoff, Alan

    1998-01-01

    A major difficulty in determining the structure of an oligomeric protein by NMR is the problem of distinguishing inter- from intraprotomer NOEs. In order to address this issue in studies of the 27 kD compact trimeric domain of the MHC class II-associated invariant chain, we compared the 13C NOESY-HSQC spectrum of a uniformly 13C-labeled trimer with the spectrum of the same trimer labeled with 13C in only one protomer, and with deuterium in the other two protomers. The spectrum of the unmixed trimer included both inter- and intraprotomer NOEs while the spectrum of the mixed trimer included only intraprotomer peaks. NOEs clearly absent from the spectrum of the mixed trimer could be confidently assigned to interprotomer interactions. Asymmetrically labeled trimers were isolated by refolding a 13C-labeled shorter form of the protein with a 2H-labeled longer form, chromatographically purifying trimers with only one short chain, and then processing with trypsin to yield only protomers with the desired N- and C-termini. In contrast to earlier studies, in which statistical mixtures of differently labeled protomers were analyzed, our procedure generated only a well-defined 1:2 oligomer, and no other mixed oligomers were present. This increased the maximum possible concentration of NMR-active protomers and thus the sensitivity of the experiments. Related methods should be applicable to many oligomeric proteins, particularly those with slow protomer exchange rates

  2. Species Differences in the Carbohydrate Binding Preferences of Surfactant Protein D

    DEFF Research Database (Denmark)

    Crouch, Erika C.; Smith, Kelly; McDonald, Barbara

    2006-01-01

    Interactions of surfactant protein D (SP-D) with micro-organisms and organic antigens involve binding to the trimeric neck plus carbohydrate recognition domain (neck+CRD). In these studies, we compared the ligand binding of homologous human, rat, and mouse trimeric neck+CRD fusion proteins, each...

  3. Species Differences in the Carbohydrate Binding Preferences of Surfactant Protein D

    DEFF Research Database (Denmark)

    Crouch, Erika C.; Smith, Kelly; McDonald, Barbara

    2006-01-01

    Interactions of surfactant protein D (SP-D) with micro-organisms and organic antigens involve binding to the trimeric neck plus carbohydrate recognition domain (neck+CRD). In these studies, we compared the ligand binding of homologous human, rat, and mouse trimeric neck+CRD fusion proteins, each ...

  4. The HIV-1 envelope glycoprotein structure: nailing down a moving target.

    Science.gov (United States)

    Ward, Andrew B; Wilson, Ian A

    2017-01-01

    Structure determination of the HIV-1 envelope glycoprotein (Env) presented a number of challenges, but several high-resolution structures have now become available. In 2013, cryo-EM and x-ray structures of soluble, cleaved SOSIP Env trimers from the clade A BG505 strain provided the first glimpses into the Env trimer fold as well as more the variable regions. A recent cryo-EM structure of a native full-length trimer without any stabilizing mutations had the same core structure, but revealed new insights and features. A more comprehensive and higher resolution understanding of the glycan shield has also emerged, enabling a more complete representation of the Env glycoprotein structure. Complexes of Env trimers with broadly neutralizing antibodies have surprisingly illustrated that most of the Env surface can be targeted in natural infection and that the neutralizing epitopes are almost all composed of both peptide and glycan components. These structures have also provided further evidence of the inherent plasticity of Env and how antibodies can exploit this flexibility by perturbing or even stabilizing the trimer to facilitate neutralization. These breakthroughs have stimulated further design and stabilization of Env trimers as well as other platforms to generate trimers that now span multiple subtypes. These Env trimers when used as immunogens, have led to the first vaccine-induced neutralizing antibodies for structural and functional analyses. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  5. C-C bond formation and related reactions at the CNC backbone in (smif)FeX (smif = 1,3-di-(2-pyridyl)-2-azaallyl): dimerizations, 3 + 2 cyclization, and nucleophilic attack; transfer hydrogenations and alkyne trimerization (X = N(TMS)2, dpma = (di-(2-pyridyl-methyl)-amide)).

    Science.gov (United States)

    Frazier, Brenda A; Williams, Valerie A; Wolczanski, Peter T; Bart, Suzanne C; Meyer, Karsten; Cundari, Thomas R; Lobkovsky, Emil B

    2013-03-18

    Molecular orbital analysis depicts the CNC(nb) backbone of the smif (1,3-di-(2-pyridyl)-2-azaallyl) ligand as having singlet diradical and/or ionic character where electrophilic or nucleophilic attack is plausible. Reversible dimerization of (smif)Fe{N(SiMe3)2} (1) to [{(Me3Si)2N}Fe]2(μ-κ(3),κ(3)-N,py2-smif,smif) (2) may be construed as diradical coupling. A proton transfer within the backbone-methylated, and o-pyridine-methylated smif of putative ((b)Me2(o)Me2smif)FeN(SiMe3)2 (8) provides a route to [{(Me3Si)2N}Fe]2(μ-κ(4),κ(4)-N,py2,C-((b)Me,(b)CH2,(o)Me2(smif)H))2 (9). A 3 + 2 cyclization of ditolyl-acetylene occurs with 1, leading to the dimer [{2,5-di(pyridin-2-yl)-3,4-di-(p-tolyl-2,5-dihydropyrrol-1-ide)}FeN(SiMe3)2]2 (11), and the collateral discovery of alkyne cyclotrimerization led to a brief study that identified Fe(N(SiMe3)2(THF) as an effective catalyst. Nucleophilic attack by (smif)2Fe (13) on (t)BuNCO and (2,6-(i)Pr2C6H3)NCO afforded (RNHCO-smif)2Fe (14a, R = (t)Bu; 14b, 2,6-(i)PrC6H3). Calculations suggested that (dpma)2Fe (15) would favorably lose dihydrogen to afford (smif)2Fe (13). H2-transfer to alkynes, olefins, imines, PhN═NPh, and ketones was explored, but only stoichiometric reactions were affected. Some physical properties of the compounds were examined, and X-ray structural studies on several dinuclear species were conducted.

  6. Crystal structure, conformational fixation and entry-related interactions of mature ligand-free HIV-1 Env.

    Science.gov (United States)

    Kwon, Young Do; Pancera, Marie; Acharya, Priyamvada; Georgiev, Ivelin S; Crooks, Emma T; Gorman, Jason; Joyce, M Gordon; Guttman, Miklos; Ma, Xiaochu; Narpala, Sandeep; Soto, Cinque; Terry, Daniel S; Yang, Yongping; Zhou, Tongqing; Ahlsen, Goran; Bailer, Robert T; Chambers, Michael; Chuang, Gwo-Yu; Doria-Rose, Nicole A; Druz, Aliaksandr; Hallen, Mark A; Harned, Adam; Kirys, Tatsiana; Louder, Mark K; O'Dell, Sijy; Ofek, Gilad; Osawa, Keiko; Prabhakaran, Madhu; Sastry, Mallika; Stewart-Jones, Guillaume B E; Stuckey, Jonathan; Thomas, Paul V; Tittley, Tishina; Williams, Constance; Zhang, Baoshan; Zhao, Hong; Zhou, Zhou; Donald, Bruce R; Lee, Lawrence K; Zolla-Pazner, Susan; Baxa, Ulrich; Schön, Arne; Freire, Ernesto; Shapiro, Lawrence; Lee, Kelly K; Arthos, James; Munro, James B; Blanchard, Scott C; Mothes, Walther; Binley, James M; McDermott, Adrian B; Mascola, John R; Kwong, Peter D

    2015-07-01

    As the sole viral antigen on the HIV-1-virion surface, trimeric Env is a focus of vaccine efforts. Here we present the structure of the ligand-free HIV-1-Env trimer, fix its conformation and determine its receptor interactions. Epitope analyses revealed trimeric ligand-free Env to be structurally compatible with broadly neutralizing antibodies but not poorly neutralizing ones. We coupled these compatibility considerations with binding antigenicity to engineer conformationally fixed Envs, including a 201C 433C (DS) variant specifically recognized by broadly neutralizing antibodies. DS-Env retained nanomolar affinity for the CD4 receptor, with which it formed an asymmetric intermediate: a closed trimer bound by a single CD4 without the typical antigenic hallmarks of CD4 induction. Antigenicity-guided structural design can thus be used both to delineate mechanism and to fix conformation, with DS-Env trimers in virus-like-particle and soluble formats providing a new generation of vaccine antigens.

  7. Herceptin conjugates linked by EDC boost direct tumor cell death via programmed tumor cell necrosis.

    Directory of Open Access Journals (Sweden)

    Jiemiao Hu

    Full Text Available Tumor-targeted antibody therapy is one of the safest biological therapeutics for cancer patients, but it is often ineffective at inducing direct tumor cell death and is ineffective against resistant tumor cells. Currently, the antitumor efficacy of antibody therapy is primarily achieved by inducing indirect tumor cell death, such as antibody-dependent cell cytotoxicity. Our study reveals that Herceptin conjugates, if generated via the crosslinker EDC (1-ethyl-3-(3-dimethylaminopropyl carbodiimide hydrochloride, are capable of engendering human epidermal growth factor receptor 2 (Her2 positive tumor cells death. Using a high-performance liquid chromatography (HPLC system, three peaks with estimated molecular weights of antibody monomer, dimer, and trimer were isolated. Both Herceptin trimer and dimer separated by HPLC induced significant levels of necrotic tumor cell death, although the trimer was more effective than the dimer. Notably, the Herceptin trimer also induced Herceptin-resistant tumor cell death. Surprisingly different from the known cell death mechanism that often results from antibody treatment, the Herceptin trimer elicited effective and direct tumor cell death via a novel mechanism: programmed cell necrosis. In Her2-positive cells, inhibition of necrosis pathways significantly reversed Herceptin trimer-induced cell death. In summary, the Herceptin trimer reported herein harbors great potential for overcoming tumor cell resistance to Herceptin treatment.

  8. Density function theoretical investigation on the Ni3PP structure and the hydrogen adsorption property of the Ni2P(0001) surface

    OpenAIRE

    Ariga, Hiroko; Kawashima, Mayumi; Takakusagi, Satoru; Asakura, Kiyotaka

    2013-01-01

    The electronic and structural properties of a phosphorus-terminated structure of Ni2P(0001) surface (Ni3PP) are investigated by density functional theory (DFT) calculations. Phosphorus adsorption largely stabilizes the Ni2P(0001) surface by creating Ni-P bonds on the Ni trimer. Atomic hydrogen can adsorb on the topmost P site although its adsorption energy is much lower than its adsorption energy on the Ni trimer site of the Ni3P2 surface. Our results suggest that the Ni trimer is the key fac...

  9. Quantification and Statistical Modeling—Part I: Breathing-Zone Concentrations of Monomeric and Polymeric 1,6-Hexamethylene Diisocyanate

    Science.gov (United States)

    Fent, Kenneth W.; Gaines, Linda G. Trelles; Thomasen, Jennifer M.; Flack, Sheila L.; Ding, Kai; Herring, Amy H.; Whittaker, Stephen G.; Nylander-French, Leena A.

    2009-01-01

    We conducted a repeated exposure-assessment survey for task-based breathing-zone concentrations (BZCs) of monomeric and polymeric 1,6-hexamethylene diisocyanate (HDI) during spray painting on 47 automotive spray painters from North Carolina and Washington State. We report here the use of linear mixed modeling to identify the primary determinants of the measured BZCs. Both one-stage (N = 98 paint tasks) and two-stage (N = 198 paint tasks) filter sampling was used to measure concentrations of HDI, uretidone, biuret, and isocyanurate. The geometric mean (GM) level of isocyanurate (1410 μg m−3) was higher than all other analytes (i.e. GM biuret, and isocyanurate, respectively. Mixed models developed in this study characterize the processes governing individual polyisocyanate BZCs. In addition, the mixed models identify ways to reduce polyisocyanate BZCs and, hence, protect painters from potential adverse health effects. PMID:19622637

  10. Rapid detection of economic adulterants in fresh milk by liquid chromatography-tandem mass spectrometry.

    Science.gov (United States)

    Abernethy, Grant; Higgs, Kerianne

    2013-05-03

    A method to aid in the detection of the economically driven adulteration of fresh milk with a range of small, nitrogen containing compounds, including melamine, ammeline, ammelide, cyanuric acid, allantoin, thiourea, urea, biuret, triuret, semicarbazide, aminotriazine, 3- and 4-aminotriazole, cyanamide, dicyandiamide, guanidine, choline, hydroxyproline, nitrate, and a range of amino acids, has been developed. (15)N2-Urea is used as an internal standard. The adulteration of milk with exogenous urea has previously been difficult to detect because of the variation in the naturally occurring levels of urea in milk. However, by monitoring the contaminants biuret and triuret, which comprise up to 1% of synthetic urea, the adulteration of milk with urea-based fertilizer can be detected. We estimate that to be economically viable, adulteration of the order of 90-4000ppm of the above adulterants would need to be added to fresh milk. For most of the compounds, an arbitrary detection threshold of 2ppm is therefore more than sufficient. For biuret, a lower detection threshold, better than 0.5ppm, is desirable and the sensitivity for biuret and triuret can be improved by the post-column addition of lithium to create lithium adducts under electrospray ionisation. Sample handling involves a two-step solvent precipitation method that is deployed in a 96-well plate format, and the hydrophilic interaction liquid chromatography uses a rapid gradient (1.2min). Three separate injections, to detect the positively charged compounds, the negatively charged compounds and amino acids and finally the lithium adducts, are used. This rapid and qualitative survey method may be deployed as a second tier screening method to quickly reduce sample numbers indicated as irregular by an FTIR based screening system, and to direct analysis to appropriate quantification methods. Copyright © 2013 Elsevier B.V. All rights reserved.

  11. . اﻟﺤﻤﺎم اﻷهﻠﻲ ؛ اﻟﻔﺘﺮة اﻟﻀﻮﺋﻴﺔ ؛ اﻟﺘﻴﺮوآﺴﻴﻦ ؛ ﺗﻜﺎﺛﺮ : ا

    African Journals Online (AJOL)

    Prélèvement sanguin par ponction de la veine brachiale. Pesée. Prélèvement sanguin par ponction de la veine brachiale. -Taille des gonades. -Poids des gonades ..... glucose in blood, using glucose oxydase with an alternative oxygen acceptor. Cli. Biochimy., Vol. 6, 1969, p.24. [23] N. Biuret, R.J. Henry, D.C. Cannon,.

  12. THE INFLUENCE OF DTFFEft.ENT PI-ANES OF NUTRITION ...

    African Journals Online (AJOL)

    All the heifers in both treatments had free access to a lick consisting af 259o yellow maize nteal, 25% biuret, 259o dicalcium phosphate, l5e, salt and lA9o sunflower oilcake meal during the winter feeding period. From October 7 the two groups were pooled and had free access to veld and a lick consisting of 50eo dicalcium.

  13. Uji Banding Pengukuran Protein Total Serum Antara Metoda Tetes Layang, Refraktometer Dan Spektrofotometer

    OpenAIRE

    Suryanto, Suryanto; Banundari, RH

    2001-01-01

    Examination of serum total protein is often needed to assess the presence :' hypoproteinemia or hyperproteinemia in various cases, and the method of Examination commonly used is photometric (Biuret) or automatisation. In View of not all regions can make this instrument available, the method of Examination using Refractometer device and Flying-drops method can become alternatives for measuring serum total protein in remote region or in many Tablic health centres and small hospitals.This reseac...

  14. Production of granules of urea, urea-ammonium sulphate and urea-potassium chloride enriched with 15N

    International Nuclear Information System (INIS)

    Bendassolli, J.A.

    1991-01-01

    Using a pearling tower it was possible to produce granulated urea, and granulated mixtures of ammonium sulphate and urea, potassium chloride and urea, Labelled in 15 N. Granulated urea with 1, 2, 3 and 4 mm of diameter was obtained using a system with a heating controller. A low concentration of biuret was observed in the granules produced ( 15 N-Labelled ( 15 NH 4' 15 NH 2 ) with variable proportion of ammonium sulphate and urea. (author)

  15. The growth kinetics of urea monocrystals from 1-propanol-aqueous solutions

    Science.gov (United States)

    Olech, A. Z.; Hodorowicz, S. A.

    1990-05-01

    The growth kinetics of urea monocrystals from 1-propanol-aqueous solutions were studied for solvents of composition 0-25 mol% of n-propanol. The measurements were carried out at 298 K for [001] and [110] directions. The crystallization rate was found to be a linear function of supercooling. The influence of biuret and ammonia on the growth rate of urea crystals is defined quantitatively and the relation between solubility and the crystallization rate is discussed.

  16. Use of refractometry for determination of psittacine plasma protein concentration.

    Science.gov (United States)

    Cray, Carolyn; Rodriguez, Marilyn; Arheart, Kristopher L

    2008-12-01

    Previous studies have demonstrated both poor and good correlation of total protein concentrations in various avian species using refractometry and biuret methodologies. The purpose of the current study was to compare these 2 techniques of total protein determination using plasma samples from several psittacine species and to determine the effect of cholesterol and other solutes on refractometry results. Total protein concentration in heparinized plasma samples without visible lipemia was analyzed by refractometry and an automated biuret method on a dry reagent analyzer (Ortho 250). Cholesterol, glucose, and uric acid concentrations were measured using the same analyzer. Results were compared using Deming regression analysis, Bland-Altman bias plots, and Spearman's rank correlation. Correlation coefficients (r) for total protein results by refractometry and biuret methods were 0.49 in African grey parrots (n=28), 0.77 in Amazon parrots (20), 0.57 in cockatiels (20), 0.73 in cockatoos (36), 0.86 in conures (20), and 0.93 in macaws (38) (Prefractometry in Amazon parrots, conures, and macaws (n=25 each, PRefractometry can be used to accurately measure total protein concentration in nonlipemic plasma samples from some psittacine species. Method and species-specific reference intervals should be used in the interpretation of total protein values.

  17. Ring cleavage and degradative pathway of cyanuric acid in bacteria.

    Science.gov (United States)

    Cook, A M; Beilstein, P; Grossenbacher, H; Hütter, R

    1985-01-01

    The degradative pathway of cyanuric acid [1,3,5-triazine-2,4,6(1H,3H,5H)-trione] was examined in Pseudomonas sp. strain D. The bacterium grew with cyanuric acid, biuret, urea or NH4+ as sole source of nitrogen, and each substrate was entirely metabolized concomitantly with growth. Enzymes from strain D were separated by chromatography on DEAE-cellulose and three reactions were examined. Cyanuric acid (1 mol) was converted stoichiometrically into 1.0 mol of CO2 and 1.1 mol of biuret, which was conclusively identified. Biuret (1 mol) was converted stoichiometrically into 1.1 mol of NH4+, about 1 mol of CO2 and 1.0 mol of urea, which was conclusively identified. Urea (1 mol) was converted into 1.9 mol of NH4+ and 1.0 mol of CO2. The reactions proceeded under aerobic or anoxic conditions and were presumed to be hydrolytic. Data indicate that the same pathway occurred in another pseudomonad and a strain of Klebsiella pneumoniae. PMID:3904735

  18. Gene Sequence and Properties of an s-Triazine Ring-Cleavage Enzyme from Pseudomonas sp. Strain NRRLB-12227

    Science.gov (United States)

    Karns, Jeffrey S.

    1999-01-01

    Pesticides based on the s-triazine ring structure are widely used in cultivation of food crops. Cleavage of the s-triazine ring is an important step in the mineralization of s-triazine compounds and hence in their complete removal from the environment. Cyanuric acid amidohydrolase cleaves cyanuric acid (2,4,6-trihydroxy-s-triazine), which yields carbon dioxide and biuret; the biuret is subject to further metabolism, which yields CO2 and ammonia. The trzD gene encoding cyanuric acid amidohydrolase was cloned into pMMB277 from Pseudomonas sp. strain NRRLB-12227, a strain that is capable of utilizing s-triazines as nitrogen sources. Hydrolysis of cyanuric acid was detected in crude extracts of Escherichia coli containing the cloned gene by monitoring the disappearance of cyanuric acid and the appearance of biuret by high-performance liquid chromatography (HPLC). DEAE and hydrophobic interaction HPLC were used to purify cyanuric acid amidohydrolase to homogeneity, and a spectrophotometric assay for the purified enzyme was developed. The purified enzyme had an apparent Km of 0.05 mM for cyanuric acid at pH 8.0. The enzyme did not cleave any other s-triazine or hydroxypyrimidine compound, although barbituric acid (2,4,6-trihydroxypyrimidine) was found to be a strong competitive inhibitor. Neither the nucleotide sequence of trzD nor the amino acid sequence of the gene product exhibited a significant level of similarity to any known gene or protein. PMID:10427042

  19. Interference of Cerebrospinal Fluid Total Protein Measurement by Povidone-Iodine Contamination

    Science.gov (United States)

    Gounden, Verena; Sacks, David B; Zhao, Zhen

    2014-01-01

    Background A falsely high cerebrospinal fluid (CSF) total protein (TP) result measured by pyrogallol red (PGR) method was suspected to be caused by preparation of the collection site with povidone-iodine (PVP-iodine) solution. Methods CSF TP was evaluated for interference in samples with different final concentrations of PVP-iodine (up to 0.25% PVP and 0.025% iodine) or iodine alone (up to 0.025% iodine) using three methods: PGR, modified biuret and benzethonium chloride (BZTC). Interference exceeding ±20% of the baseline value is considered clinically significant according the criterion defined by College of American Pathologists. Results There was a positive interference with the PGR method and a negative inference for the BZTC method in CSF samples spiked with PVP-iodine. The PVP-iodine (up to 0.25% PVP and 0.025% iodine) did not cause a clinically significant interference with the modified biuret method. PVP alone without iodine caused a positive interference with the PGR method but did not interfere with the modified biuret or the BZTC method. When the samples were spiked with iodine alone, none of the three methods was affected (change < 20%) by iodine concentration up to 0.025%. Conclusions Contamination of CSF specimens with PVP-iodine can lead to interference with CSF TP measurements using PGR or BZTC methods. PMID:25446880

  20. Mixtures of Olive Pomace with Different Nitrogen Sources for the Control of Meloidogyne spp. on Tomato

    Science.gov (United States)

    Rodríguez-Kábana, R.; Estaún, V.; Pinochet, J.; Marfá, O.

    1995-01-01

    The efficacy of mixtures of dry olive (Olea europea) pomace with biuret, guanidine, and melamine for control of root-knot nematodes (Meloidogyne spp.) on tomato (Lycopersicon esculentum) was studied in greenhouse experiments. Olive pomace (OP) applied pre-plant at 10 g/kg soil was phytotoxic. Mixtures of OP (10 g/kg soil) with biuret or guanidine at 200-300 mg/kg soil reduced or eliminated the phytotoxic effect, controlled root-knot nematodes, and increased soil esterase activity indicative of microbial activity. The addition of biuret or guanidine without OP to soil at rates <300 mg/kg soil did not control root-knot nematodes. Melamine applied at 100-400 mg/kg soil was phytotoxic as were mixtures of melamine with OP. Treatment of OP with anhydrous ammonia increased N content of the material. In another greenhouse experiment, NH₃-treated OP added to soil was not phytotoxic to tomato, suppressed root-knot nematodes, and increased soil esterase activity. Greenhouse and microplot experiments with OP plus chicken litter demonstrated the efficacy of these combination amendments to control root-knot nematodes and increase tomato yields in Meloidogyne-infested soil. PMID:19277325

  1. Ultrasound-based protein determination in maize seeds.

    Science.gov (United States)

    Drochioiu, Gabi; Ciobanu, Catalina Ionica; Bancila, Sabina; Ion, Laura; Petre, Brindusa Alina; Andries, Claudia; Gradinaru, Robert Vasile; Murariu, Manuela

    2016-03-01

    The need for a simple and accurate method for protein estimation in alcoholic extracts led to the reexamination of the optimum conditions of a colorimetric assay based on the biuret reaction. Sonication time and the other experimental parameters were optimized after kinetics study on the extraction of either zein or total proteins. Zein extraction and purity were investigated by (1)H and (13)C NMR spectroscopy, SDS-PAGE electrophoresis, and UV-visible spectrophotometry (UV-vis). A zein assay was proposed, which involves the reaction of copper ions in copper phosphate powder with zein extracted in ethanolic solutions under strong alkaline environment. Furthermore, we extended this procedure to determine total proteins in maize samples simultaneously with their ultrasonic-assisted (US) extraction with an alkaline-alcoholic solution. Proteins in both types of extracts were well characterized by UV-vis spectroscopy. However, the 545 nm absorbance of the violet-colored supernatants which is proportional to the protein content was found to be the key parameter of the improved biuret-based protein assay. Comparison of values obtained by this procedure and by Micro-Kjeldahl method was in excellent agreement. A scaled-down procedure agreed well with the standard procedure. Enhanced accuracy and repeatability was found in protein determination in maize using the modified biuret method. The optimization of reagent concentrations and incubation times were studied as well. Copyright © 2015 Elsevier B.V. All rights reserved.

  2. Interference of cerebrospinal fluid total protein measurement by povidone-iodine contamination.

    Science.gov (United States)

    Gounden, Verena; Sacks, David B; Zhao, Zhen

    2015-02-02

    A falsely high cerebrospinal fluid (CSF) total protein (TP) result measured by pyrogallol red (PGR) method was suspected to be caused by preparation of the collection site with povidone-iodine (PVP-iodine) solution. CSF TP was evaluated for interference in samples with different final concentrations of PVP-iodine (up to 0.25% PVP and 0.025% iodine) or iodine alone (up to 0.025% iodine) using three methods: PGR, modified biuret and benzethonium chloride (BZTC). Interference exceeding ±20% of the baseline value is considered clinically significant according to the criterion defined by the College of American Pathologists. There were positive interference with the PGR method and negative inference for the BZTC method in CSF samples spiked with PVP-iodine. The PVP-iodine (up to 0.25% PVP and 0.025% iodine) did not cause a clinically significant interference with the modified biuret method. PVP alone without iodine caused a positive interference with the PGR method but did not interfere with the modified biuret or the BZTC method. When the samples were spiked with iodine alone, none of the three methods was affected (change<20%) by iodine concentration up to 0.025%. Contamination of CSF specimens with PVP-iodine can lead to interference with CSF TP measurements using PGR or BZTC methods. Published by Elsevier B.V.

  3. Mixtures of Olive Pomace with Different Nitrogen Sources for the Control of Meloidogyne spp. on Tomato.

    Science.gov (United States)

    Rodríguez-Kábana, R; Estaún, V; Pinochet, J; Marfá, O

    1995-12-01

    The efficacy of mixtures of dry olive (Olea europea) pomace with biuret, guanidine, and melamine for control of root-knot nematodes (Meloidogyne spp.) on tomato (Lycopersicon esculentum) was studied in greenhouse experiments. Olive pomace (OP) applied pre-plant at 10 g/kg soil was phytotoxic. Mixtures of OP (10 g/kg soil) with biuret or guanidine at 200-300 mg/kg soil reduced or eliminated the phytotoxic effect, controlled root-knot nematodes, and increased soil esterase activity indicative of microbial activity. The addition of biuret or guanidine without OP to soil at rates <300 mg/kg soil did not control root-knot nematodes. Melamine applied at 100-400 mg/kg soil was phytotoxic as were mixtures of melamine with OP. Treatment of OP with anhydrous ammonia increased N content of the material. In another greenhouse experiment, NH-treated OP added to soil was not phytotoxic to tomato, suppressed root-knot nematodes, and increased soil esterase activity. Greenhouse and microplot experiments with OP plus chicken litter demonstrated the efficacy of these combination amendments to control root-knot nematodes and increase tomato yields in Meloidogyne-infested soil.

  4. Coil-type Fano Resonances: a Plasmonic Approach to Magnetic Sub-diffraction Confinement

    KAUST Repository

    Panaro, Simone

    2015-05-10

    Matrices of nanodisk trimers are introduced as plasmonic platforms for the generation of localized magnetic hot-spots. In Fano resonance condition, the optical magnetic fields can be squeezed in sub-wavelength regions, opening promising scenarios for spintronics.

  5. Coiled-coil oligomerization controls localization of the plasma membrane REMORINs.

    Science.gov (United States)

    Martinez, Denis; Legrand, Anthony; Gronnier, Julien; Decossas, Marion; Gouguet, Paul; Lambert, Olivier; Berbon, Mélanie; Verron, Loris; Grélard, Axelle; Germain, Veronique; Loquet, Antoine; Mongrand, Sébastien; Habenstein, Birgit

    2018-02-23

    REMORINs are nanodomain-organized proteins located in the plasma membrane and involved in cellular responses in plants. The dynamic assembly of the membrane nanodomains represents an essential tool of the versatile membrane barriers to control and modulate cellular functions. Nevertheless, the assembly mechanisms and protein organization strategies of nanodomains are poorly understood and many structural aspects are difficult to visualize. Using an ensemble of biophysical approaches, including solid-state nuclear magnetic resonance, cryo-electron microscopy and in vivo confocal imaging, we provide first insights on the role and the structural mechanisms of REMORIN trimerization. Our results suggest that the formation of REMORIN coiled-coil trimers is essential for membrane recruitment and promotes REMORIN assembly in vitro into long filaments by trimer-trimer interactions that might participate in nanoclustering into membrane domains in vivo. Copyright © 2018. Published by Elsevier Inc.

  6. Harnessing Quinone Methides: Total Synthesis of (±)‐Vaticanol A

    DEFF Research Database (Denmark)

    Jepsen, Tue Heesgaard; Thomas, Stephen; Yunqing, lin

    2014-01-01

    Although quinone methides are often postulated as intermediates in the biosynthesis of many polyphenolic natural products, deploying their power in a laboratory setting to achieve similar bond constructions has sometimes proven challenging. Herein, a total synthesis of the resveratrol trimer...

  7. Synthesis and membrane performance characterization of self-emulsified waterborne nitrocellulose dispersion modified with castor oil

    Science.gov (United States)

    Su, Xiuxia; Zhao, Qingxiao; Zhang, Dan; Dong, Wei

    2015-11-01

    Waterborne nitrocellulose dispersion modified with castor oil (CWNC) was designed and successfully synthesized by self emulsification and reaction among isophorone diisocyanate (IPDI) trimer, dimethylol propionic acid (DMPA), nitrocellulose (NC) and castor oil (C.O.). The CWNC was characterized by transmission electron microscope (TEM), Fourier transform infrared (FTIR) spectroscopy and thermal gravimetric analysis (TGA), etc. The particle size of CWNC increased with the increase of mass fraction of castor oil to total reactants, ω (C.O.). The morphology of particles is an approximate core-shell structure indicated by TEM. FTIR confirmed that the reactions (i.e. IPDI trimer and castor oil, IPDI trimer and NC) occurred, the NCO groups of IPDI trimer were consumed totally and the backbone of NC was retained. The water contact angle measurements confirmed that introduced castor oil increased hydrophobicity of the film, thereby increasing the contact angle. TGA revealed that the CWNC film had better thermal resistance.

  8. The Role of the Hendra Virus and Nipah Virus Attachment Glycoproteins in Receptor Binding and Antibody Neutralization

    Science.gov (United States)

    2014-01-31

    glycoproteins, such as HIV gp140 and Rabies glycoprotein, its ability to form trimers but not tetramers makes it ineffective for correct oligomerization of...Oldstone MB. 2000. V and C proteins of measles virus function as virulence factors in vivo. Virology 267:80-9.   216 148. Playford EG, McCall B...2005. Stable trimerization of recombinant rabies virus glycoprotein ectodomain is required for interaction with the p75NTR receptor. J Gen Virol 86

  9. Proteolytic Processing of the Human Immunodeficiency Virus Envelope Glycoprotein Precursor Decreases Conformational Flexibility

    OpenAIRE

    Haim, Hillel; Salas, Ignacio; Sodroski, Joseph

    2013-01-01

    The mature envelope glycoprotein (Env) spike on the surface of human immunodeficiency virus type 1 (HIV-1) virions is derived by proteolytic cleavage of a trimeric gp160 glycoprotein precursor. Remarkably, proteolytic processing of the HIV-1 Env precursor results in changes in Env antigenicity that resemble those associated with glutaraldehyde fixation. Apparently, proteolytic processing of the HIV-1 Env precursor decreases conformational flexibility of the Env trimeric complex, differentiall...

  10. Molecular structure and interactions of nucleic acid components in nanoparticles: ab initio calculations

    International Nuclear Information System (INIS)

    Rubin, Yu.V.; Belous, L.F.

    2012-01-01

    Self-associates of nucleic acid components (stacking trimers and tetramers of the base pairs of nucleic acids) and short fragments of nucleic acids are nanoparticles (linear sizes of these particles are more than 10 A). Modern quantum-mechanical methods and softwares allow one to perform ab initio calculations of the systems consisting of 150-200 atoms with enough large basis sets (for example, 6-31G * ). The aim of this work is to reveal the peculiarities of molecular and electronic structures, as well as the energy features of nanoparticles of nucleic acid components. We had carried out ab initio calculations of the molecular structure and interactions in the stacking dimer, trimer, and tetramer of nucleic base pairs and in the stacking (TpG)(ApC) dimer and (TpGpC) (ApCpG) trimer of nucleotides, which are small DNA fragments. The performed calculations of molecular structures of dimers and trimers of nucleotide pairs showed that the interplanar distance in the structures studied is equal to 3.2 A on average, and the helical angle in a trimer is approximately equal to 30 o : The distance between phosphor atoms in neighboring chains is 13.1 A. For dimers and trimers under study, we calculated the horizontal interaction energies. The analysis of interplanar distances and angles between nucleic bases and their pairs in the calculated short oligomers of nucleic acid base pairs (stacking dimer, trimer, and tetramer) has been carried out. Studies of interactions in the calculated short oligomers showed a considerable role of the cross interaction in the stabilization of the structures. The contribution of cross interactions to the horizontal interactions grows with the length of an oligomer. Nanoparticle components get electric charges in nanoparticles. Longwave low-intensity bands can appear in the electron spectra of nanoparticles.

  11. Molecular dynamics studies of PEGylated α-helical coiled coils and their self-assembled micelles.

    Science.gov (United States)

    Woo, Sun Young; Lee, Hwankyu

    2014-07-29

    We performed coarse-grained (CG) molecular dynamics simulations of trimeric α-helical coiled coils grafted with poly(ethylene glycol) (PEG) of different sizes and conjugate positions and the self-assembled micelle of amphiphilic trimers. The CG model for the trimeric coiled coil is verified by comparing the α-helical structure and interhelical distance with those calculated from all-atom simulations. In CG simulations of PEGylated trimers, the end-to-end distances and radii of gyration of PEGs grafted to the sides of peptides become shorter than those of free PEGs in water, which agrees with experiments. This shorter size of the grafted PEGs is also confirmed by calculating the thickness of the PEG layer, which is less than the size of the mushroom. These indicate the adsorption of PEG chains onto coiled coils since hydrophobic residues in the exterior sites of coiled coils tend to be less exposed to water and thus interact with PEGs, leading to the compact conformation of adsorbed PEGs. Simulations of the self-assembly of amphiphilic trimers show that the randomly distributed trimers self-assemble to micelles. The outer radius and hydrodynamic radius of the micelle, which were calculated respectively from radial densities and diffusion coefficients, are ∼7 nm, in agreement with the experimental value of ∼7.5 nm, while the aggregation number of amphiphilic molecules per micelle is lower than the experimentally proposed value. These simulations predict the experimentally measured size of PEGs grafted to the trimeric coiled coils and their self-assembled amphiphilic micelles and suggest that the aggregation number of the micelle may be lower, which needs to be confirmed by experiments.

  12. Preliminary crystallographic analysis of the major capsid protein P2 of the lipid-containing bacteriophage PM2

    International Nuclear Information System (INIS)

    Abrescia, Nicola G. A.; Kivelä, Hanna M.; Grimes, Jonathan M.; Bamford, Jaana K. H.; Bamford, Dennis H.; Stuart, David I.

    2005-01-01

    The viral capsid protein P2 of bacteriophage PM2 has been crystallized. Preliminary X-ray analysis demonstrates the position and orientation of the two trimers in the asymmetric unit. PM2 (Corticoviridae) is a dsDNA bacteriophage which contains a lipid membrane beneath its icosahedral capsid. In this respect it resembles bacteriophage PRD1 (Tectiviridae), although it is not known whether the similarity extends to the detailed molecular architecture of the virus, for instance the fold of the major coat protein P2. Structural analysis of PM2 has been initiated and virus-derived P2 has been crystallized by sitting-nanodrop vapour diffusion. Crystals of P2 have been obtained in space group P2 1 2 1 2, with two trimers in the asymmetric unit and unit-cell parameters a = 171.1, b = 78.7, c = 130.1 Å. The crystals diffract to 4 Å resolution at the ESRF BM14 beamline (Grenoble, France) and the orientation of the non-crystallographic threefold axes, the spatial relationship between the two trimers and the packing of the trimers within the unit cell have been determined. The trimers form tightly packed layers consistent with the crystal morphology, possibly recapitulating aspects of the arrangement of subunits in the virus

  13. Antibodies to a conformational epitope on gp41 neutralize HIV-1 by destabilizing the Env spike

    Science.gov (United States)

    Lee, Jeong Hyun; Leaman, Daniel P.; Kim, Arthur S.; Torrents de La Peña, Alba; Sliepen, Kwinten; Yasmeen, Anila; Derking, Ronald; Ramos, Alejandra; de Taeye, Steven W.; Ozorowski, Gabriel; Klein, Florian; Burton, Dennis R.; Nussenzweig, Michel C.; Poignard, Pascal; Moore, John P.; Klasse, Per Johan; Sanders, Rogier W.; Zwick, Michael B.; Wilson, Ian A.; Ward, Andrew B.

    2015-09-01

    The recent identification of three broadly neutralizing antibodies (bnAbs) against gp120-gp41 interface epitopes has expanded the targetable surface on the HIV-1 envelope glycoprotein (Env) trimer. By using biochemical, biophysical and computational methods, we map the previously unknown trimer epitopes of two related antibodies, 3BC315 and 3BC176. A cryo-EM reconstruction of a soluble Env trimer bound to 3BC315 Fab at 9.3 Å resolution reveals that the antibody binds between two gp41 protomers, and neutralizes the virus by accelerating trimer decay. In contrast, bnAb 35O22 binding to a partially overlapping quaternary epitope at the gp120-gp41 interface does not induce decay. A conserved gp41-proximal glycan at N88 was also shown to play a role in the binding kinetics of 3BC176 and 3BC315. Finally, our data suggest that the dynamic structure of the Env trimer influences exposure of bnAb epitopes.

  14. Antibody potency relates to the ability to recognize the closed, pre-fusion form of HIV Env

    Science.gov (United States)

    Guttman, Miklos; Cupo, Albert; Julien, Jean-Philippe; Sanders, Rogier W.; Wilson, Ian A.; Moore, John P.; Lee, Kelly K.

    2015-02-01

    HIV’s envelope glycoprotein (Env) is the sole target for neutralizing antibodies. The structures of many broadly neutralizing antibodies (bNAbs) in complex with truncated Env subunits or components have been reported. However, their interaction with the intact Env trimer, and the structural determinants that underlie neutralization resistance in this more native context are less well understood. Here we use hydrogen/deuterium exchange to examine the interactions between a panel of bNAbs and native-like Env trimers (SOSIP.664 trimers). Highly potent bNAbs cause only localized effects at their binding interface, while the binding of less potent antibodies is associated with elaborate changes throughout the trimer. In conjunction with binding kinetics, our results suggest that poorly neutralizing antibodies can only bind when the trimer transiently samples an open state. We propose that the kinetics of such opening motions varies among isolates, with Env from neutralization-sensitive viruses opening more frequently than Env from resistant viruses.

  15. Structures and transport dynamics of a Campylobacter jejuni multidrug efflux pump

    Energy Technology Data Exchange (ETDEWEB)

    Su, Chih-Chia; Yin, Linxiang; Kumar, Nitin; Dai, Lei; Radhakrishnan, Abhijith; Bolla, Jani Reddy; Lei, Hsiang-Ting; Chou, Tsung-Han; Delmar, Jared A.; Rajashankar, Kanagalaghatta R.; Zhang, Qijing; Shin, Yeon-Kyun; Yu, Edward W. (Cornell); (Iowa State)

    2017-08-01

    Resistance-nodulation-cell division efflux pumps are integral membrane proteins that catalyze the export of substrates across cell membranes. Within the hydrophobe-amphiphile efflux subfamily, these resistance-nodulation-cell division proteins largely form trimeric efflux pumps. The drug efflux process has been proposed to entail a synchronized motion between subunits of the trimer to advance the transport cycle, leading to the extrusion of drug molecules. Here we use X-ray crystallography and single-molecule fluorescence resonance energy transfer imaging to elucidate the structures and functional dynamics of the Campylobacter jejuni CmeB multidrug efflux pump. We find that the CmeB trimer displays a very unique conformation. A direct observation of transport dynamics in individual CmeB trimers embedded in membrane vesicles indicates that each CmeB subunit undergoes conformational transitions uncoordinated and independent of each other. On the basis of our findings and analyses, we propose a model for transport mechanism where CmeB protomers function independently within the trimer.

  16. Macrocyclic ligands for uranium complexation

    International Nuclear Information System (INIS)

    Potts, K.T.

    1990-04-01

    Macrocycles with cavity diameters in the region 4.5-5.2 Angstrom, based on computer modeling studies, have been prepared by high dilution cyclocondensation of 2-aminopyridyl substituted biurets and thiobiurets with appropriately substituted, reactive intermediates such as chlorocarbonyl isocyanate and malonyl dichloride. Detailed high field 1 H NMR studies have been used to identify all protons in these macrocycles for structural verification and in confromation studies, and by-products obtained in their synthesis as well as in complexation studies have been characterized and identified using these NMR techniques, as well as single crystal X-ray studies. 5 refs

  17. pelo método do Biureto

    Directory of Open Access Journals (Sweden)

    João Baptista Severo

    2007-01-01

    Full Text Available In this work, the aim was to minimize the interference effect provoked by pH and the bovine serum albumin (BSA and the phosphate buffer concentration on the total protein concentration determined by the biuret method, using a statistical model, established by response surface methodology (RSM. Results showed that the factors effects on protein concentration analysis were very significant and, in order to reduce these effects, it is necessary to use 0.01 M phosphate buffer at pH 7 and the protein concentration must be inferior, at 3 g/L.

  18. Thermal decomposition of dinitratobis(carbamido) uranyl

    International Nuclear Information System (INIS)

    Kobets, L.V.; Kostyukov, N.N.; Umrejko, D.S.

    1987-01-01

    Thermal stability of dinitratobis (carbamido) uranyl was investigated by the methods of DTA, TG and isothermal heatings at different temperatures. It was revealed that urea as a whole was not removed from the complex. Urea loses ammonia simultaneously with substance melting (498K); biuret and cyanuric acid form at that. Ammonia removal to gaseous phase decelerates in 573-593 K range due to its binding with formation of amination products. Decomposition of nitrate groups begins at temperatures above 570 K. Heating products were studied by the methods of vibrational spectroscopy, chemical and X-ray phase analyses. Ideas about mechanism of decomposition were considered

  19. Investigation of the behavior of urea during the pelleting of straw in a laboratory hand press

    International Nuclear Information System (INIS)

    Bergner, H.; Mueller, J.; Kijora, C.

    1983-01-01

    Pellets containing 14 C- and 15 N-labelled urea (3%) were made in a self-developed laboratory hand press at temperatures of 100 0 C, 150 0 C, and 200 0 C. Transformation and decomposition processes on a large scale only took place at a temperature of 200 0 C. Partly, urea was decomposed (formation of CO 2 and NH 3 ) and, partly, polymerization products were formed (biuret and cyanic acid). Even at a temperature of 200 0 C there were 85.5% nondecomposed urea in the dissoluble filtrate. All the products formed, including urea, very probably undergo condensation reactions with straw. (author)

  20. Reduction of nitrogen- and carbon-based pollutants

    Energy Technology Data Exchange (ETDEWEB)

    Bowers, W.E.

    1990-05-22

    This patent describes a process for educing the concentration of nitrogen oxides in an oxygen-rich effluent from the combustion of a carbonaceous fuel. It comprises: injecting a solution comprising at least one additive compound selected from the group consisting of guanidine, guanidine carbonate, biguanide, guanylurea sulfate, melamine, dicyandiamide, biuret, 1,1{prime}-azobisformamide, methylol urea, methylol urea-urea condensation product, dimethylol urea, methyl urea, and dimethyl urea, at a concentration and a temperature effective to achieve reduction in nitrogen oxide levels in the effluent.

  1. Ultraviolet-gas phase and -photocatalytic synthesis from CO and NH3. [photolysis products

    Science.gov (United States)

    Hubbard, J. S.; Voecks, G. E.; Hobby, G. L.; Ferris, J. P.; Williams, E. A.; Nicodem, D. E.

    1975-01-01

    Ammonium cyanate is identified as the major product of the photolysis of gaseous NH3-CO mixtures at 206.2 or 184.9 nm. Lesser amounts of urea, biurea, biuret semicarbazide, formamide and cyanide are observed. A series of 18 reactions underlying the formation of photolysis products is presented and discussed. Photocatalytic syntheses of C-14-urea, -formamide, and -formaldehyde are carried out through irradiation of (C-14)O and NH3 in the presence of Vycor, silica gel, or volcanic ash shale surfaces. The possible contributions of the relevant reactions to the abiotic synthesis of organic nitrogen compounds on Mars, the primitive earth, and in interstellar space are examined.

  2. Spectral-luminescent properties of the complex of organic amides with europium nitrate

    International Nuclear Information System (INIS)

    Makhmudova, Kh.I.; Akhmatova, M.R.; Davronov, M.D.; Resnyanskij, V.F.; Sulajmankulov, K.S.; Kaldybaev, K.A.; Umarov, K.U.

    2008-01-01

    In this paper the results of studying the excitation and fluorescence spectra of co-ordinating compounds of carbamide and biuret with europium nitrate, and influence of organic components on intensity of spectra of initial europium nitrate are presented. It was found that the photoluminescence of europium ions in carbamide compounds possesses a considerable brightness. This is indicative of probability of excited europium ion deactivation. Most bright emission is observed in compounds of europium nitrate with four carbamide molecules and two water ones. (authors)

  3. Formation of biologically relevant compounds of interest in chemical evolution from the radiolysis of succinonitrile solutions

    International Nuclear Information System (INIS)

    Albarran, G.; Juarez, C.; Negron-Mendoza, A.

    1991-01-01

    Low molecular weight compounds such as H 2 , CO 2 , NH 3 were identified among the radiolytic products. Irradiated samples exhibit positive biuret test. IR spectra of the dry residue confirm the presence of amide groups. These results suggest the presence of peptidic type material, which increased with the radiation dose. Other compounds identified were several di and tricarboxylic acids. The initial yield of formation of a variety of products was calculated from the concentration vs dose plots. Some of the radiolytic compounds are of biological importance and their formation is significant to chemical evolution studies. (author) 7 refs

  4. Kemin bakom framtidens avgasrening : En studie av ureasönderfall under kvävgasatmosfär

    OpenAIRE

    Le, Tan

    2011-01-01

    The purpose of this work was to provide a better understanding of urea’s decomposition and byproduct formation in an SCR system on heavy trucks. In my experimental setup with TGA-DSC-FTIR (a combination of two thermal analysis methods and a method for gas phase detection), an FTIR method for urea in the gas phase was developed for the qualitative and quantitative determination of urea and its decomposition products. Chemicals such as urea, biuret, cyanuric acid and melamine of p.a. quality we...

  5. GLUCOSE AND TOTAL PROTEIN LEVEL IN LABORATORY RATS UNDER CONDITIONS OF SHORT-TERM FASTING

    Directory of Open Access Journals (Sweden)

    Damir Suljević

    2013-09-01

    Full Text Available Glucose level (UV enzymatic method and total protein level (Biuret method were measured in the blood samples of the rats exposed to short-term starvation. We found a statistically significant increase in the glucose level in experimental animals during starvation, which is also evident in males and females in the experimental group (p <0.05, while decrease in the total protein level was not statistically significant. During starvation, more significant weight loss was observed in females compared to males.Key words: glucose, total protein, serum, Rattus

  6. Structure of Rotavirus Outer-Layer Protein VP7 Bound with a Neutralizing Fab

    Energy Technology Data Exchange (ETDEWEB)

    Aoki, Scott T.; Settembre, Ethan C.; Trask, Shane D.; Greenberg, Harry B.; Harrison, Stephen C.; Dormitzer, Philip R.; (Stanford-MED); (CH-Boston)

    2009-06-17

    Rotavirus outer-layer protein VP7 is a principal target of protective antibodies. Removal of free calcium ions (Ca{sup 2+}) dissociates VP7 trimers into monomers, releasing VP7 from the virion, and initiates penetration-inducing conformational changes in the other outer-layer protein, VP4. We report the crystal structure at 3.4 angstrom resolution of VP7 bound with the Fab fragment of a neutralizing monoclonal antibody. The Fab binds across the outer surface of the intersubunit contact, which contains two Ca{sup 2+} sites. Mutations that escape neutralization by other antibodies suggest that the same region bears the epitopes of most neutralizing antibodies. The monovalent Fab is sufficient to neutralize infectivity. We propose that neutralizing antibodies against VP7 act by stabilizing the trimer, thereby inhibiting the uncoating trigger for VP4 rearrangement. A disulfide-linked trimer is a potential subunit immunogen.

  7. Efimov states and bound state properties in selected nuclear and molecular three-body systems

    International Nuclear Information System (INIS)

    Huber, H.S.

    1978-01-01

    The search is made among selected three-body systems for possible Efimov state behavior. In order to carry out this analysis of phenomenological potentials a new mathematical approach, the FCM (Faddeev-coordinate-momentum) technique, is developed. The analysis then proceeds through the framework of the Faddeev equations by employing the UPE (unitary pole expansion) to reduce these equations to numerically feasible form. The systems chosen for analysis are the 4 He trimer and the three-α model of 12 C. Efimov states are not found in 12 C, thus answering speculation among nuclear theorists. The 4 He trimer, on the other hand, manifests Efimov states for each potential considered and the characteristics of these states are extensively analyzed. Since Efimov states are predicted by all of the phenomenological potentials considered, these states would seem to be a realistically fundamental property of the 4 He trimer system

  8. Pair condensation and bound states in fermionic systems

    International Nuclear Information System (INIS)

    Sedrakian, Armen; Clark, John W.

    2006-01-01

    We study the finite temperature-density phase diagram of an attractive fermionic system that supports two-body (dimer) and three-body (trimer) bound states in free space. Using interactions characteristic for nuclear systems, we obtain the critical temperature T c2 for the superfluid phase transition and the limiting temperature T c3 for the extinction of trimers. The phase diagram features a Cooper-pair condensate in the high-density, low-temperature domain which, with decreasing density, crosses over to a Bose condensate of strongly bound dimers. The high-temperature, low-density domain is populated by trimers whose binding energy decreases toward the density-temperature domain occupied by the superfluid and vanishes at a critical temperature T c3 >T c2

  9. Hydrolyzable Tannins of Tamaricaceous Plants. 7.1 Structures and Cytotoxic Properties of Oligomeric Ellagitannins from Leaves of Tamarix nilotica and Cultured Tissues of Tamarix tetrandra.

    Science.gov (United States)

    Orabi, Mohamed A A; Taniguchi, Shoko; Sakagami, Hiroshi; Yoshimura, Morio; Amakura, Yoshiaki; Hatano, Tsutomu

    2016-04-22

    Partially unacylated new oligomeric hydrolyzable tannins, nilotinin T2 (1, trimer) and nilotinin Q1 (2, tetramer), together with four known trimers, nilotinin T1 (3) and hirtellins T1-T3 (4-6), and a dimer, tamarixinin B (7), were isolated from the aqueous acetone extracts of leaves of Tamarix nilotica. Among them, the new trimer 1 and the known trimers 4 and 6, in addition to the partially unacylated new trimer nilotinin T3 (8), the known dimers nilotinin D3 (9) and tamarixinin C (10), and the monomer tellimagrandin I (11), were isolated from the cultured shoots of Tamarix tetrandra. The structures of the new hydrolyzable tannins were established by chromatographic analyses and extensive 1D and 2D NMR, HRESI-TOFMS, and ECD spectroscopic experiments. Among the new oligomeric tannins, the particular unacylated position of a glucose core is attributed to a possible biosynthetic route. Isolation of the same oligomeric tannins from cultured shoots of T. tetrandra emphasizes the unique biogenetic ability of the obtained cultures on production of the structurally and biologically characteristic tamaricaceous tannins commonly produced by the intact Tamarix plants. Additionally, tannins obtained in the present study together with gemin D (12) and 1,3-di-O-galloyl-4,6-O-(aS)-hexahydroxydiphenoyl-β-d-glucose (13), from our previous investigation of the leaves of T. nilotica, exhibited variable tumor-specific cytotoxic effects. The ellagitannin trimers 4, 6, and 8 and the dimer 9 exerted predominant tumor-selective cytotoxic effects with high specificity toward human promyelocytic leukemia cells.

  10. Picosecond excitation energy transfer of allophycocyanin studied in solution and in crystals.

    Science.gov (United States)

    Ranjbar Choubeh, Reza; Sonani, Ravi R; Madamwar, Datta; Struik, Paul C; Bader, Arjen N; Robert, Bruno; van Amerongen, Herbert

    2018-03-01

    Cyanobacteria perform photosynthesis with the use of large light-harvesting antennae called phycobilisomes (PBSs). These hemispherical PBSs contain hundreds of open-chain tetrapyrrole chromophores bound to different peptides, providing an arrangement in which excitation energy is funnelled towards the PBS core from where it can be transferred to photosystem I and/or photosystem II. In the PBS core, many allophycocyanin (APC) trimers are present, red-light-absorbing phycobiliproteins that covalently bind phycocyanobilin (PCB) chromophores. APC trimers were amongst the first light-harvesting complexes to be crystallized. APC trimers have two spectrally different PCBs per monomer, a high- and a low-energy pigment. The crystal structure of the APC trimer reveals the close distance (~21 Å) between those two chromophores (the distance within one monomer is ~51 Å) and this explains the ultrafast (~1 ps) excitation energy transfer (EET) between them. Both chromophores adopt a somewhat different structure, which is held responsible for their spectral difference. Here we used spectrally resolved picosecond fluorescence to study EET in these APC trimers both in crystallized and in solubilized form. We found that not all closely spaced pigment couples consist of a low- and a high-energy pigment. In ~10% of the cases, a couple consists of two high-energy pigments. EET to a low-energy pigment, which can spectrally be resolved, occurs on a time scale of tens of picoseconds. This transfer turns out to be three times faster in the crystal than in the solution. The spectral characteristics and the time scale of this transfer component are similar to what have been observed in the whole cells of Synechocystis sp. PCC 6803, for which it was ascribed to EET from C-phycocyanin to APC. The present results thus demonstrate that part of this transfer should probably also be ascribed to EET within APC trimers.

  11. Monomeric nucleoprotein of influenza A virus.

    Directory of Open Access Journals (Sweden)

    Sylvie Chenavas

    2013-03-01

    Full Text Available Isolated influenza A virus nucleoprotein exists in an equilibrium between monomers and trimers. Samples containing only monomers or only trimers can be stabilized by respectively low and high salt. The trimers bind RNA with high affinity but remain trimmers, whereas the monomers polymerise onto RNA forming nucleoprotein-RNA complexes. When wild type (wt nucleoprotein is crystallized, it forms trimers, whether one starts with monomers or trimers. We therefore crystallized the obligate monomeric R416A mutant nucleoprotein and observed how the domain exchange loop that leads over to a neighbouring protomer in the trimer structure interacts with equivalent sites on the mutant monomer surface, avoiding polymerisation. The C-terminus of the monomer is bound to the side of the RNA binding surface, lowering its positive charge. Biophysical characterization of the mutant and wild type monomeric proteins gives the same results, suggesting that the exchange domain is folded in the same way for the wild type protein. In a search for how monomeric wt nucleoprotein may be stabilized in the infected cell we determined the phosphorylation sites on nucleoprotein isolated from virus particles. We found that serine 165 was phosphorylated and conserved in all influenza A and B viruses. The S165D mutant that mimics phosphorylation is monomeric and displays a lowered affinity for RNA compared with wt monomeric NP. This suggests that phosphorylation may regulate the polymerisation state and RNA binding of nucleoprotein in the infected cell. The monomer structure could be used for finding new anti influenza drugs because compounds that stabilize the monomer may slow down viral infection.

  12. Structure of the first PDZ domain of human PSD-93

    DEFF Research Database (Denmark)

    Fiorentini, Monica; Nielsen, Ann Kallehauge; Kristensen, Ole

    2009-01-01

    The crystal structure of the PDZ1 domain of human PSD-93 has been determined to 2.0 A resolution. The PDZ1 domain forms a crystallographic trimer that is also predicted to be stable in solution. The main contributions to the stabilization of the trimer seem to arise from interactions involving th...... as well as of the closely related human PSD-95 PDZ1 shows that they are very similar in terms of amino-acid composition. However, the cleft is significantly narrower in PSD-95. This could be part of the basis of peptide selectivity between PSD-93 PDZ1 and PSD-95 PDZ1....

  13. Two zeolite-type frameworks in one metal-organic framework with Zn24 @Zn104 cube-in-sodalite architecture.

    Science.gov (United States)

    Bu, Fei; Lin, Qipu; Zhai, Quanguo; Wang, Le; Wu, Tao; Zheng, Shou-Tian; Bu, Xianhui; Feng, Pingyun

    2012-08-20

    Two in one: A metal-organic framework obtained from three different inorganic building blocks (tetrameric Zn(4) O, trimeric Zn(3) OH, and monomeric Zn) posseses a nested cage-in-cage and framework-in-framework architecture. 24 Zn(4) O tetramers and eight Zn monomers form a sodalite cage into which a cubic cage made from eight Zn(3) (OH) trimers is nestled. Eight monomeric Zn(2+) centers interconnect these two cages. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  14. Control of the active site structure of giant bilayer hemoglobin from the Annelid Eisenia foetida using hierarchic assemblies

    Science.gov (United States)

    Girasole, Marco; Arcovito, Alessandro; Marconi, Augusta; Davoli, Camilla; Congiu-Castellano, Agostina; Bellelli, Andrea; Amiconi, Gino

    2005-12-01

    The active site structure of the oxygenated derivative of the main subassemblies (whole protein, dodecamers, and trimers) of the giant haemoglobin from Eisenia foetida has been characterized by x-ray absorption near edge structure spectroscopy. The data revealed a remarkable effect of the hierarchic assemblies on the active site of the subunit. Specifically, the whole protein has the same site structure of the dodecamer, while a sharp conformational transition occurs when the dodecamer is disassembled into trimers (and monomers) revealing that constraints due to the protein matrix determine the active site geometry and, consequently, the protein function in these large complexes.

  15. Exploring the "fold-in" strategy toward the construction of a highly-strained triazasumanene skeleton.

    Science.gov (United States)

    Chen, Fengkun; Tanaka, Takayuki; Osuka, Atsuhiro

    2017-02-28

    The synthesis of tribenzotriazasumanene was attempted on the basis of the "fold-in" synthesis of cyclic pyrrole trimer 5 and its analogues under oxidative and reductive conditions. While unexpectedly triply-fused product 6 was obtained in the oxidation of 5 with FeCl 3 and AgOTf, the reductive coupling of hexabromo-tri-N-methylpyrrole trimer 9 furnished partially fused product 10. These results indicate the potential of the "fold-in" strategy that gave complementary outcomes depending on the reaction conditions.

  16. Investigation into chromophore excited-state coupling in allophycocyanin

    Science.gov (United States)

    Zheng, Xiguang; Zhao, Fuli; Wang, He Z.; Gao, Zhaolan; Yu, Zhenxin; Zhu, Jinchang; Xia, Andong; Jiang, Lijin

    1994-08-01

    Both theoretical and experimental studies are presented on chromophore excited-state coupling in linker-free allophycocyanin (APC), one of the antenna phycobiliproteins in algal photosynthesis. A three-site-coupling model has been introduced to describe the exciton interaction mechanism amoung the excited (beta) chromophore in APC, and the exciton energy splitting is estimated. Picosecond polarized fluorescence experiments both on monomeric and trimeric APC isolated from alga Spirulina platensis have been performed. The experimental results show that APC monomer and trimer exhibit remarkedly different spectropic characteristics, and satisfy the suggestion of strong excited- state coupling among chromophores in APC.

  17. Investigation of γ-ray induced polymer formation in the carboranes

    International Nuclear Information System (INIS)

    Klingen, T.J.; Mathur, M.A.

    1977-01-01

    The γ-ray radiolysis of pure 1-phenyl-o-carborane in the solid state at 31 0 C resulted in the formation of hydrogen, methane, ethylene and benzene gases, as well as solid products which were determined to be adduct dimer, trimer and tetramer of 1-phenyl-o-carborane and the cyclic trimer of o-carborane. The formation of the adduct oligomeric products has been shown to be a chain process which predominates at low total doses. Whereas, the reaction path leading to decomposition of 1-phenyl-o-carborane has been shown to be a non-chain process that predominates at high total doses. (author)

  18. Escherichia coli Purine Nucleoside Phosphorylase II, the Product of the xapA Gene

    DEFF Research Database (Denmark)

    Dandanell, Gert; Szczepanowski, R.H.; Kierdaszuk, B.

    2005-01-01

    , but has been reported to migrate as a hexamer and to accept xanthosine with comparable efficiency to guanosine and inosine, the usual physiological substrates for trimeric PNPs. Here, we present a detailed biochemical characterization and the crystal structure of E. coli PNP-II. In three different crystal...... the monoanionic form of xanthine. A single amino acid exchange, tyrosine 191 to leucine, is sufficient to convert E. coli PNP-II into an enzyme with the specificity of conventional trimeric PNPs, but the reciprocal mutation in human PNP, valine 195 to tyrosine, does not elicit xanthosine phosphorylase activity...

  19. The radiolysis of aqueous ammonium cyanide - Compounds of interest to chemical evolution studies

    International Nuclear Information System (INIS)

    Draganic, Z.D.; Jovanovic, S.

    1980-01-01

    Oxygen-free aqueous solutions of NH4CN (0.1 M, pH 9) were exposed to gamma rays from a Co-60 source, the mixture of nonvolatile products was fractionated, and the fractions were analyzed. The procedures were chosen to make effective investigations of radiolytic products, and to minimize the contributions of chemical changes which are known to occur in aqueous solution in the absence of ionizing radiation. It has been found that the main constituents are: urea, 25.9% an oligomer, very likely oligoimine (18.4%) and several fractions (about 50%) which release amino acids on hydrolysis. These fractions differ considerably, as shown by amino acid assay, enzymatic digestion, IR spectra, and biuret reaction. All these tests were found to be positive for two fractions in two further fractions the enzymatic cleavage was absent, but other tests were positive. Negative enzymatic and biuret tests, and no bands characteristic of amide or peptide, were found for a fraction whose hydrolysate consisted of 55% glycine. Although most of the isolated materials were found to be composite, the results of the analyses were sufficient for getting a reliable over-all picture of the chemical action of the ionizing radiation. The role of free radicals in reactions leading to the formations of radiolytic products was considered

  20. Radiation chemistry of aqueous solutions of acetonitrile and propionitrile in the megarad dose range

    International Nuclear Information System (INIS)

    Draganic, Z.D.; Draganic, I.G.; Shushtarian, M.J.

    1976-01-01

    A study was made of the radiolytic behavior of dilute, neutral, oxygen-free aqueous solutions of CH 3 CN and C 2 H 5 CN. Small-molecular products were identified as RCHO, NH 3 , CO 2 , and H 2 . The decomposition of nitrile is followed by high yields of formation of the nonvolatile nitrogen-containing compounds, G(N). The γ-irradiated solutions exhibit a positive biuret reaction. Several amino acids were identified among radiolytic products, and glycine and alanine were found to be the most abundant for CH 3 CN and C 2 H 5 CH, respectively. Their yields increased after strong acid hydrolysis of the irradiated samples. The free radicals formed by reactions of RCN with H, OH, and e/sub aq/- were found to be important for the phenomena observed. It is suggested that the positive biuret reaction, ir spectra, and the release of amino acids on acid hydrolysis provide some evidence on the formation of peptidic materials and might be of interest for the evaluation of the role that ionizing radiation might have played in prebiotic chemical evolution in aqueous media