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

Popp, R.A.; Bradshaw, B.S.; Hirsch, G.P.

Embryonic hemoglobins in heterozygous ..cap alpha..-thalassemic and normal fetuses were compared to study the effects of the deficient ..cap alpha.. chain on the synthesis of hemoglobins in the nucleated embryonic erythrocytes derived from the fetal yolk sac. Visual inspection of embryonic hemoglobins following acrylamide gel electrophoresis suggested that less hemoglobin EII (..cap alpha../sub 2/y/sub 2/) was formed in ..cap alpha..-thalassemic heterozygotes between 12/sup 1///sub 2/ and 14/sup 1///sub 2/ days of gestation. Quantitation of in vitro synthesis between 11/sup 1///sub 2/ and 13/sup 1///sub 2/ days of gestation confirmed that EII was synthesized less rapidly in ..cap alpha..-thalassemic fetuses. Inmore » contrast, the synthesis of EIII (..cap alpha../sub 2/z/sub 2/) was higher in ..cap alpha..-thalassemic than in normal fetuses at 12/sup 1///sub 2/ and 13/sup 1///sub 2/ days of gestation. Measurements of synthesis of individual chains in EI (x/sub 2/y/sub 2/) and EII showed that x-chain synthesis was normal and that ..cap alpha..-chain synthesis was deficient in ..cap alpha..-thalassemic fetuses at 11/sup 1///sub 2/ and 12/sup 1///sub 2/ days of gestation. There is still no proof for close linkage of x- and ..cap alpha..-chain genes in chromosome 11. Differences in the electrophoretic patterns of embryonic hemoglobins of ..cap alpha..-thalassemic and normal fetuses can be explained by normal synthesis of x chains, deficient synthesis of ..cap alpha.. chains, and a higher affinity of z than y for the reduced amount of ..cap alpha.. chain present in the nucleated embryonic erythrocytes of ..cap alpha..-thalassemic mice.« less

Jellyfish mesogloea collagen. Characterization of molecules as alpha 1 alpha 2 alpha 3 heterotrimers.

PubMed

Miura, S; Kimura, S

1985-12-05

The mesogloea collagen of a primitive animal, the jellyfish Stomolophus nomurai, belonging to the class Scyphozoa in the Coelenterata, was studied with respect to its chain structure. Most of the mesogloea collagen was solubilized by limited digestion with pepsin and isolated by selective precipitation at 0.9 m NaCl in 0.5 M acetic acid. Upon denaturation, the pepsin-solubilized collagen produced three distinct alpha chains, alpha 1, alpha 2, and alpha 3, in comparable amounts which were separable by CM-cellulose chromatography. The nonidentity of these alpha chains was confirmed by amino acid and carbohydrate analyses and peptide mapping. Furthermore, the introduction of intramolecular cross-links into native molecules by formaldehyde yielded a large proportion of gamma 123 chain with chain structure alpha 1 alpha 2 alpha 3, as judged by chromatographic behavior and peptide maps. We concluded that mesogloea collagen is comprised of alpha 1 alpha 2 alpha 3 heterotrimers and is chemically like vertebrate Type V collagen. On the other hand, sea anemone mesogloea collagen from the class Anthozoa was previously reported to comprise (alpha)3 homotrimers (Katzman, R. L., and Kang, A. H. (1972) J. Biol. Chem. 247, 5486-5489). On the basis of these findings, we assume that alpha 1 alpha 2 alpha 3 heterotrimers arose in evolution with the divergence of Scyphozoa and Anthozoa.

Overproduction of alpha chains provides a proton-insensitive component to the bluefish hemoglobin system.

PubMed

Bonaventura, Celia; Godette, Gerald; Stevens, Robert; Brenowitz, Michael; Henkens, Robert

2005-12-09

Expression of alpha and beta chains and their post-translational assembly into alpha(2)beta(2) tetramers is fundamental to the formation and function of most vertebrate hemoglobins. There is a strong evolutionary bias that favors expression of equal amounts of the two types of chains, because cooperativity, pH sensitivity, and anionic control of function occurs only for the alpha(2)beta(2) tetramers. Remarkably, an over-production of alpha chains, as in the pathological condition known as beta thalassemia in humans, is adaptive rather than pathological in the bluefish hemoglobin system. The thalassemia of the bluefish is a novel means of providing for oxygen uptake and delivery when low pH conditions incapacitate the highly pH-sensitive Root effect hemoglobins of the fish. Although fish often have pH-insensitive along with highly pH-sensitive hemoglobins, having pH-insensitive alpha chain monomers in circulation is an unusual structural variation. The role of bluefish alpha chains in oxygen transport is enabled by their remarkably lower oxygen affinity relative to human alpha chains. This is the first reported case of a thalassemic condition that is maintained in a species as an adaptive advantage.

Gene encoding the human. beta. -hexosaminidase. beta. chain: Extensive homology of intron placement in the. alpha. - and. beta. -chain genes

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

Proia, R.L.

1988-03-01

Lysosomal {beta}-hexosaminidase is composed of two structurally similar chains, {alpha} and {beta}, that are the products of different genes. Mutations in either gene causing {beta}-hexosaminidase deficiency result in the lysosomal storage disease GM2-gangliosidosis. To enable the investigation of the molecular lesions in this disorder and to study the evolutionary relationship between the {alpha} and {beta} chains, the {beta}-chain gene was isolated, and its organization was characterized. The {beta}-chain coding region is divided into 14 exons distributed over {approx}40 kilobases of DNA. Comparison with the {alpha}-chain gene revealed that 12 of the 13 introns interrupt the coding regions at homologous positions.more » This extensive sharing of intron placement demonstrates that the {alpha} and {beta} chains evolved by way of the duplication of a common ancestor.« less

Inhibition of laminin alpha 1-chain expression leads to alteration of basement membrane assembly and cell differentiation

PubMed Central

1996-01-01

The expression of the constituent alpha 1 chain of laminin-1, a major component of basement membranes, is markedly regulated during development and differentiation. We have designed an antisense RNA strategy to analyze the direct involvement of the alpha 1 chain in laminin assembly, basement membrane formation, and cell differentiation. We report that the absence of alpha 1-chain expression, resulting from the stable transfection of the human colonic cancer Caco2 cells with an eukaryotic expression vector comprising a cDNA fragment of the alpha 1 chain inserted in an antisense orientation, led to (a) an incorrect secretion of the two other constituent chains of laminin-1, the beta 1/gamma 1 chains, (b) the lack of basement membrane assembly when Caco2-deficient cells were cultured on top of fibroblasts, assessed by the absence of collagen IV and nidogen deposition, and (c) changes in the structural polarity of cells accompanied by the inhibition of an apical digestive enzyme, sucrase-isomaltase. The results demonstrate that the alpha 1 chain is required for secretion of laminin-1 and for the assembly of basement membrane network. Furthermore, expression of the laminin alpha 1-chain gene may be a regulatory element in determining cell differentiation. PMID:8609173

A study of excess H-alpha emission in chromospherically active M dwarf

NASA Technical Reports Server (NTRS)

Young, Arthur; Skumanich, Andrew; Stauffer, John R.; Harlan, Eugene; Bopp, Bernard W.

1989-01-01

Spectroscopic observations from three observatories are combined to study the properties of the excess H-alpha emission which characterizes the most chromospherically active subset of the M dwarf stars, known as the dMe stars. It is demonstrated that the excess H-alpha luminosity from these stars is a monotonically decreasing function of their (R-I) color, and evidence is presented which suggests that the product of the mean surface brightness and the mean filling factor of the emissive regions is essentially constant with color. Another significant result of the study is a linear correlation between the excess luminosity in H-alpha and the coronal X-ray luminosity.

Regulation of hepatic branched-chain alpha-keto acid dehydrogenase complex in rats fed a high-fat diet

USDA-ARS?s Scientific Manuscript database

Objective: Branched-chain alpha-keto acid dehydrogenase complex (BCKDC) regulates branched-chain amino acid (BCAA) metabolism at the level of branched chain alpha-ketoacid (BCKA) catabolism. It has been demonstrated that the activity of hepatic BCKDC is markedly decreased in type 2 diabetic animal...

Structural analyses of polymorphic transitions of sn-1, 3-distearoyl-2-oleoylglycerol (SOS) and sn-1, 3-dioleoyl-2-stearoylglycerol (OSO): assessment on steric hindrance of unsaturated and saturated acyl chain interactions.

PubMed

Yano, J; Sato, K; Kaneko, F; Small, D M; Kodali, D R

1999-01-01

Polymorphic transformations in two saturated-unsaturated mixed acid triacylglycerols, SOS (sn -1,3-distearoyl-2-oleoylglycerol) and OSO (sn -1,3-dioleoyl-2-stearoylglycerol), have been studied by FT-IR spectroscopy using deuterated specimens in which stearoyl chains are fully deuterated. A reversible phase transition between sub alpha and alpha and a series of irreversible transitions (alpha-->gamma-->beta'-->beta (beta2, beta1) for SOS and alpha-->beta'-->beta for OSO) were studied with an emphasis on the conformational ordering process of stearoyl and oleoyl chains. The alpha-->sub alpha reversible transition was due to the orientational change of stearoyl chains in the lateral directions from the hexagonal subcell to a perpendicularly packed one. As the first stage of the series of irreversible transitions from alpha to beta, the conformational ordering of saturated chains took place in the alpha-->gamma transition of SOS and in the alpha-->beta' transition of OSO; one stearoyl chain in SOS and OSO takes the all-trans conformation and the second stearoyl chain in SOS takes the bent conformation like those observed in the most stable beta-type. As the final stage, the ordering of unsaturated chains occurred in the beta'-->beta transition both for SOS and OSO. A conversion in the layered structure from bilayer to trilayer was also accompanied by the conformational ordering in the alpha-->gamma transition of SOS and in the beta'-->beta transition of OSO.

Alport Syndrome Diagnosis

MedlinePlus

... the presence or absence of the type IV collagen alpha-3, alpha-4 and alpha-5 chains ( ... linked Alport syndrome) is suspected. The type IV collagen alpha-5 chain (COL4A5) is normally present in ...

New lipid family that forms inverted cubic phases in equilibrium with excess water: molecular structure-aqueous phase structure relationship for lipids with 5,9,13,17-tetramethyloctadecyl and 5,9,13,17-tetramethyloctadecanoyl chains.

PubMed

Yamashita, Jun; Shiono, Manzo; Hato, Masakatsu

2008-10-02

With a view to discovering a new family of lipids that form inverted cubic phases, the aqueous phase behavior of a series of lipids with isoprenoid-type hydrophobic chains has been examined over a temperature range from -40 to 65 degrees C by using optical microscopy, DSC (differential scanning calorimetry), and SAXS (small-angle X-ray scattering) techniques. The lipids examined are those with 5,9,13,17-tetramethyloctadecyl and 5,9,13,17-tetramethyloctadecanoyl chains linked to a series of headgroups, that is, erythritol, pentaerythritol, xylose, and glucose. All of the lipid/water systems displayed a "water + liquid crystalline phase" two-phase coexistence state when sufficiently diluted. The aqueous phase structures of the most diluted liquid crystalline phases in equilibrium with excess water depend both on the lipid molecular structure and on the temperature. Given an isoprenoid chain, the preferred phase consistently follows a phase sequence of an H II (an inverted hexagonal phase) to a Q II (an inverted bicontinuous cubic phase) to an L alpha (a lamellar phase) as A* (cross-section area of the headgroup) increases. For a given lipid/water system, the phase sequence observed as the temperature increases is L alpha to Q II to H II. The present study allowed us to find four cubic phase-forming lipid species, PEOC 18+4 [mono- O-(5,9,13,17-tetramethyloctadecyl)pentaerythritol], beta-XylOC 18+4 [1- O-(5,9,13,17-tetramethyloctadecyl)-beta- d-xylopyranoside], EROCOC 17+4 [1- O-(5,9,13,17-tetramethyloctadecanoyl)erythritol], and PEOCOC 17+4 [mono- O-(5,9,13,17-tetramethyloctadecanoyl)pentaerythritol]. The values of T K (hydrated solid-liquid crystalline phase transition temperature) of the cubic phase-forming lipids are all below 0 degrees C. Quantitative analyses of the lipid molecular structure-aqueous phase structure relationship in terms of the experimentally evaluated "surfactant parameter" allow us to rationally select an optimum combination of hydrophilic/hydrophobic part of a lipid molecule that will form a desired phase in a desired temperature range.

Coordinated balancing of muscle oxidative metabolism through PGC-1{alpha} increases metabolic flexibility and preserves insulin sensitivity

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

Summermatter, Serge; Troxler, Heinz; Santos, Gesa

2011-04-29

Highlights: {yields} PGC-1{alpha} enhances muscle oxidative capacity. {yields} PGC-1{alpha} promotes concomitantly positive and negative regulators of lipid oxidation. {yields} Regulator abundance enhances metabolic flexibility and balances oxidative metabolism. {yields} Balanced oxidation prevents detrimental acylcarnitine and ROS generation. {yields} Absence of detrimental metabolites preserves insulin sensitivity -- Abstract: The peroxisome proliferator-activated receptor {gamma} coactivator 1{alpha} (PGC-1{alpha}) enhances oxidative metabolism in skeletal muscle. Excessive lipid oxidation and electron transport chain activity can, however, lead to the accumulation of harmful metabolites and impair glucose homeostasis. Here, we investigated the effect of over-expression of PGC-1{alpha} on metabolic control and generation of insulin desensitizing agentsmore » in extensor digitorum longus (EDL), a muscle that exhibits low levels of PGC-1{alpha} in the untrained state and minimally relies on oxidative metabolism. We demonstrate that PGC-1{alpha} induces a strictly balanced substrate oxidation in EDL by concomitantly promoting the transcription of activators and inhibitors of lipid oxidation. Moreover, we show that PGC-1{alpha} enhances the potential to uncouple oxidative phosphorylation. Thereby, PGC-1{alpha} boosts elevated, yet tightly regulated oxidative metabolism devoid of side products that are detrimental for glucose homeostasis. Accordingly, PI3K activity, an early phase marker for insulin resistance, is preserved in EDL muscle. Our findings suggest that PGC-1{alpha} coordinately coactivates the simultaneous transcription of gene clusters implicated in the positive and negative regulation of oxidative metabolism and thereby increases metabolic flexibility. Thus, in mice fed a normal chow diet, over-expression of PGC-1{alpha} does not alter insulin sensitivity and the metabolic adaptations elicited by PGC-1{alpha} mimic the beneficial effects of endurance training on muscle metabolism in this context.« less

[Perissodactyla: the primary structure of hemoglobins from the lowland tapir (Tapirus terrestris): glutamic acid in position 2 of the beta chains].

PubMed

Mazur, G; Braunitzer, G

1984-09-01

The hemoglobins from a lowland tapir (Tapirus terrestris) were analysed and the complete primary structure is described. The globin chains were separated on CM cellulose column in 8M urea and the amino-acid sequences were determined in the liquid phase sequenator. The results show that globin consists of two alpha chains (alpha I and alpha II) and beta major and beta minor components. The alpha chains differ only at one position: alpha I contains aspartic acid and alpha II glycine. The beta chains are heterogeneous: aspartic and glutamic acid were found at position beta 21 and beta 73 of the beta major components and asparagine and serine at position beta 139. In the beta minor components four positions were found with more than one amino acid, namely beta 2, beta 4, beta 6 and beta 56. The sequences are compared with those of man, horse and rhinoceros. Four residues of horse methemoglobin, which are involved in the alpha 1 beta 1 contacts are substituted in tapir hemoglobins. In the alpha chains: alpha 107(G14)Ser----Val, alpha 111-(G18) Val----Leu, alpha 115(GH3) Asn----Asp or Gly; in the beta chains: beta 116(G18) Arg----Gln. The amino acid at beta 2 of the major components is glutamic acid while glutamine and histidine are found in the minor components. Although glutamic acid, a binding site for ATP, does not interact with 2,3-bisphosphoglycerate, glutamine and histidine in the minor components are responsible for the slight effect of 2,3-bisphosphoglycerate on tapir hemoglobin.

Cecal parameters of rats fed diets containing grapefruit polyphenols and inulin as single supplements or in a combination.

PubMed

Zduńczyk, Zenon; Juśkiewicz, Jerzy; Estrella, Isabel

2006-09-01

We compared the effects of grapefruit flavonoids and inulin, as single dietary components or in a combination, on cecal fermentation in rats adapted to a semipurified diet. The experimental diets contained 0.3% flavonoid extract and 5% or 10% inulin and a combination of both supplements. The large bowel metabolism assessment was based on cecal parameters: bulk effect, pH, microbial enzymes activity, and short-chain fatty acid production. Both supplements induced significant enlargement of the cecal digesta weight. Acidification of cecal digesta was more pronounced, with a higher inulin addition to the diet. Cecal pH was the highest with the flavonoid-rich diets and lowest in the case of a simultaneous addition of flavonoids and a high content of inulin. The flavonoid extract applied as a single dietary supplement was observed to decrease the activity of bacterial beta-glucosidase and beta- and alpha-galactosidases in the cecal digesta. In contrast, addition of the grapefruit extract to inulin-containing diets increased the activity of alpha-glucosidase, alpha-galactosidase, and beta-galactosidase. Great accumulation of cecal digesta in rats consuming the flavonoid-diet caused a considerable increase in the short-chain fatty acid pool, mainly acetic acid. Inulin added to the diet decreased the excessive enlargement of digesta caused by dietary flavonoids. Dietary addition of inulin to the flavonoid-diet also normalized hydration of cecal digesta and significantly decreased the pH of digesta. The presence of polyphenols in the inulin-containing diets did not change total short-chain fatty acid production in the cecum of rats. Our results suggested that simultaneous intake of inulin and polyphenols can decrease the detrimental effects of the latter on cecal fermentation.

Characterization of heterotrimeric collagen molecules in a sea-pen (Cnidaria, Octocorallia).

PubMed

Tillet-Barret, E; Franc, J M; Franc, S; Garrone, R

1992-01-15

The collagen of a primitive invertebrate, the sea-pen Veretillum Cnidaria, Octocorallia), was studied with respect to its molecular-chain composition. The soft extracellular tissues (mesoglea) were solubilized by limited pepsin proteolysis and the collagen was isolated by selective precipitation at 0.7 M NaCl under acidic conditions. The pepsinized molecules were 260 nm in length, as demonstrated by electron microscope studies of rotary-shadowed molecules and of the segment-long-spacing crystallites obtained by dialysis against ATP. SDS/PAGE of the extract produced two main bands susceptible to bacterial collagenase, designated as the alpha 1 and alpha 2 chain, which were differentiated clearly by their CNBr cleavage products and the higher glycosylation rate of the alpha 2 chain. The latter finding corresponds with the high hydroxylysine content of the alpha 2 chain. The alpha 1/alpha 2 chain ratio observed in SDS/PAGE and the fact that only one peak was obtained by concanavalin-A affinity chromatography of a non-denatured 0.7 M NaCl extract demonstrate the alpha 1 [alpha 2]2 molecular structure of this collagen. These results contrast with data on the structure of other coelenterates (i.e. [alpha]3 for sea anemone collagen molecules and alpha 1 alpha 2 alpha 3 for jellyfish collagen molecules). They are discussed in relation to the evolution of collagen.

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

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

Fardella, C.E.; Hum, D.W.; Miller, W.L.

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

Primary structure of the hemoglobin beta-chain of rose-ringed parakeet (Psittacula krameri).

PubMed

Islam, A; Persson, B; Zaidi, Z H; Jörnvall, H

1989-08-01

The primary structure of Rose-ringed Parakeet hemoglobin beta-chain was established, completing the analysis of this hemoglobin. Comparison with other avian beta-chains show variations smaller than those for the corresponding alpha-chains. There are 11 amino acid exchanges in relationship to the only other characterized psittaciform beta-chain, and a total of 35 positions are affected by differences among all avian beta-chains analyzed (versus 61 for the alpha-chains). At three positions, the Psittacula beta-chain has residues unique to this species. Three alpha 1 beta 1 contacts are modified, by substitutions at positions beta 51, beta 116, and beta 125.

Salt bridge residues between I-Ak dimer of dimers alpha-chains modulate antigen presentation.

PubMed

Yadati, S; Nydam, T; Demian, D; Wade, T K; Gabriel, J L; Barisas, B G; Wade, W F

1999-03-15

Class II dimers of dimers are predicted to have functional significance in antigen presentation. The putative contact amino acids of the I-Ak class II dimer of dimers have been identified by molecular modeling based on the DR1 crystal structure (Nydam et al., Int. Immunol. 10, 1237,1998). We have previously reported the role in antigen presentation of dimer of dimers contact amino acids located in the C-terminal domains of the alpha- and beta-chains of class II. Our calculations show that residues Ealpha89 and Ralpha145 in the alpha2-domain form an inter alpha-chain salt bridge between pairs of alphabeta-heterodimers. Other residues, Qalpha92 and Nalpha115, may be involved in close association in that part of the alpha-chain. We investigated the role of these amino acids on class II expression and antigen presentation. Class II composed of an Ealpha89K substituted alpha-chain paired with a wt beta-chain exhibited inhibited antigen presentation and expression of alpha-chain serologic epitopes. In contrast, mutation of Ralpha145E had less affect on antigen presentation and did not affect I-Ak serologic epitopes. Interchanging charges of the salt bridge residues by expressing both Ralpha145E and Ealpha89K on the same chain obviated the large negative effect of the Ealpha89K mutation on antigen presentation but not on the serologic epitopes. Our results are similar for those reported for mutation of DR3's inter-chain salt bridge with the exception that double mutants did not moderate the DR3 defect. Interestingly, the amino acids differences between I-A and DR change the location of the inter-chain salt bridges. In DR1 these residues are located at positions Ealpha88 and Kalpha111; in I-Ak these residues are located at position Ealpha89 and Ralpha145. Inter alpha-chain salt bridges are thus maintained in various class II molecules by amino acids located in different parts of the alpha2-domain. This conservation of structure suggests that considerable functional importance may attach to the ionic interactions.

Biochemical characterization of a recombinant short-chain NAD(H)-dependent dehydrogenase/reductase from Sulfolobus acidocaldarius.

PubMed

Pennacchio, Angela; Giordano, Assunta; Pucci, Biagio; Rossi, Mosè; Raia, Carlo A

2010-03-01

The gene encoding a novel alcohol dehydrogenase that belongs to the short-chain dehydrogenases/reductases (SDRs) superfamily was identified in the aerobic thermoacidophilic crenarchaeon Sulfolobus acidocaldarius strain DSM 639. The saadh gene was heterologously overexpressed in Escherichia coli, and the protein (SaADH) was purified to homogeneity and characterized. SaADH is a tetrameric enzyme consisting of identical 28,978-Da subunits, each composed of 264 amino acids. The enzyme has remarkable thermophilicity and thermal stability, displaying activity at temperatures up to 75 degrees C and a 30-min half-inactivation temperature of ~90 degrees C, and shows good tolerance to common organic solvents. SaADH has a strict requirement for NAD(H) as the coenzyme, and displays a preference for the reduction of alicyclic, bicyclic and aromatic ketones and alpha-keto esters, but is poorly active on aliphatic, cyclic and aromatic alcohols, and shows no activity on aldehydes. The enzyme catalyses the reduction of alpha-methyl and alpha-ethyl benzoylformate, and methyl o-chlorobenzoylformate with 100% conversion to methyl (S)-mandelate [17% enantiomeric excess (ee)], ethyl (R)-mandelate (50% ee), and methyl (R)-o-chloromandelate (72% ee), respectively, with an efficient in situ NADH-recycling system which involves glucose and a thermophilic glucose dehydrogenase. This study provides further evidence supporting the critical role of the D37 residue in discriminating NAD(H) from NAD(P)H in members of the SDR superfamily.

Fibrinogen Lincoln: a new truncated alpha chain variant with delayed clotting.

PubMed

Ridgway, H J; Brennan, S O; Gibbons, S; George, P M

1996-04-01

A patient referred for preoperative investigation of prolonged bleeding and easy bruising was found to have increased thrombin and reptilase times; however, the thrombin catalysed release of fibrinopeptides A and B was normal. Analysis of five other family members, spanning three generations, indicated that three had a similar defect and suggested autosomal dominant inheritance. Non-reducing SDS-PAGE of purified fibrinogen from affected individuals showed that the 340 kD form of their fibrinogen ran as a doublet. SSCP (single-stranded conformational polymorphism) analysis of exon 5 of the A alpha gene, which encodes the C-terminal half of the chain, confirmed the presence of a mutation. Cycle sequencing of PCR amplified DNA revealed a 13 base pair deletion (nt 4758-4770), resulting in a frame-shift at Ala 475, which translates as four new amino acids before terminating at a new stop codon (-476His-Cys-Leu-Ala-Stop). The presence of a circulating truncated A alpha chain was confirmed when SDS-PAGE gels were probed with an alpha chain specific antisera; which showed that the variant A alpha chain comigrated with gamma chains. The truncation results in a variant A alpha chain with a deletion of 131 amino acids (480-610), and four new amino acids at the C-terminal.

Absence of Fc epsilonRI alpha chain results in upregulation of Fc gammaRIII-dependent mast cell degranulation and anaphylaxis. Evidence of competition between Fc epsilonRI and Fc gammaRIII for limiting amounts of FcR beta and gamma chains.

PubMed Central

Dombrowicz, D; Flamand, V; Miyajima, I; Ravetch, J V; Galli, S J; Kinet, J P

1997-01-01

In mouse mast cells, both Fc epsilonRI and Fc gammaRIII are alpha beta gamma2 tetrameric complexes in which different alpha chains confer IgE or IgG ligand recognition while the signaling FcR beta and gamma chains are identical. We used primarily noninvasive techniques (changes in body temperature, dye extravasation) to assess systemic anaphylactic responses in nonanesthetized wild-type, Fc epsilonRI alpha chain -/- and FcR gamma chain -/- mice. We confirm that systemic anaphylaxis in mice can be mediated largely through IgG1 and Fc gammaRIII and we provide direct evidence that these responses reflect activation of Fc gammaRIII rather than Fc gammaRI. Furthermore, we show that Fc gammaRIII-dependent responses are more intense in normal than in congenic mast cell-deficient KitW/KitW-v mice, indicating that Fc gammaRIII responses have mast cell-dependent and -independent components. Finally, we demonstrate that the upregulation of cell surface expression of Fc gammaRIII seen in Fc epsilonRI alpha chain -/- mice corresponds to an increased association of Fc gammaRIII alpha chains with FcR beta and gamma chains and is associated with enhanced Fc gammaRIII-dependent mast cell degranulation and systemic anaphylactic responses. Therefore, the phenotype of the Fc epsilonRI alpha chain -/- mice suggests that expression of Fc epsilonRI and Fc gammaRIII is limited by availability of the FcR beta and gamma chains and that, in normal mice, changes in the expression of one receptor (Fc epsilonRI) may influence the expression of functional responses dependent on the other (Fc gammaRIII). PMID:9062349

Three mouse models of human thalassemia.

PubMed Central

Martinell, J; Whitney, J B; Popp, R A; Russell, L B; Anderson, W F

1981-01-01

Three types of mice with globin gene mutations, called 352HB, 27HB, and Hbath-J, appear to be true animal models of human thalassemia. Expression of the alpha-globin genes in three stocks of mice, each one heterozygous for one of the alpha-globin mutations, was examined at the polypeptide, RNA, and DNA levels. alpha-Globin polypeptide chains, relative to beta-globin chains in heterozygous thalassemic mice, are present at approximately 80% of normal. The ratios of alpha-globin to beta-globin RNA sequences are also 75-80% of normal, exactly reflecting the alpha-globin to beta-globin chain ratios. In the case of mutant 352HB, at least one alpha-globin gene is deleted. Thalassemic mouse erythroid cells appear to compensate partially for the loss of half of their alpha-globin genes. Images PMID:6946454

Structural and functional characterization of a novel homodimeric three-finger neurotoxin from the venom of Ophiophagus hannah (king cobra).

PubMed

Roy, Amrita; Zhou, Xingding; Chong, Ming Zhi; D'hoedt, Dieter; Foo, Chun Shin; Rajagopalan, Nandhakishore; Nirthanan, Selvanayagam; Bertrand, Daniel; Sivaraman, J; Kini, R Manjunatha

2010-03-12

Snake venoms are a mixture of pharmacologically active proteins and polypeptides that have led to the development of molecular probes and therapeutic agents. Here, we describe the structural and functional characterization of a novel neurotoxin, haditoxin, from the venom of Ophiophagus hannah (King cobra). Haditoxin exhibited novel pharmacology with antagonism toward muscle (alphabetagammadelta) and neuronal (alpha(7), alpha(3)beta(2), and alpha(4)beta(2)) nicotinic acetylcholine receptors (nAChRs) with highest affinity for alpha(7)-nAChRs. The high resolution (1.5 A) crystal structure revealed haditoxin to be a homodimer, like kappa-neurotoxins, which target neuronal alpha(3)beta(2)- and alpha(4)beta(2)-nAChRs. Interestingly however, the monomeric subunits of haditoxin were composed of a three-finger protein fold typical of curaremimetic short-chain alpha-neurotoxins. Biochemical studies confirmed that it existed as a non-covalent dimer species in solution. Its structural similarity to short-chain alpha-neurotoxins and kappa-neurotoxins notwithstanding, haditoxin exhibited unique blockade of alpha(7)-nAChRs (IC(50) 180 nm), which is recognized by neither short-chain alpha-neurotoxins nor kappa-neurotoxins. This is the first report of a dimeric short-chain alpha-neurotoxin interacting with neuronal alpha(7)-nAChRs as well as the first homodimeric three-finger toxin to interact with muscle nAChRs.

Tyrosine sulfation in precursors of collagen V.

PubMed

Fessler, L I; Brosh, S; Chapin, S; Fessler, J H

1986-04-15

Radioactive labeling of p-collagens V, collagens V, and, to a small extent, of procollagen V occurred when [35S]sulfate was incubated with tendons or primary tendon cell cultures, or blood vessels and crops of 17- to 19-day-old chick embryos, or with lung slices from neonatal rats. Most or all of this label is in the form of 1 or more sulfated tyrosine residues/chain of p alpha 1(V), alpha 1(V), p alpha 1'(V), alpha 1'(V), p alpha 2(V), and alpha 2(V), and it remains attached through purification by dialysis, ammonium sulfate precipitation, CsCl-GdnCl2 equilibrium buoyant density and velocity sedimentations, ion-exchange chromatography, and sodium dodecyl sulfate gel electrophoresis. Radioactive tyrosine sulfate was identified in alkaline hydrolysates of these collagen V chains, after labeling the tissues with either [35S]sulfate or [3H]tyrosine, by electrophoretic and chromatographic comigration with a tyrosine sulfate standard. Tunicamycin A1, which inhibits the attachment of N-linked complex carbohydrate, did not interfere with the sulfation process. The tyrosine sulfate is located in a noncollagenous domain, which is probably adjacent to the amino end of the collagen helix, and is retained throughout the physiological proteolytic processing of procollagens V. After digestion with Staphylococcus aureus V8 protease, 35S-labeled p alpha 1(V) and alpha 1(V) chains gave the same map of labeled peptides, and this differed from the map given by p alpha 1'(V) and alpha 1'(V) chains. Little sulfation of p alpha 2(V) and alpha 2(V) chains occurs. The implications of these observations for the structure and properties of procollagens V and their derivatives are considered.

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

Martinell, J.; Whitney, J.B.; Popp, R.A.

Three types of mice with globin gene mutations, called 352HB, 27HB, and Hba/sup th-J/, appear to be true animal models of human thalassemia. Expression of the ..cap alpha..-globin genes in three stocks of mice, each one heterozygous for one of the ..cap alpha..-globin mutations, was examined at the polypeptide, RNA, and DNA levels. ..cap alpha..-globin polypeptide chains, relative to ..gamma..-globin chains in heterozygous thalassemic mice, are present at approximately 80% of normal. The ratios of ..cap alpha..-globin to ..gamma..-globin RNA sequences are also 75 to 80% normal, exactly reflecting the ..cap alpha..-globin to ..gamma..-globin chain ratios. In the case ofmore » mutant 352HB, at least one ..cap alpha..-globin gene is deleted. Thalassemic mouse erythroid cells appear to compensate partially for the loss of half of their ..cap alpha..-globin genes.« less

Cloning and purification of alpha-neurotoxins from king cobra (Ophiophagus hannah).

PubMed

He, Ying-Ying; Lee, Wei-Hui; Zhang, Yun

2004-09-01

Thirteen complete and three partial cDNA sequences were cloned from the constructed king cobra (Ophiophagus hannah) venom gland cDNA library. Phylogenetic analysis of nucleotide sequences of king cobra with those from other snake venoms revealed that obtained cDNAs are highly homologous to snake venom alpha-neurotoxins. Alignment of deduced mature peptide sequences of the obtained clones with those of other reported alpha-neurotoxins from the king cobra venom indicates that our obtained 16 clones belong to long-chain neurotoxins (seven), short-chain neurotoxins (seven), weak toxin (one) and variant (one), respectively. Up to now, two out of 16 newly cloned king cobra alpha-neurotoxins have identical amino acid sequences with CM-11 and Oh-6A/6B, which have been characterized from the same venom. Furthermore, five long-chain alpha-neurotoxins and two short-chain alpha-neurotoxins were purified from crude venom and their N-terminal amino acid sequences were determined. The cDNAs encoding the putative precursors of the purified native peptide were also determined based on the N-terminal amino acid sequencing. The purified alpha-neurotoxins showed different lethal activities on mice.

Identification of functional domains within the alpha and beta subunits of beta-hexosaminidase A through the expression of alpha-beta fusion proteins.

PubMed

Tse, R; Wu, Y J; Vavougios, G; Hou, Y; Hinek, A; Mahuran, D J

1996-08-20

There are three human beta-hexosaminidase isozymes which are composed of all possible dimeric combinations of an alpha and/or a beta subunit; A (alpha beta), and B (beta beta), and S (alpha alpha). The amino acid sequences of the two subunits are 60% identical. The homology between the two chains varies with the middle > the carboxy-terminal > > the amino-terminal portions. Although dimerization is required for activity, each subunit contains its own active site and differs in its substrate specificity and thermal stability. The presence of the beta subunit in hexosaminidase A also influences the substrate specificity of the alpha subunit; e.g., in vivo only the A heterodimer can hydrolyze GM2 ganglioside. In this report, we localize functional regions in the two subunits by cellular expression of alpha/beta fusion proteins joined at adjacently aligned residues. First, a chimeric alpha/beta chain was made by replacing the least well-conserved amino-terminal section of the beta chain with the corresponding alpha section. The biochemical characteristics of this protein were nearly identical to hexosaminidase B. Therefore, the most dissimilar regions in the subunits are not responsible for their dissimilar biochemical properties. A second fusion protein was made that also included the more homologous middle section of the alpha chain. This protein expressed the substrate specificity unique to isozymes containing an alpha subunit (A and S). We conclude that the region responsible for the ability of the alpha subunit to bind negatively charged substrates is located within residues alpha 132-283. Interestingly, the remaining carboxy-terminal section from the beta chain, beta 316-556, was sufficient to allow this chimera to hydrolyze GM2 ganglioside with 10% the specific activity of heterodimeric hexosaminidase A. Thus, the carboxy-terminal section of each subunit is likely involved in subunit-subunit interactions.

Enantiomeric excesses in meteoritic amino acids

NASA Technical Reports Server (NTRS)

Cronin, J. R.; Pizzarello, S.

1997-01-01

Gas chromatographic-mass spectral analyses of the four stereoisomers of 2-amino-2,3-dimethylpentanoic acid (dl-alpha-methylisoleucine and dl-alpha-methylalloisoleucine) obtained from the Murchison meteorite show that the L enantiomer occurs in excess (7.0 and 9.1%, respectively) in both of the enantiomeric pairs. Similar results were obtained for two other alpha-methyl amino acids, isovaline and alpha-methylnorvaline, although the alpha hydrogen analogs of these amino acids, alpha-amino-n-butyric acid and norvaline, were found to be racemates. With the exception of alpha-amino-n-butyric acid, these amino acids are either unknown or of limited occurrence in the biosphere. Because carbonaceous chondrites formed 4.5 billion years ago, the results are indicative of an asymmetric influence on organic chemical evolution before the origin of life.

On the chain length dependence of local correlations in polymer melts and a perturbation theory of symmetric polymer blends.

PubMed

Morse, David C; Chung, Jun Kyung

2009-06-14

The self-consistent field (SCF) approach to the thermodynamics of dense polymer liquids is based on the idea that short-range correlations in a polymer liquid are almost independent of how monomers are connected into polymers over larger scales. Some limits of this idea are explored in the context of a perturbation theory for symmetric polymer blends. We consider mixtures of two structurally identical polymers, A and B, in which the AB monomer pair interaction differs slightly from the AA and BB interactions by an amount proportional to a parameter alpha. An expansion of the free energy to first order in alpha yields an excess free energy of mixing per monomer of the form alphaz(N)phi(A)phi(B) in both lattice and continuum models, where z(N) is a measure of the number of intermolecular near neighbors per monomer in a one-component (alpha=0) reference liquid with chains of length N. The quantity z(N) decreases slightly with increasing N because the concentration of intramolecular near neighbors is slightly higher for longer chains, creating a slightly deeper intermolecular correlation hole. We predict that z(N)=z(infinity)[1+betaN(-1/2)], where N is an invariant degree of polymerization and beta=(6/pi)(3/2) is a universal coefficient. This and related predictions about the slight N dependence of local correlations are confirmed by comparison to simulations of a continuum bead-spring model and to published lattice Monte Carlo simulations. We show that a renormalized one-loop theory for blends correctly describes this N dependence of local liquid structure. We also propose a way to estimate the effective interaction parameter appropriate for comparisons of simulation data to SCF theory and to coarse-grained theories of corrections to SCF theory, which is based on an extrapolation of perturbation theory to the limit N-->infinity.

Lack of association between atopic eczema and the genetic variants of interleukin-4 and the interleukin-4 receptor alpha chain gene: heterogeneity of genetic backgrounds on immunoglobulin E production in atopic eczema patients.

PubMed

Tanaka, K; Sugiura, H; Uehara, M; Hashimoto, Y; Donnelly, C; Montgomery, D S

2001-10-01

The genetic background of atopic eczema might be heterogeneous and there is a possibility that immunoglobulin (Ig)E responsiveness in patients with atopic eczema is controlled separately from the development of atopic eczema. Although both interleukin (IL)-4 and the IL-4 receptor alpha chain have an important role for IgE production and are therefore possible candidate genes for atopy, it has not been clarified whether these genes play any roles in atopic eczema patients who have normal IgE productivity. We aimed to assess whether the polymorphisms of the IL-4 gene and the IL-4 receptor alpha chain gene play any roles in atopic eczema patients, particularly in patients who have normal IgE productivity. We determined the genotype with regard to polymorphisms in the genes for IL-4 and the IL-4 receptor alpha chain (- 589C/T of IL-4; Ile50Val, Ala375Glu and Arg551Gln of IL-4 receptor alpha chain) in patients with atopic eczema using the fluorogenic 5' nuclease assay. IL-4 and the IL-4 receptor alpha chain genotypes were not significantly associated with either total patients with atopic eczema or atopic eczema patients who had normal IgE productivity. The distribution of genotypes of IL-4-589C/T differed by the serum IgE levels in patients with atopic eczema. These results suggest that the polymorphisms in the IL-4 gene and the IL-4 receptor alpha chain gene play no role in the development of atopic eczema in patients who have normal IgE productivity.

Primary structure of the hemoglobin alpha-chain of rose-ringed parakeet (Psittacula krameri).

PubMed

Islam, A; Beg, O U; Persson, B; Zaidi, Z H; Jörnvall, H

1988-10-01

The structure of the hemoglobin alpha-chain of Rose-ringed Parakeet was determined by sequence degradations of the intact subunit, the CNBr fragments, and peptides obtained by digestion with staphylococcal Glu-specific protease and trypsin. Using this analysis, the complete alpha-chain structure of 21 avian species is known, permitting comparisons of the protein structure and of avian relationships. The structure exhibits differences from previously established avian alpha-chains at a total of 61 positions, five of which have residues unique to those of the parakeet (Ser-12, Gly-65, Ser-67, Ala-121, and Leu-134). The analysis defines hemoglobin variation within an additional avian order (Psittaciformes), demonstrates distant patterns for evaluation of relationships within other avian orders, and lends support to taxonomic conclusions from molecular data.

Analysis of the major epitope of the alpha2 chain of bovine type I collagen in children with bovine gelatin allergy.

PubMed

Hori, Hisae; Hattori, Shunji; Inouye, Sakae; Kimura, Akinori; Irie, Shinkichi; Miyazawa, Hiroshi; Sakaguchi, Masahiro

2002-10-01

Anaphylaxis to measles, mumps, and rubella vaccines has been reported. It has been found that most of these reactions to live vaccines are caused by type I allergy with the bovine gelatin present in the vaccines as an allergen. Gelatin mainly includes denatured type I collagen, which consists of alpha1 and alpha2 chains. We previously reported that allergic reactions to gelatin are caused by the type I collagen alpha2 (alpha2[I]) chain. To aid in the development of gelatin that has little or no allergenicity in human subjects, we investigated epitopes of bovine alpha2(I) chain with use of IgE in gelatin-sensitive children. Serum samples were collected from 15 patients who had systemic allergic reactions to vaccines and high levels of specific IgE to bovine gelatin. Eleven overlapping recombinant proteins that cover bovine alpha2(I) were prepared with a bacterial expression vector. We examined IgE reactivity to these recombinant proteins by means of ELISA. Fifteen peptides covering a major reactive recombinant protein were synthesized. The IgE-reacting epitope was identified by means of IgE-ELISA inhibition with these synthetic peptides and pooled serum from the patients. We found that of the 15 patients, 13 showed IgE reactivity to a recombinant protein (no. 3) spanning the central region of the collagenous domain ((418)Gly-(662)Pro). Furthermore, all 13 patients showed IgE reactivity to the 4-kd recombinant protein (no. 3a) spanning the region from (461)Pro to (500)Glu. In IgE-ELISA inhibition we found that a minimum IgE epitope of gelatin allergen was composed of the 10-amino-acid sequence (485)Ile-Pro-Gly-Glu-Phe-Gly-Leu-Pro-Gly-Pro(494). This sequence is not observed in the human type I collagen alpha1 and alpha2 chains, nor is it found in the bovine type I collagen alpha1 chain. We found that Ile-Pro-Gly-Glu-Phe-Gly-Leu-Pro-Gly-Pro is a major IgE epitope of the alpha2 chain of bovine type I collagen in patients with gelatin allergy. The degree of anaphylaxis to gelatin in vaccines might be reduced by digestion of this IgE-binding site in gelatin.

Direct binding of F actin to the cytoplasmic domain of the alpha 2 integrin chain in vitro

NASA Technical Reports Server (NTRS)

Kieffer, J. D.; Plopper, G.; Ingber, D. E.; Hartwig, J. H.; Kupper, T. S.

1995-01-01

The transmembrane integrins have been shown to interact with the cytoskeleton via noncovalent binding between cytoplasmic domains (CDs) of integrin beta chains and various actin binding proteins within the focal adhesion complex. Direct or indirect integrin alpha chain CD binding to the actin cytoskeleton has not been reported. We show here that actin, as an abundant constituent of focal adhesion complex proteins isolated from fibroblasts, binds strongly and specifically to alpha 2 CD, but not to alpha 1 CD peptide. Similar specific binding to alpha 2 CD peptide was seen for highly purified F actin, free of putative actin-binding proteins. The bound complex of actin and peptide was visualized directly by coprecipitation, and actin binding was abrogated by removal of a five amino acid sequence from the alpha 2 CD peptide. Our findings may explain the earlier observation that, while integrins alpha 2 beta 1 and alpha 1 beta 1 both bind to collagen, only alpha 2 beta 1 can mediate contraction of extracellular collagen matrices.

Expression and function of CD8 alpha/beta chains on rat and human mast cells.

PubMed

Kim, Mi-Sun; Kim, Sung-Hoon; Lee, Hye-Jung; Kim, Hyung-Min

2004-03-01

The expression and functional role of CD8 glycoprotein, a marker of cytotoxic/suppressor T lymphocytes and NK cells, were not studied on freshly isolated connective tissue type rat peritoneal mast cells, a rat mucosal type mast cell line (RBL 2H3), or human mast cell line (HMC-1). We used the reverse transcription-polymerase chain reaction (RT-PCR) and Western blot analysis, immunohistochemistry and enzyme-linked immunosorbent assay. RT-PCR and Western blot analysis identified the presence of CD8 alpha/beta chains on the mast cells, and immunohistochemistry confirmed CD8alpha expression on rat or human mast cells. Functional studies demonstrated that stimulation of CD8 alpha/beta chains on rat mast cells induced the secretion of tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6), which are regarded as important mediators during infection. However, co-stimulation with stem cell factor had no effect on CD8-induced mediator secretion. Our findings demonstrate novel biological roles of CD8 molecules in mast cells.

Excess entropy scaling for the segmental and global dynamics of polyethylene melts.

PubMed

Voyiatzis, Evangelos; Müller-Plathe, Florian; Böhm, Michael C

2014-11-28

The range of validity of the Rosenfeld and Dzugutov excess entropy scaling laws is analyzed for unentangled linear polyethylene chains. We consider two segmental dynamical quantities, i.e. the bond and the torsional relaxation times, and two global ones, i.e. the chain diffusion coefficient and the viscosity. The excess entropy is approximated by either a series expansion of the entropy in terms of the pair correlation function or by an equation of state for polymers developed in the context of the self associating fluid theory. For the whole range of temperatures and chain lengths considered, the two estimates of the excess entropy are linearly correlated. The scaled bond and torsional relaxation times fall into a master curve irrespective of the chain length and the employed scaling scheme. Both quantities depend non-linearly on the excess entropy. For a fixed chain length, the reduced diffusion coefficient and viscosity scale linearly with the excess entropy. An empirical reduction to a chain length-independent master curve is accessible for both dynamic quantities. The Dzugutov scheme predicts an increased value of the scaled diffusion coefficient with increasing chain length which contrasts physical expectations. The origin of this trend can be traced back to the density dependence of the scaling factors. This finding has not been observed previously for Lennard-Jones chain systems (Macromolecules, 2013, 46, 8710-8723). Thus, it limits the applicability of the Dzugutov approach to polymers. In connection with diffusion coefficients and viscosities, the Rosenfeld scaling law appears to be of higher quality than the Dzugutov approach. An empirical excess entropy scaling is also proposed which leads to a chain length-independent correlation. It is expected to be valid for polymers in the Rouse regime.

Dissociation of branched-chain alpha-keto acid dehydrogenase kinase (BDK) from branched-chain alpha-keto acid dehydrogenase complex (BCKDC) by BDK inhibitors.

PubMed

Murakami, Taro; Matsuo, Masayuki; Shimizu, Ayako; Shimomura, Yoshiharu

2005-02-01

Branched-chain alpha-keto acid dehydrogenase kinase (BDK) phosphorylates and inactivates the branched-chain alpha-keto acid dehydrogenase complex (BCKDC), which is the rate-limiting enzyme in the branched-chain amino acid catabolism. BDK has been believed to be bound to the BCKDC. However, recent our studies demonstrated that protein-protein interaction between BDK and BCKDC is one of the factors to regulate BDK activity. Furthermore, only the bound form of BDK appears to have its activity. In the present study, we examined effects of BDK inhibitors on the amount of BDK bound to the BCKDC using rat liver extracts. The bound form of BDK in the extracts of liver from low protein diet-fed rats was measured by an immunoprecipitation pull down assay with or without BDK inhibitors. Among the BDK inhibitors. alpha-ketoisocaproate, alpha-chloroisocaproate, and a-ketoisovalerate released the BDK from the complex. Furthermore, the releasing effect of these inhibitors on the BDK appeared to depend on their inhibition constants. On the other hand, clofibric acid and thiamine pyrophosphate had no effect on the protein-protein interaction between two enzymes. These results suggest that the dissociation of the BDK from the BCKDC is one of the mechanisms responsible for the action of some inhibitors to BDK.

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

Popp, R.A.; Bradshaw, B.S.; Skow, L.C.

Human alpha thalassemia is a congential disease causing a deficiency in the synthesis of alpha chains of hemoglobin. Homozygous individuals (hydrops fetalis) usually die during late pregnancy. Alpha thalassemia in mice has been induced by X-irradiation of males. Clinical symptoms in heterozygous mice are similar to those in man, e.g. microcytosis, reticulocytosis, poikilocytosis and hypochromia. Genetic studies showed that all viable alpha thalassemic progeny of matings of alpha thalassemic females and males were heterozygotes. Examination of preimplantation blastocysts flushed from the uterus of alpha thalassemic females at 86 hours after mating with alpha thalassemic males showed that about three-fourths ofmore » the embryos were composed of more than 32 blastomeres and had reached the early blastocyst stage while the remaining one-fourth of the embryos were composed of 32 or less blastomeres and were still at the morula stage of development. About one-fourth of the implantation sites did not contain live fetuses at 11 to 15 days of development. Histological examination at 5 to 8 days of gestation showed that the homozygous alpha thalassemic embryos implanted and developed to the late blastocyst stage when they became necrotic. At 11 to 15 days of development, the primitive nucleated erythrocytes appeared to be normal. However, the anucleated erythrocytes, which differentiate in the fetal liver, of alpha thalassemic fetuses contained abnormal eosinophilic inclusions that may be aggregations of beta chain polypeptides. The electrophoretic pattern of hemoglobins from alpha thalassemic fetuses and adults are distinguishable from those of their normal littermates. The differences can be explained on the basis of deficient alpha chain synthesis and the different affinities of the various kinds of alpha and non-alpha chains during the assembly of polypeptides of the tetrameric hemoglobin molecule.« less

Mouse interleukin-2 structure-function studies: substitutions in the first alpha-helix can specifically inactivate p70 receptor binding and mutations in the fifth alpha-helix can specifically inactivate p55 receptor binding.

PubMed Central

Zurawski, S M; Zurawski, G

1989-01-01

The function of two alpha-helical regions of mouse interleukin-2 were analyzed by saturation substitution analysis. The functional parts of the first alpha-helix (A) was defined as residues 31-39 by the observation that proline substitutions within this region inactivate the protein. Four residues within alpha-helix A, Leu31, Asp34, Leu35 and Leu38, were found to be crucial for biological activity. Structural modeling suggested that these four residues are clustered on one face of alpha-helix A. Residues 31 and 35 had to remain hydrophobic for the molecule to be functional. At residue 38 there was a preference for hydrophobic side chain residues, while at residue 34 some small side chain residues as well as acidic or amide side chain residues were functionally acceptable. Inactivating changes at residue 34 had no effect upon the ability of the protein to interact with the p55 receptor. Disruption of the fifth alpha-helix (E), which had little effect upon biological activity, resulted in an inability of the protein to interact with the p55 receptor. Mutagenesis of the alpha-helix E region demonstrated that alpha-helicity and the nature of the side chain residues in this region were unimportant for biological activity. The region immediately proximal to alpha-helix E was important only for the single intramolecular disulfide linkage. PMID:2583124

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

Rouyer-Fessard, P.; Garel, M.C.; Domenget, C.

The soluble pool of alpha hemoglobin chains present in blood or bone marrow cells was measured with a new affinity method using a specific probe, beta A hemoglobin chain labeled with ({sup 3}H)N-ethylmaleimide. This pool of soluble alpha chains was 0.067 {plus minus} 0.017% of hemoglobin in blood of normal adult, 0.11 {plus minus} 0.03% in heterozygous beta thalassemia and ranged from 0.26 to 1.30% in homozygous beta thalassemia intermedia. This elevated pool of soluble alpha chains observed in human beta thalassemia intermedia decreased 33-fold from a value of 10% of total hemoglobin in bone marrow cells to 0.3% inmore » the most dense red blood cells. The amount of insoluble alpha chains was measured by using the polyacrylamide gel electrophoresis in urea and Triton X-100. In beta thalassemia intermedia the amount of insoluble alpha chains was correlated with the decreased spectrin content of red cell membrane and was associated with a decrease in ankyrin and with other abnormalities of the electrophoretic pattern of membrane proteins. The loss and topology of the reactive thiol groups of membrane proteins was determined by using ({sup 3}H)N-ethylmaleimide added to membrane ghosts prior to urea and Triton X-100 electrophoresis. Spectrin and ankyrin were the major proteins with the most important decrease of thiol groups.« less

Decreased enzyme activity and contents of hepatic branched-chain alpha-keto acid dehydrogenase complex subunits in a rat model for type 2 diabetes mellitus.

PubMed

Bajotto, Gustavo; Murakami, Taro; Nagasaki, Masaru; Sato, Yuzo; Shimomura, Yoshiharu

2009-10-01

The mitochondrial branched-chain alpha-keto acid dehydrogenase complex (BCKDC) is responsible for the committed step in branched-chain amino acid catabolism. In the present study, we examined BCKDC regulation in Otsuka Long-Evans Tokushima Fatty (OLETF) rats both before (8 weeks of age) and after (25 weeks of age) the onset of type 2 diabetes mellitus. Long-Evans Tokushima Otsuka (LETO) rats were used as controls. Plasma branched-chain amino acid and branched-chain alpha-keto acid concentrations were significantly increased in young and middle-aged OLETF rats. Although the hepatic complex was nearly 100% active in all animals, total BCKDC activity and protein abundance of E1alpha, E1beta, and E2 subunits were markedly lower in OLETF than in LETO rats at 8 and 25 weeks of age. In addition, hepatic BCKDC activity and protein amounts were significantly decreased in LETO rats aged 25 weeks than in LETO rats aged 8 weeks. In skeletal muscle, E1beta and E2 proteins were significantly reduced, whereas E1alpha tended to increase in OLETF rats. Taken together, these results suggest that (1) whole-body branched-chain alpha-keto acid oxidation capacity is extremely reduced in OLETF rats independently of diabetes development, (2) the aging process decreases BCKDC activity and protein abundance in the liver of normal rats, and (3) differential posttranscriptional regulation for the subunits of BCKDC may exist in skeletal muscle.

Inferring High-Confidence Human Protein-Protein Interactions

DTIC Science & Technology

2012-01-01

comprised proteins that had the same specific func- tion or were subunits of the same protein complex, such as branched chain keto acid E1 alpha (BCKDHA...and branched chain keto acid E1 beta (BCKDHB) [3,29], and dynein cytoplasmic 2 intermediate chain 1 (D2LIC) and dynein cytoplasmic 2 heavy chain 1...474.3 28.0 1337.0 BCKDHA 5 Branched chain keto acid dehydro. E1, alpha BCKDHB 4 Branched chain keto acid dehydro. E1, beta 4 471.4 29.0 1337.5 ARTN 2

cap alpha. -D-Mannopyranosylmethyl-P-nitrophenyltriazene effects on the degradation and biosynthesis of N-linked oligosaccharide chains on. cap alpha. /sub 1/-acid glycoprotein by liver cells

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

Docherty, P.A.; Aronson, N.N. Jr.

1986-05-01

The effects of ..cap alpha..-D-mannopyranosylmethyl-p-nitrophenyltriazene (..cap alpha..-ManMNT) on the degradation and processing of oligosaccharide chains on ..cap alpha../sub 1/-acid glycoprotein (AGP) were studied. Addition of the triazene to a perfused liver blocked the complete degradation of endocytosed N-acetyl (/sup 14/C)glucosamine-labeled asialo-AGP and caused the accumulation of Man/sub 2/GlcNAc/sub 1/ fragments in the lysosome-enriched fraction of the liver homogenate. This compound also reduced the reincorporation of lysosomally-derived (/sup 14/C)GlcNAc into newly secreted glycoproteins. Cultured hepatocytes treated with the inhibitor synthesized and secreted fully-glycosylated AGP. However, the N-linked oligosaccharide chains on AGP secreted by the ..cap alpha..-ManMNT-treated hepatocytes remained sensitive to digestionmore » with endoglycosidase H, were resistant to neuraminidase, and consisted of Man/sub 9-7/GlcNAc/sub 2/ structures as analyzed by high resolution Bio-Gel P-4 chromatography. As measured by their resistance to cleavage by endoglycosidase H, the normal processing of all six carbohydrate chains on AGP to the complex form did not completely resume until nearly 24 h after triazene treatment. Since ManMNT is likely to irreversibly inactivate ..cap alpha..-D-mannosidases, the return of AGP to secretory forms with complex chains after 24 h probably resulted from synthesis of new processing enzymes.« less

The Buccaneer software for automated model building. 1. Tracing protein chains.

PubMed

Cowtan, Kevin

2006-09-01

A new technique for the automated tracing of protein chains in experimental electron-density maps is described. The technique relies on the repeated application of an oriented electron-density likelihood target function to identify likely C(alpha) positions. This function is applied both in the location of a few promising ;seed' positions in the map and to grow those initial C(alpha) positions into extended chain fragments. Techniques for assembling the chain fragments into an initial chain trace are discussed.

Chromosomal locations of mouse immunoglobulin genes.

PubMed Central

Valbuena, O; Marcu, K B; Croce, C M; Huebner, K; Weigert, M; Perry, R P

1978-01-01

The chromosomal locations of the structural genes coding for the constant portions of mouse heavy (H) and light chain immunoglobulins were studied by molecular hybridization techniques. Complementary DNA probes containing the constant-region sequences of kappa and lambdaI light chain and alpha, gamma2b, and mu heavy chain mRNAs were annealed to a large excess of DNA from a series of eight mouse-human hybrid cell lines that are deficient for various mouse chromosomes. The lines were scored as positive when a high proportion of a probe annealed and negative when an insignificant proportion annealed. Some lines were clearly negative for H and lambda and clearly positive for kappa. Others were positive or intermediate for lambda, positive for kappa and negative for H. Still others, including a line that was selected for the absence of the mouse X chromosome, were positive for all immunoglobulin species. These results demonstrate that the Clambda, Ckappa, and CH genes are located on different autosomes in the mouse. In contrast, the three heavy-chain families exhibited consistently uniform hybridization results, suggesting that the genes for Calpha, Cgamma, and Cmu are located on the same chromosome. A comparison of karyotypic data with hybridization data has limited the possible locations of the Ig genes to only a few chromosomes. PMID:96442

Identification of the NC1 domain of {alpha}3 chain as critical for {alpha}3{alpha}4{alpha}5 type IV collagen network assembly.

PubMed

LeBleu, Valerie; Sund, Malin; Sugimoto, Hikaru; Birrane, Gabriel; Kanasaki, Keizo; Finan, Elizabeth; Miller, Caroline A; Gattone, Vincent H; McLaughlin, Heather; Shield, Charles F; Kalluri, Raghu

2010-12-31

The network organization of type IV collagen consisting of α3, α4, and α5 chains in the glomerular basement membrane (GBM) is speculated to involve interactions of the triple helical and NC1 domain of individual α-chains, but in vivo evidence is lacking. To specifically address the contribution of the NC1 domain in the GBM collagen network organization, we generated a mouse with specific loss of α3NC1 domain while keeping the triple helical α3 chain intact by connecting it to the human α5NC1 domain. The absence of α3NC1 domain leads to the complete loss of the α4 chain. The α3 collagenous domain is incapable of incorporating the α5 chain, resulting in the impaired organization of the α3α4α5 chain-containing network. Although the α5 chain can assemble with the α1, α2, and α6 chains, such assembly is incapable of functionally replacing the α3α4α5 protomer. This novel approach to explore the assembly type IV collagen in vivo offers novel insights in the specific role of the NC1 domain in the assembly and function of GBM during health and disease.

Translation and assembly of HLA-DR antigens in Xenopus oocytes injected with mRNA from a human B-cell line.

PubMed Central

Long, E O; Gross, N; Wake, C T; Mach, J P; Carrel, S; Accolla, R; Mach, B

1982-01-01

HLA-DR antigens are polymorphic cell surface glycoproteins, expressed primarily in B lymphocytes and macrophages, which are thought to play an important role in the immune response. Two polypeptide chains, alpha and beta, are associated at the cell surface, and a third chain associates with alpha and beta intracellularly. RNA isolated from the human B-cell line Raji was injected in Xenopus laevis oocytes. Immunoprecipitates of translation products with several monoclonal antibodies revealed the presence of HLA-DR antigens similar to those synthesized in Raji cells. One monoclonal antibody was able to bind the beta chain after dissociation of the three polypeptide chains with detergent. The presence of all three chains was confirmed by two-dimensional gel electrophoresis. The glycosylation pattern of the three chains was identical to that observed in vivo, as evidenced in studies using tunicamycin, an inhibitor of N-linked glycosylation. The presence of alpha chains assembled with beta chains in equimolar ratio was further demonstrated by amino-terminal sequencing. An RNA fraction enriched for the three mRNAs, encoding alpha, beta, and intracellular chains, was isolated. This translation-assembly system and the availability of monoclonal antibodies make it possible to assay for mRNA encoding specific molecules among the multiple human Ia-like antigens. Images Fig. 1. Fig. 2. Fig. 3. Fig. 4. Fig. 5. PMID:6821356

pH regulation of mitochondrial branch chain alpha-keto acid transport and oxidation in rat heart mitochondria.

PubMed

Hutson, S M

1987-07-15

The kinetics of branched chain alpha-keto acid uptake and efflux were studied as a function of varied external and matrix pH. Matrix pH was determined by the distribution of 5,5'-dimethyloxazolidine-2,4-dione. When rat heart mitochondria were incubated under transport conditions at pH 7.0 with succinate as respiratory substrate, the matrix pH was significantly greater than 8.0. Matrix pH remained greater than or equal to 8.0 when the medium pH was varied from 6.3 to 8.3, and it was lowered below 8.0 by addition of 5 mM phosphate or uncoupler. No pH gradient was detectable when mitochondria were incubated in the presence of valinomycin and uncoupler. Efflux of alpha-ketoisocaproate or alpha-ketoisovalerate from rat heart mitochondria obeyed first order kinetics. Varying the external pH from 6.6 to 8.3 had no significant effect on efflux, and at an external pH of 7.0, the first order rate constant for efflux was not affected by decreasing the matrix pH. On the other hand, exchange was sensitive to changes in medium but not matrix pH. The K0.5 for external branched chain alpha-keto acid was lowered by changing the medium pH from 7.6 to 6.3. At medium pH values greater than or equal to 8.0 both K0.5 and Vmax were affected. Uptake was determined either by measuring initial rates or was calculated after measuring the first order approach to a final equilibrium value. Unlike efflux, uptake was sensitive to changes in both external and matrix pH. The rate of branched chain alpha-keto acid uptake was stimulated by decreasing the medium pH from 8.3 to 6.3 and by alkalinization of the mitochondrial matrix. The estimated external pK for proton binding was 6.9. The data indicate that the branched chain alpha-keto acid transporter is asymmetric, that is, binding sites for substrate on the inside and outside of the mitochondrial membrane are not identical. alpha-Ketoisocaproate oxidation was measured at 37 degrees C in isolated mitochondria over the pH range of 6.6 to 8.1. Changes in the rate of branched chain alpha-keto acid oxidation, particularly when ATP was added (increase delta pH), were found to parallel the pH effects observed on branched chain alpha-keto acid uptake. Therefore, transport, and by implication oxidation, can be regulated by pH changes within the physiological range. Furthermore, intracellular pH may affect the degree of compartmentation between the cytosolic and mitochondrial branched chain alpha-keto acid pools.

29 CFR 1926.1412 - Inspections.

Code of Federal Regulations, 2013 CFR

2013-07-01

... conditions, and proper operation. (vii) Chains and chain drive sprockets for excessive wear of sprockets and excessive chain stretch. (viii) Travel steering, brakes, and locking devices, for proper operation. (ix... decals originally supplied with the equipment by the manufacturer or otherwise required under this...

29 CFR 1926.1412 - Inspections.

Code of Federal Regulations, 2014 CFR

2014-07-01

... conditions, and proper operation. (vii) Chains and chain drive sprockets for excessive wear of sprockets and excessive chain stretch. (viii) Travel steering, brakes, and locking devices, for proper operation. (ix... decals originally supplied with the equipment by the manufacturer or otherwise required under this...

29 CFR 1926.1412 - Inspections.

Code of Federal Regulations, 2012 CFR

2012-07-01

... conditions, and proper operation. (vii) Chains and chain drive sprockets for excessive wear of sprockets and excessive chain stretch. (viii) Travel steering, brakes, and locking devices, for proper operation. (ix... decals originally supplied with the equipment by the manufacturer or otherwise required under this...

29 CFR 1926.1412 - Inspections.

Code of Federal Regulations, 2011 CFR

2011-07-01

... conditions, and proper operation. (vii) Chains and chain drive sprockets for excessive wear of sprockets and excessive chain stretch. (viii) Travel steering, brakes, and locking devices, for proper operation. (ix... decals originally supplied with the equipment by the manufacturer or otherwise required under this...

Hydration of alcohol clusters in 1-propanol-water mixture studied by quasielastic neutron scattering and an interpretation of anomalous excess partial molar volume.

PubMed

Misawa, M; Inamura, Y; Hosaka, D; Yamamuro, O

2006-08-21

Quasielastic neutron scattering measurements have been made for 1-propanol-water mixtures in a range of alcohol concentration from 0.0 to 0.167 in mole fraction at 25 degrees C. Fraction alpha of water molecules hydrated to fractal surface of alcohol clusters in 1-propanol-water mixture was obtained as a function of alcohol concentration. Average hydration number N(ws) of 1-propanol molecule is derived from the value of alpha as a function of alcohol concentration. By extrapolating N(ws) to infinite dilution, we obtain values of 12-13 as hydration number of isolated 1-propanol molecule. A simple interpretation of structural origin of anomalous excess partial molar volume of water is proposed and as a result a simple equation for the excess partial molar volume is deduced in terms of alpha. Calculated values of the excess partial molar volumes of water and 1-propanol and the excess molar volume of the mixture are in good agreement with experimental values.

Molecular cloning and characterization of adult Sparus aurata hemoglobin genes.

PubMed

Campo, Salvatore; Nastasi, Giancarlo; Fedeli, Donatella; D'Ascola, Angela; Campo, Giuseppe M; Avenoso, Angela; Ferlazzo, Alida; Calatroni, Alberto; Falcioni, Giancarlo

2010-04-01

Among Teleosts, Sparus aurata occupies a prominent place in the gastronomic and economic fields of the Mediterranean basin and other geographic districts. The knowledge of its molecular structures and functional features, such as hemoglobin, may be helpful to understand the adaptive biochemical mechanisms that allow this fish to live under extreme conditions, including fish farming. In Sparus aurata red blood cells two different alpha and one beta hemoglobin genes have been identified. The alpha1 gene codifies a putative protein of 144 amino acids, the alpha2 gene produces a protein of 143 amino acids, and the beta gene encodes a chain of 148 amino acids. Comparative analysis of various hemoglobins indicates that allosteric regulation can be modified by the substitution of one or a few key residues. The comparison of the percentage sequence differences for alpha and beta chains in fishes indicates that evolutionary relationships between different species may be helpful to understand the mechanisms of their differentiation from other vertebrates. Hemoglobin alpha and beta chains of about 50 teleostean temperate and Antarctic fishes were analyzed to build phylogenetic trees using different algorithms: the neighbor-joining method, the maximum likelihood approach, and the Bayesian inference computation. Sparus aurata alpha chains are positioned in a paraphyletic cluster, which includes the same subunit of Chrysophrys auratus and Seriola quinqueradiata, whereas the beta chain is on an homophyletic branch with that of Chrysophrys auratus. Therefore, the phylogenetic approach suggests that both Sparus aurata hemoglobin alpha genes are paralogues and may have derived from a duplication event.

Alternative splicing produces transcripts coding for alpha and beta chains of a hetero-dimeric phosphagen kinase.

PubMed

Ellington, W Ross; Yamashita, Daisuke; Suzuki, Tomohiko

2004-06-09

Glycocyamine kinase (GK) catalyzes the reversible phosphorylation of glycocyamine (guanidinoacetate), a reaction central to cellular energy homeostasis in certain animals. GK is a member of the phosphagen kinase enzyme family and appears to have evolved from creatine kinase (CK) early in the evolution of multi-cellular animals. Prior work has shown that GK from the polychaete Neanthes (Nereis) diversicolor exits as a hetero-dimer in vivo and that the two polypeptide chains (termed alpha and beta) are coded for by unique transcripts. In the present study, we demonstrate that the GK from a congener Nereis virens is also hetero-dimeric and is coded for by alpha and beta transcripts, which are virtually identical to the corresponding forms in N. diversicolor. The GK gene from N. diversicolor was amplified by PCR. Sequencing of the PCR products showed that the alpha and beta chains are the result of alternative splicing of the GK primary mRNA transcript. These results also strongly suggest that this gene underwent an early tandem exon duplication event. Full-length cDNAs for N. virens GKalpha and GKbeta were individually ligated into expression vectors and the resulting constructs used to transform Escherichia coli expression hosts. Regardless of expression conditions, minimal GK activity was observed in both GKalpha and GKbeta constructs. Inclusion bodies for both were harvested, unfolded in urea and alpha chains, beta chains and mixtures of alpha and beta chains were refolded by sequential dialysis. Only modest amounts of GK activity were observed when alpha and beta were refolded individually. In contrast, when refolded the alpha and beta mixture yielded highly active hetero-dimers, as validated by size exclusion chromatography, electrophoresis and mass spectrometry, with a specific activity comparable to that of natural GK. The above evidence suggests that there is a preference for hetero-dimer formation in the GKs from these two polychaetes. The evolution of the alternate splicing and an additional exon in these GKs, producing alpha and beta transcripts, can be viewed as a possible compensation for a mutation(s) in the original gene, which most likely coded for a homo-dimeric protein.

Bauhinia championii flavone inhibits apoptosis and autophagy via the PI3K/Akt pathway in myocardial ischemia/reperfusion injury in rats

PubMed Central

Jian, Jie; Xuan, Feifei; Qin, Feizhang; Huang, Renbin

2015-01-01

This study aimed to determine the effects of Bauhinia championii flavone (BCF) on myocardial ischemia/reperfusion injury (MI/RI) in rats and to explore potential mechanisms. The MI/RI model in rats was established by ligating the left anterior descending coronary artery for 30 minutes, then reperfusing for 3 hours. BCF at 20 mg/kg was given 20 minutes prior to ischemia via sublingual intravenous injection, with 24 μg/kg phosphoinositide 3-kinase inhibitor (PI3K; wortmannin) as a control. The creatine kinase-MB and nitric oxide content were assessed by colorimetry. The levels of mitochondrial permeability transition pores and tumor necrosis factor alpha were determined by an enzyme-linked immunosorbent assay. Cardiomyocyte apoptosis was detected by the terminal deoxynucleotidyl transferase dUTP nick end labeling assay. Additionally, the expression of PI3K, endothelial nitric oxide synthase, caspase-3, and Beclin1 was analyzed by fluorescence quantitative polymerase chain reaction and Western blotting, respectively. Akt and microtubule-associated protein 1 light chain 3-II protein levels were also evaluated. Pretreatment with BCF significantly decreased the levels of creatine kinase-MB, tumor necrosis factor alpha, and mitochondrial permeability transition pores, but increased the nitric oxide content. Furthermore, BCF inhibited apoptosis, downregulated caspase-3, Beclin1, and microtubule-associated protein 1 light chain 3-II, upregulated PI3K, and increased the protein levels of phosphorylated Akt and endothelial nitric oxide synthase. However, all of the previously mentioned effects of BCF were blocked when BCF was coadministered with wortmannin. In conclusion, these observations indicated that BCF has cardioprotective effects against MI/RI by reducing cell apoptosis and excessive autophagy, which might be related to the activation of the PI3K/Akt signaling pathway. PMID:26604691

Bauhinia championii flavone inhibits apoptosis and autophagy via the PI3K/Akt pathway in myocardial ischemia/reperfusion injury in rats.

PubMed

Jian, Jie; Xuan, Feifei; Qin, Feizhang; Huang, Renbin

2015-01-01

This study aimed to determine the effects of Bauhinia championii flavone (BCF) on myocardial ischemia/reperfusion injury (MI/RI) in rats and to explore potential mechanisms. The MI/RI model in rats was established by ligating the left anterior descending coronary artery for 30 minutes, then reperfusing for 3 hours. BCF at 20 mg/kg was given 20 minutes prior to ischemia via sublingual intravenous injection, with 24 μg/kg phosphoinositide 3-kinase inhibitor (PI3K; wortmannin) as a control. The creatine kinase-MB and nitric oxide content were assessed by colorimetry. The levels of mitochondrial permeability transition pores and tumor necrosis factor alpha were determined by an enzyme-linked immunosorbent assay. Cardiomyocyte apoptosis was detected by the terminal deoxynucleotidyl transferase dUTP nick end labeling assay. Additionally, the expression of PI3K, endothelial nitric oxide synthase, caspase-3, and Beclin1 was analyzed by fluorescence quantitative polymerase chain reaction and Western blotting, respectively. Akt and microtubule-associated protein 1 light chain 3-II protein levels were also evaluated. Pretreatment with BCF significantly decreased the levels of creatine kinase-MB, tumor necrosis factor alpha, and mitochondrial permeability transition pores, but increased the nitric oxide content. Furthermore, BCF inhibited apoptosis, downregulated caspase-3, Beclin1, and microtubule-associated protein 1 light chain 3-II, upregulated PI3K, and increased the protein levels of phosphorylated Akt and endothelial nitric oxide synthase. However, all of the previously mentioned effects of BCF were blocked when BCF was coadministered with wortmannin. In conclusion, these observations indicated that BCF has cardioprotective effects against MI/RI by reducing cell apoptosis and excessive autophagy, which might be related to the activation of the PI3K/Akt signaling pathway.

Expression of a partially deleted gene of human type II procollagen (COL2A1) in transgenic mice produces a chondrodysplasia

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

Vandenberg, P.; Khillan, J.S.; Prockop, D.J.

A minigene version of the human gene for type II procollagen (COL2AI) was prepared that lacked a large central region containing 12 of the 52 exons and therefore 291 of the 1523 codons of the gene. The construct was modeled after sporadic in-frame deletions of collagen genes that cause synthesis of shortened pro{alpha} chains that associate with normal pro{alpha} chains and thereby cause degradation of the shortened and normal pro{alpha} chains through a process called procollagen suicide. The gene construct was used to prepare five lines of transgenic mice expressing the minigene. A large proportion of the mice expressing themore » minigene developed a phenotype of a chondrodysplasia with dwarfism, short and thick limbs, a short snout, a cranial bulge, a cleft palate, and delayed mineralization of bone. A number of mice died shortly after birth. Microscopic examination of cartilage revealed decreased density and organization of collagen fibrils. In cultured chondrocytes from the transgenic mice, the minigene was expressed as shortened pro{alpha}1(II) chains that were disulfide-linked to normal mouse pro{alpha}1(II) chains. Therefore, the phenotype is probably explained by depletion of the endogenous mouse type II procollagen through the phenomenon of procollagen suicide.« less

Cardiac compartment-specific overexpression of a modified retinoic acid receptor produces dilated cardiomyopathy and congestive heart failure in transgenic mice.

PubMed

Colbert, M C; Hall, D G; Kimball, T R; Witt, S A; Lorenz, J N; Kirby, M L; Hewett, T E; Klevitsky, R; Robbins, J

1997-10-15

Retinoids play a critical role in cardiac morphogenesis. To examine the effects of excessive retinoid signaling on myocardial development, transgenic mice that overexpress a constitutively active retinoic acid receptor (RAR) controlled by either the alpha- or beta-myosin heavy chain (MyHC) promoter were generated. Animals carrying the alpha-MyHC-RAR transgene expressed RARs in embryonic atria and in adult atria and ventricles, but developed no signs of either malformations or disease. In contrast, beta-MyHC-RAR animals, where expression was activated in fetal ventricles, developed a dilated cardiomyopathy that varied in severity with transgene copy number. Characteristic postmortem lesions included biventricular chamber dilation and left atrial thrombosis; the incidence and severity of these lesions increased with increasing copy number. Transcript analyses showed that molecular markers of hypertrophy, alpha-skeletal actin, atrial natriuretic factor and beta-MyHC, were upregulated. Cardiac performance of transgenic hearts was evaluated using the isolated perfused working heart model as well as in vivo, by transthoracic M-mode echocardiography. Both analyses showed moderate to severe impairment of left ventricular function and reduced cardiac contractility. Thus, expression of a constitutively active RAR in developing atria and/ or in postnatal ventricles is relatively benign, while ventricular expression during gestation can lead to significant cardiac dysfunction.

Characterization of N-linked oligosaccharides assembled on secretory recombinant glucose oxidase and cell wall mannoproteins from the methylotrophic yeast Hansenula polymorpha.

PubMed

Kim, Moo Woong; Rhee, Sang Ki; Kim, Jeong-Yoon; Shimma, Yoh-ichi; Chiba, Yasunori; Jigami, Yoshifumi; Kang, Hyun Ah

2004-03-01

Presently almost no information is available on the oligosaccharide structure of the glycoproteins secreted from the methylotrophic yeast Hansenula polymorpha, a promising host for the production of recombinant proteins. In this study, we analyze the size distribution and structure of N-linked oligosaccharides attached to the recombinant glycoprotein glucose oxidase (GOD) and the cell wall mannoproteins obtained from H. polymorpha. Oligosaccharide profiling showed that the major oligosaccharide species derived from the H. polymorpha-secreted recombinant GOD (rGOD) had core-type structures (Man(8-12)GlcNAc(2)). Analyses using anti-alpha 1,3-mannose antibody and exoglycosidases specific for alpha 1,2- or alpha 1,6-mannose linkages revealed that the mannose outer chains of N-glycans on the rGOD have very short alpha 1,6 extensions and are mainly elongated in alpha 1,2-linkages without a terminal alpha 1,3-linked mannose addition. The N-glycans released from the H. polymorpha mannoproteins were shown to contain mostly mannose in their outer chains, which displayed almost identical size distribution and structure to those of H. polymorpha-derived rGOD. These results strongly indicate that the outer chain processing of N-glycans by H. polymorpha significantly differs from that by Saccharomyces cerevisiae, thus generating much shorter mannose outer chains devoid of terminal alpha 1,3-linked mannoses.

Secondary reduction of alpha7B integrin in laminin alpha2 deficient congenital muscular dystrophy supports an additional transmembrane link in skeletal muscle.

PubMed

Cohn, R D; Mayer, U; Saher, G; Herrmann, R; van der Flier, A; Sonnenberg, A; Sorokin, L; Voit, T

1999-03-01

The integrins are a large family of heterodimeric transmembrane cellular receptors which mediate the association between the extracellular matrix (ECM) and cytoskeletal proteins. The alpha7beta1 integrin is a major laminin binding integrin in skeletal and cardiac muscle and is thought to be involved in myogenic differentiation and migration processes. The main binding partners of the alpha7 integrin are laminin-1 (alpha1-beta1-gamma1), laminin-2 (alpha2-beta1-gamma1) and laminin-4 (alpha2-beta2-gamma1). Targeted deletion of the gene for the alpha7 integrin subunit (ITGA7) in mice leads to a novel form of muscular dystrophy. In the present study we have investigated the expression of two alternative splice variants, the alpha7B and beta1D integrin subunits, in normal human skeletal muscle, as well as in various forms of muscular dystrophy. In normal human skeletal muscle the expression of the alpha7 integrin subunit appeared to be developmentally regulated: it was first detected at 2 years of age. In contrast, the beta1D integrin could be detected in immature and mature muscle in the sarcolemma of normal fetal skeletal muscle at 18 weeks gestation. The expression of alpha7B integrin was significantly reduced at the sarcolemma in six patients with laminin alpha2 chain deficient congenital muscular dystrophy (CMD) (age >2 years). However, this reduction was not correlated with the amount of laminin alpha2 chain expressed. In contrast, the expression of the laminin alpha2 chain was not altered in the skeletal muscle of the alpha7 knock-out mice. These data argue in favor that there is not a tight correlation between the expression of the alpha7 integrin subunit and that of the laminin alpha2 chain in either human or murine dystrophic muscle. Interestingly, in dystrophinopathies (Duchenne and Becker muscular dystrophy; DMD/BMD) expression of alpha7B was upregulated irrespective of the level of dystrophin expression as shown by a strong sarcolemmal staining pattern even in young boys (age <2 years). The expression of the beta1D integrin subunit was not altered in any of our patients with different types of muscular dystrophy. In contrast, sarcolemmal expression of beta1D integrin was significantly reduced in the alpha7 integrin knock-out mice, whereas the expression of the components of the DGC was not altered. The secondary loss of alpha7B in laminin alpha2 chain deficiency defines a biochemical change in the composition of the plasma membrane resulting from a primary protein deficiency in the basal lamina. These findings, in addition to the occurrence of a muscular dystrophy in alpha7 deficient mice, implies that the alpha7B integrin is an important laminin receptor within the plasma membrane which plays a significant role in skeletal muscle function and stability.

Activity in X-ray-selected late-type stars

NASA Technical Reports Server (NTRS)

Takalo, Leo O.; Nousek, J. A.

1988-01-01

A spectroscopic study has been conducted of nine X-ray bright late-type stars selected from two Einstein X-ray surveys: the Columbia Astrophysical Laboratory Survey (five stars) and the CFA Medium Sensitivity Survey (MSS; four stars). Spectral classes were determined and radial and V sin(i) velocities were measured for the stars. Four of the Columbia Survey stars were found to be new RS CVn-type binaries. The fifth Columbia survey star was found to be an active G dwarf star without evidence for binarity. None of the four MSS stars were found to be either binaries or optically active stars. Activity in these stars was assessed by measuring the excess emission in H-alpha and the Ca II IRT (8498, 8542) lines in comparison with inactive stars of similar spectral types. A correlation was found between X-ray luminosity and V sin(i) and H-alpha line excess. The measured excess line emission in H-alpha was also correlated with V sin(i) but not with the IRT line excess.

Induction of IgA B cell differentiation of bone marrow-derived B cells by Peyer's patch autoreactive helper T cells.

PubMed

Kihira, T; Kawanishi, H

1995-08-01

The objective of this study was to demonstrate in vitro that bone marrow-derived pro/pre-B cells bearing mu mRNA can switch their Ig heavy-chain isotype to that of alpha mRNA-expressing B cells after contact with Peyer's patches-derived activated autoreactive CD4+ T cells. Bone marrow-derived pro/pre-B cells and activated autoreactive Peyer's patch, mesenteric lymph node, or spleen CD4+ T cells were co-cultured in the presence of recombinant (r) IL-2, rIL-7, and Con A for 3 days. The mixed cultured cells were isolated for preparation of total RNA. Dot/slot hybridization, using murine C mu (pu3741) and C alpha (P alpha J558) Ig heavy-chain cDNA probes, detected C mu and C alpha Ig heavy-chain mRNA transcripts. The magnitude of each mRNA expression was measured demsitometrically. In addition, the secreted class-specific Ig contents from the co-cultured supernatants were measured. The results indicate that activated autoreactive Peyer's patch and mesenteric lymph node CD4+ T cells provide a specific Ig heavy-chain switch from mu to alpha (Peyer's patch CD4+ T cells > mesenteric lymph node CD4+ T cells) in bone marrow-derived pro/pre-B cells and also assist to develop IgA-secreting plasma cells. The alpha heavy-chain switch and IgA production do not occur in the presence of activated autoreactive spleen CD4+ T cells. These results support the view that autoreactive gut Peyer's patch CD4+ T cells, at least, regulate IgA B cell heavy-chain switching and terminal differentiation during gut mucosal B cell development.

Effects of dietary excesses of branched-chain amino acids on the metabolism and tissue composition of lake trout (Salvelinus namaycush)

USGS Publications Warehouse

Hughes, S.G.; Rumsey, G.L.; Nesheim, M.C.

1984-01-01

1. Excesses of either leucine, isoleucine or valine were fed in separate experiments to determine if the branched-chain amino acid antagonism reported in other animals occur in trout (Salvelinus namaycush).2. Parameters measured include growth rate, feed utilization efficiency, plasma and muscle-free amino acids, carcass composition and branched-chain amino acid aminotransferase.3. Dietary excesses of leucine or isoleucine caused an increase in the valine requirement.4. The inability of leucine and isoleucine supplementations to ameliorate the effects of excess dietary valine are interpreted as a valine toxicity rather than an antagonism.

Maple syrup urine disease: The E1{beta} gene of human branched-chain {alpha}-ketoacid dehydrogenase complex has 11 rather than 10 exons, and the 3{prime} UTR in one of the two E1{beta} mRNAs arises from intronic sequences

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

Chuang, J.L.; Chuang, D.T.; Cox, R.P.

1996-06-01

Maple syrup urine disease (MSUD) or branched-chain ketoaciduria is caused by a deficiency in the mitochondrial branched-chain {alpha}-ketoacid dehydrogenase (BCKAD) complex. The clinical manifestations are characterized by accumulation of branched chain amino and {alpha}-ketoacids, which leads to severe cerebral edema with seizures, ketoacidosis, and mental retardation. The BCKAD complex comprises three catalytic components, i.e., a decarboxylase (E1) consisting of two E1{alpha} (M{sub r} = 46,000) and two E1{Beta} (M{sub r} = 37,500) subunits, a transacylase (E2) that contains 24 lipoic acid-bearing subunits, and a dehydrogenase (E3), which is a homodimeric flavoprotein. MSUD is genetically heterogeneous, since mutations in the E1{alpha}more » subunit (type IA MSUD), the E1{Beta} subunit (type IB), the E2 subunit (type II) and the E3 subunit (type III) have been described. The functional consequences of certain mutations in the BCKAD complex have been studied. 23 refs., 3 figs.« less

A dynamic alpha-beta inter-subunit agonist signaling complex is a novel feedback mechanism for regulating L-type Ca2+ channel opening.

PubMed

Zhang, Rong; Dzhura, Igor; Grueter, Chad E; Thiel, William; Colbran, Roger J; Anderson, Mark E

2005-09-01

L-type Ca2+ channels are macromolecular protein complexes in neurons and myocytes that open in response to cell membrane depolarization to supply Ca2+ for regulating gene transcription and vesicle secretion and triggering cell contraction. L-type Ca2+ channels include a pore-forming alpha and an auxiliary beta subunit, and alpha subunit openings are regulated by cellular Ca2+ through a mechanism involving the Ca2+-sensing protein calmodulin (CaM) and CaM binding motifs in the alpha subunit cytoplasmic C terminus. Here we show that these CaM binding motifs are "auto-agonists" that increase alpha subunit openings by binding the beta subunit. The CaM binding domains are necessary and sufficient for the alpha subunit C terminus to bind the beta subunit in vitro, and excess CaM blocks this interaction. Addition of CaM binding domains to native cardiac L-type Ca2+ channels in excised cell membrane patches increases openings, and this agonist effect is prevented by excess CaM. Recombinant LTCC openings are also increased by exogenous CaM binding domains by a mechanism requiring the beta subunit, and excess CaM blocks this effect. Thus, the bifunctional ability of the alpha subunit CaM binding motifs to competitively associate with the beta subunit or CaM provides a novel paradigm for feedback control of cellular Ca2+ entry.

[Vitamin E activity of some alpha-tocopherol derivatives and their effect on the ubiquinone level in rat liver in vitro].

PubMed

Donchenko, G V; Kovalenko, V N; Zolotashko, O M; Makovetskiĭ, V P; Basalkevich, E D; Sivachek, T E; Svishchuk, A A; Khalmuradov, A G

1979-01-01

An addition of alpha-tocopherol (I) and its synthetic derivatives (alpha-tocopheryl quinone (II), its short-chained analog (III), alpha-tocopherol lactone (IV), and short-chained alpha-tocopheryl acetate (V)) to the homogenized liver of vitamin E deficient rats resulted in a significant increase of ubiquinone after 2 hour incubation. Activity of the above derivatives (II-V) was not associated directly with their transformation into I or with a noticeable increase of the I content. There is a certain correlation between the chemical structure and the level of vitamin E activity of alpha-tocopherol derivatives that led to an increase in the ubiquinone content and prevented the decrease of tissue respiration and termination of pregnancy in rats.

Suppression of the chain nuclear fusion reaction based on the p+{sup 11}B reaction because of the deceleration of alpha particles

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

Shmatov, M. L., E-mail: M.Shmatov@mail.ioffe.ru

2016-09-15

It is shown that a rapid deceleration of alpha particles in matter of electron temperature up to 100 keV leads a strong suppression of the chain nuclear fusion reaction on the basis of the p+{sup 11}B reaction with the reproduction of fast protons in the α+{sup 11}B and n+{sup 10}B reactions. The statement that the chain nuclear fusion reaction based on the p+{sup 11}B reaction with an acceleration of {sup 11}B nuclei because of elastic alpha-particle scattering manifests itself in experiments at the PALS (Prague Asterix Laser System) facility is analyzed.

Excessive amounts of mu heavy chain block B-cell development.

PubMed

Zhu, Lingqiao; Chang, Cheong-Hee; Dunnick, Wesley

2011-09-01

Antigen-independent B-cell development occurs in several stages that depend on the expression of Ig heavy and light chain. We identified a line of mice that lacked mature B cells in the spleen. This mouse line carried approximately 11 copies of a transgene of the murine heavy chain constant region locus, and B-lineage cells expressed excessive amounts of the intracellular μ heavy chain. B-cell development failed in the bone marrow at the pro/pre B-cell transition, and examination of other lines with various copy numbers of the same transgene suggested that deficiencies in B-cell development increased with increased transgene copy number. Expression of a transgenic (Tg) light chain along with the Tg μ heavy chain led to minimal rescue of B-cell development in the bone marrow and B cells in the spleen. There are several potential mechanisms for the death of pro/pre B cells as a consequence of excess heavy chain expression.

Characterization of the major cyanogen bromide fragment of alpha-A crystallin

NASA Technical Reports Server (NTRS)

Ifeanyi, F.; Takemoto, L.; Spooner, B. S. (Principal Investigator)

1991-01-01

Alpha crystallin from the bovine lens has been digested with cyanogen bromide, and the major fragment (CB-1) has been purified using reverse phase HPLC. Characterization of this fragment by Edman degradation and antisera to synthetic peptides indicates that it originates from alpha-A crystallin, but lacks the N-terminal methionine and the last 35 amino acids from the C-terminus of the molecule. The purified CB-1 fragment binds as well as native alpha crystallin to lens membrane, but is unable to self-assemble into the correct size of high molecular weight oligomeric complexes characteristic of the intact alpha-A chain. Together, these results demonstrate that the alpha-A chain is comprised of at least two functional domains, one of which is involved in binding of alpha-A crystallin to lens membrane, and another which is necessary for correct self-assembly of the molecule into high molecular weight oligomers.

Three-dimensional crystal structure of recombinant murine interferon-beta.

PubMed Central

Senda, T; Shimazu, T; Matsuda, S; Kawano, G; Shimizu, H; Nakamura, K T; Mitsui, Y

1992-01-01

The crystal structure of recombinant murine interferon-beta (IFN-beta) has been solved by the multiple isomorphous replacement method and refined to an R-factor of 20.5% against 2.6 A X-ray diffraction data. The structure shows a variant of the alpha-helix bundle with a new chain-folding topology, which seems to represent a basic structural framework of all the IFN-alpha and IFN-beta molecules belonging to the type I family. Functionally important segments of the polypeptide chain, as implied through numerous gene manipulation studies carried out so far, are spatially clustered indicating the binding site(s) to the receptor(s). Comparison of the present structure with those of other alpha-helical cytokine proteins, including porcine growth hormone, interleukin 2 and interferon gamma, indicated either a topological similarity in chain folding or a similar spatial arrangement of the alpha-helices. Images PMID:1505514

Effect of swainsonine on the processing of the asparagine-linked carbohydrate chains of alpha 1-antitrypsin in rat hepatocytes. Evidence for the formation of hybrid oligosaccharides.

PubMed

Gross, V; Tran-Thi, T A; Vosbeck, K; Heinrich, P C

1983-03-25

The biosynthesis of the proteinase inhibitor alpha 1-antitrypsin has been studied in rat hepatocyte primary cultures. Newly synthesized alpha 1-antitrypsin was found in hepatocytes as a glycoprotein of an apparent molecular weight of 49,000 carrying oligosaccharide side chains of the high mannose type. In the hepatocyte medium a secreted alpha 1-antitrypsin of an apparent molecular weight of 54,000 could be identified as a glycoprotein with carbohydrate chains of the complex type. Pulse-chase experiments revealed a precursor-product relationship for the two forms of alpha 1-antitrypsin. When the hepatocytes were treated with swainsonine, an intracellular form of alpha 1-antitrypsin with an apparent molecular weight of 49,000 indistinguishable from that of control cells was found. However, the alpha 1-antitrypsin secreted from swainsonine-treated hepatocytes was different from that present in control media. It was characterized by a lower apparent molecular weight (51,000), a higher amount of [3H]mannose incorporation, half as much incorporation of [3H]galactose, and the same amount of [3H]fucose incorporation compared to alpha 1-antitrypsin of control media. In contrast to the 54,000 complex type alpha 1-antitrypsin from control media the 51,000 alpha 1-antitrypsin from the medium of swainsonine-treated cells was found to be susceptible to the action of endoglucosaminidase H, even when fucose was attached to the proximal GlcNAc residue. alpha 1-Antitrypsin secreted from swainsonine-treated cells combines features usually associated with either high mannose or complex type oligosaccharides and therefore represents a hybrid structure. In spite of its effect on the carbohydrate part of alpha 1-antitrypsin swainsonine did not impair the secretion of the incompletely processed glycoprotein.

Phenotypic and functional characterization of T cells from patients with myasthenia gravis.

PubMed Central

Mokhtarian, F; Pino, M; Ofosu-Appiah, W; Grob, D

1990-01-01

A study of cell surface phenotypes of PBL of myasthenia gravis (MG) patients showed that their T cells had a significantly higher percentage of 4B4+ T cells (the helper/inducer subset) than age- and sex-matched controls. The PBL of MG patients proliferated significantly higher than those of normal subjects (NS) in response to the purified alpha chain of the acetylcholine receptor (AChR). Anti-AChR antibody was present in sera of 88% of MG and none of the NS. The PBL B cells from MG only, when cultured with autologous T cells and stimulated with either pokeweed mitogen (69%), or AChR-alpha chain (38%), secreted antibody to AChR-alpha chain, whereas T and B cells alone secreted no antibody. T cells from PBL of MG patients were more readily cloned than T cells of NS, by limiting dilution, in the presence of recombinant IL-2 and in the absence of AChR-alpha chain. About 50% of T cell clones from MG patients, compared to none from NS, proliferated to AChR-alpha chain. This response was HLA-DR restricted. MG T cell clones did not display significant cytotoxic activity, as compared to control T cell clones. Our results indicate that in MG, 4B4+ regulatory T cells play their role in the pathogenesis of MG, not by cytotoxicity, but more likely by their ability to stimulate specific antibody production by B cells. Images PMID:1979338

Risk of Lung Cancer Mortality in Nuclear Workers from Internal Exposure to Alpha Particle-emitting Radionuclides

PubMed Central

Atkinson, Will; Bérard, Philippe; Bingham, Derek; Birchall, Alan; Blanchardon, Eric; Bull, Richard; Guseva Canu, Irina; Challeton-de Vathaire, Cécile; Cockerill, Rupert; Do, Minh T.; Engels, Hilde; Figuerola, Jordi; Foster, Adrian; Holmstock, Luc; Hurtgen, Christian; Laurier, Dominique; Puncher, Matthew; Riddell, Anthony E.; Samson, Eric; Thierry-Chef, Isabelle; Tirmarche, Margot; Vrijheid, Martine; Cardis, Elisabeth

2017-01-01

Background: Carcinogenic risks of internal exposures to alpha-emitters (except radon) are poorly understood. Since exposure to alpha particles—particularly through inhalation—occurs in a range of settings, understanding consequent risks is a public health priority. We aimed to quantify dose–response relationships between lung dose from alpha-emitters and lung cancer in nuclear workers. Methods: We conducted a case–control study, nested within Belgian, French, and UK cohorts of uranium and plutonium workers. Cases were workers who died from lung cancer; one to three controls were matched to each. Lung doses from alpha-emitters were assessed using bioassay data. We estimated excess odds ratio (OR) of lung cancer per gray (Gy) of lung dose. Results: The study comprised 553 cases and 1,333 controls. Median positive total alpha lung dose was 2.42 mGy (mean: 8.13 mGy; maximum: 316 mGy); for plutonium the median was 1.27 mGy and for uranium 2.17 mGy. Excess OR/Gy (90% confidence interval)—adjusted for external radiation, socioeconomic status, and smoking—was 11 (2.6, 24) for total alpha dose, 50 (17, 106) for plutonium, and 5.3 (−1.9, 18) for uranium. Conclusions: We found strong evidence for associations between low doses from alpha-emitters and lung cancer risk. The excess OR/Gy was greater for plutonium than uranium, though confidence intervals overlap. Risk estimates were similar to those estimated previously in plutonium workers, and in uranium miners exposed to radon and its progeny. Expressed as risk/equivalent dose in sieverts (Sv), our estimates are somewhat larger than but consistent with those for atomic bomb survivors. See video abstract at, http://links.lww.com/EDE/B232. PMID:28520643

Identification of a major continuous epitope of human alpha crystallin

NASA Technical Reports Server (NTRS)

Takemoto, L.; Emmons, T.; Spooner, B. S. (Principal Investigator)

1992-01-01

Human lens proteins were digested with trypsin or V8 protease, and the resulting peptides resolved on a C18 reverse phase column. Fractions from this column were probed with polyclonal antiserum made against the whole alpha crystallin molecule. Peptides in the seropositive fraction were purified to homogeneity, then characterized by mass spectral analysis and partial Edman degradation. The tryptic and V8 digests contained only one seropositive peptide that was derived from the C-terminal region of the alpha-A molecule. To determine the exact boundaries of the epitope, various size analogues of this region were synthesized and probed with anti-alpha serum. Together, these studies demonstrate that the major continuous epitope of the alpha-A chain includes the sequence KPTSAPS, corresponding to residues 166-172 of the human alpha-A crystallin chain.

Cardiac compartment-specific overexpression of a modified retinoic acid receptor produces dilated cardiomyopathy and congestive heart failure in transgenic mice.

PubMed Central

Colbert, M C; Hall, D G; Kimball, T R; Witt, S A; Lorenz, J N; Kirby, M L; Hewett, T E; Klevitsky, R; Robbins, J

1997-01-01

Retinoids play a critical role in cardiac morphogenesis. To examine the effects of excessive retinoid signaling on myocardial development, transgenic mice that overexpress a constitutively active retinoic acid receptor (RAR) controlled by either the alpha- or beta-myosin heavy chain (MyHC) promoter were generated. Animals carrying the alpha-MyHC-RAR transgene expressed RARs in embryonic atria and in adult atria and ventricles, but developed no signs of either malformations or disease. In contrast, beta-MyHC-RAR animals, where expression was activated in fetal ventricles, developed a dilated cardiomyopathy that varied in severity with transgene copy number. Characteristic postmortem lesions included biventricular chamber dilation and left atrial thrombosis; the incidence and severity of these lesions increased with increasing copy number. Transcript analyses showed that molecular markers of hypertrophy, alpha-skeletal actin, atrial natriuretic factor and beta-MyHC, were upregulated. Cardiac performance of transgenic hearts was evaluated using the isolated perfused working heart model as well as in vivo, by transthoracic M-mode echocardiography. Both analyses showed moderate to severe impairment of left ventricular function and reduced cardiac contractility. Thus, expression of a constitutively active RAR in developing atria and/ or in postnatal ventricles is relatively benign, while ventricular expression during gestation can lead to significant cardiac dysfunction. PMID:9329959

Contribution of alpha3(IV)alpha4(IV)alpha5(IV) Collagen IV to the Mechanical Properties of the Glomerular Basement Membrane

NASA Astrophysics Data System (ADS)

Gyoneva, Lazarina

The glomerular basement membrane (GBM) is a vital part of the blood-urine filtration barrier in the kidneys. In healthy GBMs, the main tension-resisting component is alpha3(IV)alpha4(IV)alpha5(IV) type IV collagen, but in some diseases it is replaced by other collagen IV isoforms. As a result, the GBM becomes leaky and disorganized, ultimately resulting in kidney failure. Our goal is to understanding the biomechanical aspects of the alpha3(IV)alpha4(IV)alpha5(IV) chains and how their absence could be responsible for (1) the initial injury to the GBM and (2) progression to kidney failure. A combination of experiments and computational models were designed for that purpose. A model basement membrane was used to compare experimentally the distensibility of tissues with the alpha3(IV)alpha4(IV)alpha5(IV) chains present and missing. The experiments showed basement membranes containing alpha3(IV)alpha4(IV)alpha5(IV) chains were less distensible. It has been postulated that the higher level of lateral cross-linking (supercoiling) in the alpha3(IV)alpha4(IV)alpha5(IV) networks contributes additional strength/stability to basement membranes. In a computational model of supercoiled networks, we found that supercoiling greatly increased the stiffness of collagen IV networks but only minimally decreased the permeability, which is well suited for the needs of the GBM. It is also known that the alpha3(IV)alpha4(IV)alpha5(IV) networks are more protected from enzymatic degradation, and we explored their significance in GBM remodeling. Our simulations showed that the more protected network was needed to prevent the system from entering a dangerous feedback cycle due to autoregulation mechanisms in the kidneys. Overall, the work adds to the evidence of biomechanical differences between the alpha3(IV)alpha4(IV)alpha5(IV) networks and other collagen IV networks, points to supercoiling as the main source of biomechanical differences, discusses the suitability of alpha3(IV)alpha4(IV)alpha5(IV) networks to meet the mechanics and permeability needs of the GBM, and explores the role of biomechanics and enzymatic digestion in GBM remodeling.

Interaction of a chick skin collagen fragment (alpha1-CB5) with human platelets. Biochemical studies during the aggregation and release reaction.

PubMed

Chiang, T M; Beachey, E H; Kang, A H

1975-09-10

The denatured alpha1(I) chain and the cyanogen bromide peptide, alpha1(I)-CB5, of chick skin collagen cause the release of serotonin and leakage of lactic dehydrogenase from human platelets in a manner similar to the release reaction mediated by adenosine diphosphate and native collagen. These peptides also cause a decrease in the level of adenosine 3':5'-monophosphate (cAMP) in platelets. Adenylate cyclase activity of platelets is partially inhibited by these peptides as well as by native collagen, ADP, and epinephrine, but cAMP phosphodiesterase activity is unaltered by these substances. In contrast, the level of platelet guanosine 3':5'-monophosphate (cGMP) is increased by the collagen peptides as well as the other aggregating agents. The increase is associated with increased guanylate cyclase, but normal cGMP phosphodiesterase activities of platelets. Optical rotatory and viscometric measurements of the alpha1 chains and alpha1-CB5 of chick skin in 0.01 M phosphate/0.15 M sodium chloride, pH 7.4, at various temperatures as a function of time indicate that no detectable renaturation occurs at 37 degrees for at least 30 min of observation. Molecular sieve chromatography of alpha1-CB5 in the phosphate buffer at 37 degrees shows that its elution position is identical to that performed under denaturing conditions (at 45 degrees) with no evidence of higher molecular weight aggregates, and the alpha1-CB5 glycopeptide fraction eluting from the column at the position of its monomer retains the platelet aggregating activity. Additionally, electron microscopic examination of the platelet-rich plasma that had been reacted with these peptides fail to show any ordered collagen structures. These data indicate that the denatured alpha1 chain and alpha1-CB5 glycopeptide of chick skin collagen mediate platelet aggregation through the "physiologic" release reaction in a manner similar to that induced by other aggregating agents such as ADP, epinephrine, or native collagen, and support the conclusion that the aggregating activity of the alpha1 chain and alpha1-CB5 is not likely to be due to the formation of polymerized products.

A Proposed Mechanism for the Thermal Denaturation of a Recombinant Bacillus Halmapalus Alpha-amylase - the Effect of Calcium Ions

NASA Technical Reports Server (NTRS)

Nielsen, Anders D.; Pusey, Marc L.; Fuglsang, Claus C.; Westh, Peter

2003-01-01

The thermal stability of a recombinant alpha-amylase from Bacillus halmapalus alpha-amylase (BHA) has been investigated using circular dichroism spectroscopy (CD) and differential scanning calorimetry (DSC). This alpha-amylase is homologous to other Bacillus alpha-amylases where previous crystallographic studies have identified the existence of 3 calcium binding sites in the structure. Denaturation of BHA is irreversible with a Tm of approximately 89 C, and DSC thermograms can be described using a one-step irreversible model. A 5 C increase in T(sub m) in the presence of 10 fold excess CaCl2 was observed. However, a concomitant increase in the tendency to aggregate was also observed. The presence of 30-40 fold excess calcium chelator (EDTA or EGTA) results in a large destabilization of BHA corresponding to about 40 C lower T(sub m), as determined by both CD and DSC. Ten fold excess EGTA reveals complex DSC thermograms corresponding to both reversible and irreversible transitions, which possibly originate from different populations of BHA:calcium complexes. The observations in the present study have, in combination with structural information of homologous alpha-amylases, provided the basis for the proposal of a simple denaturation mechanism of BHA. The proposed mechanism describes the irreversible thermal denaturation of different BHA:calcium complexes and the calcium binding equilibrium involved. Furthermore, the model accounts for a temperature induced reversible structural change associated with calcium binding.

Altered procollagen gene expression in mid-gestational mouse excisional wounds.

PubMed

Goldberg, Stephanie R; Quirk, Gerald L; Sykes, Virginia W; Kordula, Tomasz; Lanning, David A

2007-11-01

Many pathologic conditions are characterized by excessive tissue contraction and scar formation. Previously, we developed a murine model of excisional wound healing in which mid-gestational wounds heal scarlessly compared with late-gestational wounds. We theorized that variations in procollagen gene expression may contribute to the scarless and rapid closure. Time-dated pregnant FVB strain mice underwent laparotomy and hysterotomy on embryonic days 15 (E15) and 18 (E18). Full-thickness, excisional wounds (3 mm) were made on each of 4 fetuses per doe and then harvested at 32, 48, or 72 h. Control tissue consisted of age-matched normal fetal skin. Procollagen types 1alpha1, 1alpha2, and 3 gene expressions were measured by real-time polymerase chain reaction and normalized to glyceraldehyde-3-phosphate dehydrogenase. Trichrome staining was also performed. Procollagen 1alpha1 expression was decreased in E15 wounds at 32 h compared with their normal skin groups. Procollagen types 1alpha2 and 3 expressions were both increased in the E15 groups compared with the E18 groups at 48 h. At 72 h, the E15 wounds had a collagen density similar to the surrounding normal skin while E18 wounds exhibited increased collagen deposition in a disorganized pattern. This study demonstrates that the pattern of gene expression for types 1 and 3 collagen varies between mid- and late-gestational mouse excisional wounds. These alterations in procollagen expression may contribute to a pattern of collagen deposition in the mid-gestational fetuses that is more favorable for scarless healing with less type 1 and more type 3 collagen.

Electrostatic persistence length.

PubMed

Fixman, Marshall

2010-03-11

The persistence length is calculated for polyelectrolyte chains with fixed bond lengths and bond angles (pi-theta), and a potential energy consisting of the screened Coulomb interaction between beads, potential wells alpha phi(i)2 for the dihedral angles phi(i), and coupling terms beta phi(i) phi(i+/-1). This model defines a librating chain that reduces in appropriate limits to the freely rotating or wormlike chains, it can accommodate local crumpling or extreme stiffness, and it is easy to simulate. A planar-quadratic (pq), analytic approximation is based on an expansion of the electrostatic energy in eigenfunctions of the quadratic form that describes the backbone energy, and on the assumption that the quadratic form not only is positive but also adequately confines the chain in an infinite phase space of dihedral angles to the physically unique part with all |phi(i)| < pi. The pq approximation is available under these weak constraints, but the simulations confirm its quantitative accuracy only under the expected condition that alpha is large, that is, for very stiff chains. Stiff chains can also be simulated with small alpha and small theta and compared to an OSF approximation suitably generalized to chains with finite rather than vanishing theta, and increasing agreement with OSF is found the smaller is theta. The two approximations, one becoming exact as alpha --> infinity with fixed theta, the other as theta --> 0 with fixed alpha, are quantitatively similar in behavior, both giving a persistence length P = P0 + aD2 for stiff chains, where D is the Debye length. However, the coefficient apq is about twice the value of aOSF. Under other conditions the simulations show that P may or not be linear in D2 at small or moderate D, depending on the magnitudes of alpha, beta, theta, and the charge density but always becomes linear at large D. Even at a moderately low charge density, corresponding to fewer than 20% of the beads being charged, and with strong crumpling induced by large beta, increasing D dissolves blobs and recovers a linear dependence of P on D2, although a lower power of D gives an adequate fit at moderate D. For the class of models considered, it is concluded that the only universal feature is the asymptotic linearity of P in D2, regardless of flexibility or stiffness.

Enzymatic preparation of. cap alpha. - and. beta. -deuterated or tritiated amino acids with l-methionine. gamma. -lyase

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

Esaki, N.; Sawada, S.; Tanaka, H.

L-Methionine ..gamma..-lyase catalyzes the exchange of ..cap alpha..- and ..beta..-hydrogens of L-methionine and S-methyl-L-cysteine with deuterium or tritium of solvents. The rate of ..cap alpha..-hydrogen exchange with deuterium was about 40 times faster than that of the elimination reactions. The deuterium and tritium were exchanged also with the ..cap alpha..- and ..beta..-hydrogens of the straight-chain amino acids which do not undergo the elimination: L-alanine, L-..cap alpha..-aminobutyrate, L-norvaline, and L-norleucine. No exchange occurs for the D-isomers, acidic L-amino acids, basic L-amino acids, and branched-chain L-amino acids, although ..cap alpha..-hydrogen of glycine, L-trypotophan, and L-phenylalanine is exchanged slowly. These enzymatic hydrogen-exchange reactionsmore » facilitate specific labeling of the L-amino acids with deuterium and tritium.« less

Transforming growth factor-{beta} inhibits CCAAT/enhancer-binding protein expression and PPAR{gamma} activity in unloaded bone marrow stromal cells

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

Ahdjoudj, S.; Kaabeche, K.; Holy, X.

2005-02-01

The molecular mechanisms regulating the adipogenic differentiation of bone marrow stromal cells in vivo remain largely unknown. In this study, we investigated the regulatory effects of transforming growth factor beta-2 (TGF-{beta}2) on transcription factors involved in adipogenic differentiation induced by hind limb suspension in rat bone marrow stromal cells in vivo. Time course real-time quantitative reverse-transcription polymerase chain reaction (RT-PCR) analysis of gene expression showed that skeletal unloading progressively increases the expression of CCAAT/enhancer-binding protein (C/EBP){alpha} and C/EBP{beta} {alpha} at 5 days in bone marrow stromal cells resulting in increased peroxisome proliferator-activated receptor {gamma} (PPAR{gamma}2) transcripts at 7 days. TGF-{beta}2more » administration in unloaded rats corrected the rise in C/EBP{alpha} and C/EBP{beta} transcripts induced by unloading in bone marrow stromal cells. This resulted in inhibition of PPAR{gamma}2 expression that was associated with increased Runx2 expression. Additionally, the inhibition of C/EBP{alpha} and C/EBP{beta} expression by TGF-{beta}2 was associated with increased PPAR{gamma} serine phosphorylation in bone marrow stromal cells, a mechanism that inhibits PPAR{gamma} transactivating activity. The sequential inhibitory effect of TGF-{beta}2 on C/EBP{alpha}, C/EBP{beta}, and PPAR{gamma}2 resulted in reduced LPL expression and abolition of bone marrow stromal cell adipogenic differentiation, which contributed to prevent bone loss induced by skeletal unloading. We conclude that TGF-{beta}2 inhibits the excessive adipogenic differentiation of bone marrow stromal cells induced by skeletal unloading by inhibiting C/EBP{alpha}, C/EBP{beta}, and PPAR{gamma} expression and activity, which provides a sequential mechanism by which TGF-{beta}2 regulates adipogenic differentiation of bone marrow stromal cells in vivo.« less

{sup 110,116}Cd({alpha},{alpha}){sup 110,116}Cd elastic scattering and systematic investigation of elastic {alpha} scattering cross sections along the Z=48 isotopic and N=62 isotonic chains

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

Kiss, G. G.; Fueloep, Zs.; Gyuerky, Gy.

2011-06-15

The elastic scattering cross sections for the reactions {sup 110,116}Cd({alpha},{alpha}){sup 110,116}Cd at energies above and below the Coulomb barrier are presented to provide a sensitive test for the {alpha}-nucleus optical potential parameter sets. Additional constraints for the optical potential are taken from the analysis of elastic scattering excitation functions at backward angles which are available in literature. Moreover, the variation of the elastic {alpha} scattering cross sections along the Z=48 isotopic and N=62 isotonic chain is investigated by the study of the ratios of the {sup 106,110,116}Cd({alpha},{alpha}){sup 106,110,116}Cd scattering cross sections at E{sub cm{approx_equal}}15.6and18.8 MeV and the ratio of themore » {sup 110}Cd({alpha},{alpha}){sup 110}Cd and {sup 112}Sn({alpha},{alpha}){sup 112}Sn reaction cross sections at E{sub cm{approx_equal}}18.8 MeV, respectively. These ratios are sensitive probes for the {alpha}-nucleus optical potential parametrizations. The potentials under study are a basic prerequisite for the prediction of {alpha}-induced reaction cross sections (e.g., for the calculation of stellar reaction rates in the astrophysical p or {gamma} process).« less

Processing of MOPC 315 immunoglobulin A oligosaccharides: evidence for endoplasmic reticulum and trans Golgi alpha 1,2-mannosidase activity

PubMed Central

1984-01-01

The processing of asparagine-linked oligosaccharides on the alpha- chains of an immunoglobulin A (IgA) has been investigated using MOPC 315 murine plasmacytoma cells. These cells secrete IgA containing complex-type oligosaccharides that were not sensitive to endo-beta-N- acetylglucosaminidase H. In contrast, oligosaccharides present on the intracellular alpha-chain precursor were of the high mannose-type, remaining sensitive to endo-beta-N-acetylglucosaminidase H despite a long intracellular half-life of 2-3 h. The major [3H]mannose-labeled alpha-chain oligosaccharides identified after a 20-min pulse were Man8GlcNAc2 and Man9GlcNAc2. Following chase incubations, the major oligosaccharide accumulating intracellularly was Man6GlcNAc2, which was shown to contain a single alpha 1,2-linked mannose residue. Conversion of Man6GlcNAc2 to complex-type oligosaccharides occurred at the time of secretion since appreciable amounts of Man5GlcNAc2 or further processed structures could not be detected intracellularly. The subcellular locations of the alpha 1,2-mannosidase activities were studied using carbonyl cyanide m-chlorophenylhydrazone and monensin. Despite inhibiting the secretion of IgA, these inhibitors of protein migration did not effect the initial processing of Man9GlcNAc2 to Man6GlcNAc2. Furthermore, no large accumulation of Man5GlcNAc2 occurred, indicating the presence of two subcellular locations of alpha 1,2-mannosidase activity involved in oligosaccharide processing in MOPC 315 cells. Thus, the first three alpha 1,2-linked mannose residues were removed shortly after the alpha-chain was glycosylated, most likely in rough endoplasmic reticulum, since this processing occurred in the presence of carbonyl cyanide m-chlorophenylhydrazone. However, the removal of the final alpha 1,2-linked mannose residue as well as subsequent carbohydrate processing occurred just before IgA secretion, most likely in the trans Golgi complex since processing of Man6GlcNAc2 to Man5GlcNAc2 was greatly inhibited in the presence of monensin. PMID:6420419

Processing of MOPC 315 immunoglobulin A oligosaccharides: evidence for endoplasmic reticulum and trans Golgi alpha 1,2-mannosidase activity.

PubMed

Hickman, S; Theodorakis, J L; Greco, J M; Brown, P H

1984-02-01

The processing of asparagine-linked oligosaccharides on the alpha-chains of an immunoglobulin A (IgA) has been investigated using MOPC 315 murine plasmacytoma cells. These cells secrete IgA containing complex-type oligosaccharides that were not sensitive to endo-beta-N-acetylglucosaminidase H. In contrast, oligosaccharides present on the intracellular alpha-chain precursor were of the high mannose-type, remaining sensitive to endo-beta-N-acetylglucosaminidase H despite a long intracellular half-life of 2-3 h. The major [3H]mannose-labeled alpha-chain oligosaccharides identified after a 20-min pulse were Man8GlcNAc2 and Man9GlcNAc2. Following chase incubations, the major oligosaccharide accumulating intracellularly was Man6GlcNAc2, which was shown to contain a single alpha 1,2-linked mannose residue. Conversion of Man6GlcNAc2 to complex-type oligosaccharides occurred at the time of secretion since appreciable amounts of Man5GlcNAc2 or further processed structures could not be detected intracellularly. The subcellular locations of the alpha 1,2-mannosidase activities were studied using carbonyl cyanide m-chlorophenylhydrazone and monensin. Despite inhibiting the secretion of IgA, these inhibitors of protein migration did not effect the initial processing of Man9GlcNAc2 to Man6GlcNAc2. Furthermore, no large accumulation of Man5GlcNAc2 occurred, indicating the presence of two subcellular locations of alpha 1,2-mannosidase activity involved in oligosaccharide processing in MOPC 315 cells. Thus, the first three alpha 1,2-linked mannose residues were removed shortly after the alpha-chain was glycosylated, most likely in rough endoplasmic reticulum, since this processing occurred in the presence of carbonyl cyanide m-chlorophenylhydrazone. However, the removal of the final alpha 1,2-linked mannose residue as well as subsequent carbohydrate processing occurred just before IgA secretion, most likely in the trans Golgi complex since processing of Man6GlcNAc2 to Man5GlcNAc2 was greatly inhibited in the presence of monensin.

Solubilization of cyclohexane in aqueous solutions of sodium. cap alpha. -alkyl alkanoates

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

Sagitani, H.; Suzuki, T.; Nagai, M.

1982-01-01

The effect of branched alkyl chain length and the position of the COONa group on the solubilizing power of n-alkane sodium carboxylates was studied. The lipophilic property and the amount of solubilized cyclohexane increased with the branched chain length of branched soaps, and with the change of the position of the -COONa group from 3 to 7 in the alkyl chain of pentadecane -3, -5, and -7 sodium carboxylates. Alpha-branched soaps having proper branched alkyl chains were better solubilizers for cyclohexane than straight chain compounds. The amount of cyclohexane solublized by C/sub 10/ H/sub 21/ CH(C/sub 6/H/sub 13/) COONa wasmore » about three times greater than the amount solubilized by C/sub 17/ H/sub 35/ COONa. There was a marked increase in the solubilization of cyclohexane replacing ..cap alpha..-branched fatty acid soaps with optimum amount of cosurfactants such as C/sub 8/H/sub 17/ (OCH/sub 2/CH/sub 2/)/sub 2/OH. Namely, solubilization increased markedly at the optimum hydrophile-lipophile balance of mixed surfactant. 21 references.« less

Characterization of a possible amyloidogenic precursor in glutamine-repeat neurodegenerative diseases.

PubMed

Armen, Roger S; Bernard, Brady M; Day, Ryan; Alonso, Darwin O V; Daggett, Valerie

2005-09-20

Several neurodegenerative diseases are linked to expanded repeats of glutamine residues, which lead to the formation of amyloid fibrils and neuronal death. The length of the repeats correlates with the onset of Huntington's disease, such that healthy individuals have <38 residues and individuals with >38 repeats exhibit symptoms. Because it is difficult to obtain atomic-resolution structural information for poly(l-glutamine) (polyQ) in aqueous solution experimentally, we performed molecular dynamics simulations to investigate the conformational behavior of this homopolymer. In simulations of 20-, 40-, and 80-mer polyQ, we observed the formation of the "alpha-extended chain" conformation, which is characterized by alternating residues in the alpha(L) and alpha(R) conformations to yield a sheet. The structural transition from disordered random-coil conformations to the alpha-extended chain conformation exhibits modest length and temperature dependence, in agreement with the experimental observation that aggregation depends on length and temperature. We propose that fibril formation in polyQ may occur through an alpha-sheet structure, which was proposed by Pauling and Corey. Also, we propose an atomic-resolution model of how the inhibitory peptide QBP1 (polyQ-binding peptide 1) may bind to polyQ in an alpha-extended chain conformation to inhibit fibril formation.

Gastric form of alpha chain disease.

PubMed Central

Coulbois, J; Galian, P; Galian, A; Couteaux, B; Danon, F; Rambaud, J

1986-01-01

A case of alpha chain disease, involving the stomach only, is reported in an Algerian man suffering from epigastric pains. Upper digestive tract fibreoptic endoscopy showed two antral ulcers and an ulcerative gastritis pattern, which promptly disappeared with cimetidine treatment. Antral biopsies at a distance from the ulcers, but not of the ulcer crater itself, disclosed a dense infiltration of antral lamina propria by mature or sometimes atypical plasma cells. On transmural surgical antral biopsy, the infiltrate spread to the superficial part of the submucosa. No other localisation of the disease was found in spite of multiple biopsies obtained by endoscopy, with a peroral capsule and during staging laparotomy. The alpha chain disease protein was absent from serum and urine, but found in the gastric juice and in the cytoplasma of the cellular infiltrate (alpha 1 subclass). A complete clinical, endoscopic, histological and immunological remission was observed after a six months' course of oral tetracycline. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 Fig. 5 Fig. 6 PMID:3087826

Three-dimensional structure of holo 3 alpha,20 beta-hydroxysteroid dehydrogenase: a member of a short-chain dehydrogenase family.

PubMed Central

Ghosh, D; Weeks, C M; Grochulski, P; Duax, W L; Erman, M; Rimsay, R L; Orr, J C

1991-01-01

The x-ray structure of a short-chain dehydrogenase, the bacterial holo 3 alpha,20 beta-hydroxysteroid dehydrogenase (EC 1.1.1.53), is described at 2.6 A resolution. This enzyme is active as a tetramer and crystallizes with four identical subunits in the asymmetric unit. It has the alpha/beta fold characteristic of the dinucleotide binding region. The fold of the rest of the subunit, the quaternary structure, and the nature of the cofactor-enzyme interactions are, however, significantly different from those observed in the long-chain dehydrogenases. The architecture of the postulated active site is consistent with the observed stereospecificity of the enzyme and the fact that the tetramer is the active form. There is only one cofactor and one substrate-binding site per subunit; the specificity for both 3 alpha- and 20 beta-ends of the steroid results from the binding of the steroid in two orientations near the same cofactor at the same catalytic site. Images PMID:1946424

Synthesis and studies of polypeptide materials: Enantioselective polymerization of gamma-benzyl glutamate-N-carboxyanhydride and synthesis of optically active poly(beta-peptides)

NASA Astrophysics Data System (ADS)

Cheng, Jianjun

A class of zero-valent transition metal complexes have been developed by Deming et al for the controlled polymerization of alpha-aminoacid-N-carboxyanhydrides (alpha-NCAs). This discovery provided a superior starting point for the development of enantioselective polymerizations of racemic alpha-NCAs. Bidentate chiral ligands were synthesized and tested for their abilities to induce enantioselective polymerization of gamma-benzyl-glutamate NCA (Glu NCA) when they were coordinated to zero-valent nickel complexes. When optically active 2-pyridinyl oxazoline ligands were mixed with bis(1,5-cyclooctadiene)nickel in THF, chiral nickel complexes were formed that selectively polymerized one enantiomer of Glu NCA over the other. The highest selectivity was observed with the nickel complex of (S)-4-tert-butyl-2-pyridinyl oxazoline, which gave a ratio of enantiomeric polymerization rate constants (kD/kL) of 5.2. It was found that subtle modification of this ligand by incorporation of additional substituents had a substantial impact on initiator enantioselectivities. In separate efforts, methodology was developed for the general synthesis of optically active beta-aminoacid-N-carboxyanhydrides (beta-NCAs) via cyclization of Nbeta-Boc- or Nbeta-Cbz-beta-amino acids using phosphorus tribromide. The beta-NCA molecules could be polymerized in good yields using strong bases or transition metal complexes to give optically active poly(beta-peptides) bearing proteinogenic side chains. The resulting poly(beta-peptides), which have moderate molecular weights, adopt stable helical conformations in solution. Poly(beta-homoglutamate and poly(beta-homolysine), the side-chain deprotected polymers, were found to display pH dependent helix-coil conformation transitions in aqueous solution, similar to their alpha-analogs. A novel method for poly(beta-aspartate) synthesis was developed via the polymerization of L-aspartate alkyl ester beta lactams using metal-amido complexes. Poly(beta-aspartates) bearing short ethylene glycol side chains were obtained with controlled molecular weights and narrow molecular weight distributions when Sc(N(TMS)2)3 was used as initiator for the beta-lactam polymerizations. Polymer chain lengths could be controlled by both stoichiometry and monomer conversion, characteristic of a living polymerization system. Di- and tri-block copoly(beta-peptides) with desired chain lengths were also synthesized using this method. It was found that these techniques were generally applicable for the synthesis of poly(beta-peptides), bearing other proteinogetic side chains. Synthesis and studies of polypeptide materials were extended to unexplored areas by incorporation of both alpha- and beta-amino acid residues into single polymer chains. Two sequence specific polypeptides bearing alternating beta-alpha, or beta-alpha-alpha amino acid residues were synthesized. Both polymers were found to adopt unprecedented stable conformations in solution.

Structural investigation of C4b-binding protein by molecular modeling: localization of putative binding sites.

PubMed

Villoutreix, B O; Härdig, Y; Wallqvist, A; Covell, D G; García de Frutos, P; Dahlbäck, B

1998-06-01

C4b-binding protein (C4BP) contributes to the regulation of the classical pathway of the complement system and plays an important role in blood coagulation. The main human C4BP isoform is composed of one beta-chain and seven alpha-chains essentially built from three and eight complement control protein (CCP) modules, respectively, followed by a nonrepeat carboxy-terminal region involved in polymerization of the chains. C4BP is known to interact with heparin, C4b, complement factor I, serum amyloid P component, streptococcal Arp and Sir proteins, and factor VIII/VIIIa via its alpha-chains and with protein S through its beta-chain. The principal aim of the present study was to localize regions of C4BP involved in the interaction with C4b, Arp, and heparin. For this purpose, a computer model of the 8 CCP modules of C4BP alpha-chain was constructed, taking into account data from previous electron microscopy (EM) studies. This structure was investigated in the context of known and/or new experimental data. Analysis of the alpha-chain model, together with monoclonal antibody studies and heparin binding experiments, suggests that a patch of positively charged residues, at the interface between the first and second CCP modules, plays an important role in the interaction between C4BP and C4b/Arp/Sir/heparin. Putative binding sites, secondary-structure prediction for the central core, and an overall reevaluation of the size of the C4BP molecule are also presented. An understanding of these intermolecular interactions should contribute to the rational design of potential therapeutic agents aiming at interfering specifically some of these protein-protein interactions.

Characterization of a novel debranching enzyme from Nostoc punctiforme possessing a high specificity for long branched chains

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

Choi, Ji-Hye; Lee, Heeseob; Kim, Young-Wan

2009-01-09

A novel debranching enzyme from Nostoc punctiforme PCC73102 (NPDE) exhibits hydrolysis activity toward both {alpha}-(1,6)- and {alpha}-(1,4)-glucosidic linkages. The action patterns of NPDE revealed that branched chains are released first, and the resulting maltooligosaccharides are then hydrolyzed. Analysis of the reaction with maltooligosaccharide substrates labeled with {sup 14}C-glucose at the reducing end shows that NPDE specifically liberates glucose from the reducing end. Kinetic analyses showed that the hydrolytic activity of NPDE is greatly affected by the length of the substrate. The catalytic efficiency of NPDE increased considerably upon using substrates that can occupy at least eight glycone subsites such asmore » maltononaose and maltooctaosyl-{alpha}-(1,6)-{beta}-cyclodextrin. These results imply that NPDE has a unique subsite structure consisting of -8 to +1 subsites. Given its unique subsite structure, side chains shorter than maltooctaose in amylopectin were resistant to hydrolysis by NPDE, and the population of longer side chains was reduced.« less

T cell receptor (TCR) structure of autologous melanoma-reactive cytotoxic T lymphocyte (CTL) clones: tumor-infiltrating lymphocytes overexpress in vivo the TCR beta chain sequence used by an HLA-A2- restricted and melanocyte-lineage-specific CTL clone

PubMed Central

1993-01-01

HLA-A2+ melanomas express common melanoma-associated antigens (Ags) recognized in vitro by autologous cytotoxic T lymphocytes (CTL). However, it is not known whether tumor Ags can drive in vivo a selective accumulation/expansion of Ag-specific, tumor-infiltrating T lymphocytes (TIL). Therefore, to evaluate this possibility, 39 CTL clones isolated from several independent mixed lymphocyte tumor cultures (MLTC) of TIL and peripheral blood lymphocytes (PBL) of an HLA- A2+ melanoma patient and selected for T cell receptor (TCR)-dependent, HLA-restricted tumor lysis, were used for analysis of TCR alpha and beta chain structure by the cDNA polymerase chain reaction (PCR) technique with variable gene-specific primers followed by sequencing. Despite absence of oligoclonality in fresh TIL and PBL, as well as in T cells of day 28 MLTC (day of cloning), sequence analysis of TCR alpha and beta chains of TIL clones revealed a dominance of a major category of melanoma-specific, HLA-A2-restricted T cells expressing a V alpha 8.2/J alpha AP511/C alpha and V beta 2.1/D beta 1/J beta 1.1/C beta 1 TCR. The same TCR was also found in 2 out of 14 PBL clones. The other PBL clones employed a V alpha 2.1 gene segment associated with either V beta 13.2, 14, or w22. Clones A81 (V alpha 2.1/J alpha IGRJ alpha 04/C alpha and V beta 14/D beta 1/J beta 1.2/C beta 1) and A21 (V alpha 8.2/J alpha AP511/C alpha and V beta 2.1/D beta 1/J beta 1.1/C beta 1), representative of the two most frequent TCR of PBL and TIL, respectively, expressed different lytic patterns, but both were HLA-A2 restricted and lysed only HLA-A2+ melanomas and normal melanocytes, thus indicating recognition of two distinct HLA-A2-associated and tissue-related Ags. Finally, by the inverse PCR technique, the specific TCR beta chain (V beta 2.1/D beta 1/J beta 1.1/C beta 1) expressed by the dominant TIL clone was found to represent 19 and 18.4% of all V beta 2 sequences expressed in the fresh tumor sample and in the purified TIL, respectively, but < 0.19% of V beta 2+ sequences expressed in PBL. These results are consistent with the hypothesis that a clonal expansion/accumulation of a melanocyte-lineage-specific and HLA-A2-restricted T cell clone occurred in vivo at the site of tumor growth. PMID:8376931

Identification of the antigenic determinants of factors 8, 9, and 34 of genus Candida.

PubMed

Kobayashi, H; Oyamada, H; Suzuki, A; Shibata, N; Suzuki, S; Okawa, Y

1996-10-21

We investigated the antigenic determinants of factors 8, 9, and 34 of the genus Candida among pathogenic yeasts by enzyme-linked immunosorbent assay (ELISA) using mannans of Saccharomyces cerevisiae wild type and mutant types, mnn 1-mnn 4 and mnn 2. Results of ELISA including antisera against the antigenic factors of genus Candida (Candida Check, latron; FAbs) indicated that these three types of mannan distinctly react with FAbs 34, 8 and 9, respectively. To identify the recognition sites of these FAbs, we compared the ability of various oligosaccharides to inhibit the binding of the mannans to FAbs. The results indicated that FAb 34 preferentially recognizes linear side chains containing a non-reducing terminal alpha-1,3-linked mannose residue, Man(alpha)1 --> 3Man(alpha)1 --> (2Man(alpha)1 --> )n(2Man) (n > or = 0), and that one of the recognition sites of FAb 9 is linear alpha-1,6-linked oligomannosyl series, Man(alpha)1 --> (6Man(alpha)1 --> )n(6Man) (n > or = 2). On the other hand, the recognition site of FAb 8 apparently consisted of two alpha-1,2-linked oligomannosyl side chains and an alpha-1,6-linked mannose residue that originated from the mannan backbone, Man(alpha)1 --> 2Man(alpha)1 --> 2(Man(alpha)1 -->2Man(alpha)1 --> 6)Man.

Alpha-galactosidase versus active charcoal for improving sonographic visualization of abdominal organs in patients with excessive intestinal gas

PubMed Central

Maconi, G.; Bolzacchini, E.; Radice, E.; Marzocchi, M.; Badini, M.

2012-01-01

Background and aims Intestinal gas is a frequent cause of poor visualization during gastrointestinal ultrasound (US). The enzyme alpha-galactosidase may reduce intestinal gas production, thereby improving abdominal US visualization. We compared the efficacies of alpha-galactosidase and active charcoal in improving US visualization in patients with previous unsatisfactory abdominal US scans caused by excessive intestinal gas. Materials and methods: 45 patients with poor visualization of at least one target organ: pancreas, hepatic lobes (score 0–2) or common bile duct (CBD) (score 0–1) were enrolled in a prospective randomized, crossover, observer-blinded study. The patients received alpha-galactosidase (Sinaire Forte, Promefarm, Milan, Italy) 600 GalU t.i.d. for 2 days before abdominal US plus 900 GalU the morning of exam or active charcoal 448 mg t.i.d., for 2 days before the exam plus 672 mg the morning of the exam. Visualization was graded as follows: 0 = none (complete gas interference); 1 = severe interference, 2 = moderate interference, 3 = mild interference; 4 = complete (no gas interference). Results: 42 patients completed the study. Both alpha-galactosidase and active charcoal improved the visualization of target organs. Visualization of the right hepatic lobe, CBD and pancreatic tail was significantly improved (vs. baseline) only by alpha-galactosidase (p < 0.01). Scores ≥3 for all parts of the pancreas and both hepatic lobes were achieved in only 12.5% of the patients after both treatments. Both products were well tolerated. Conclusion: Alpha-galactosidase and active charcoal can improve US visualization of abdominal organs in patients whose scans are frequently unsatisfactory due to excessive intestinal gas. Visualization of the pancreatic tail and right hepatic lobe was significantly improved only by alpha-galactosidase. However, both treatments allowed adequate visualization of all target organs during the same examination only in a few patients. PMID:23730387

Alpha chain hemoglobins with electrophoretic mobility similar to that of hemoglobin S in a newborn screening program.

PubMed

Silva, Marcilene Rezende; Sendin, Shimene Mascarenhas; Araujo, Isabela Couto de Oliveira; Pimentel, Fernanda Silva; Viana, Marcos Borato

2013-01-01

To characterize alpha-chain variant hemoglobins with electric mobility similar to that of hemoglobin S in a newborn screening program. β(S) allele and alpha-thalassemia deletions were investigated in 14 children who had undefined hemoglobin at birth and an electrophoretic profile similar to that of hemoglobin S when they were six months old. Gene sequencing and restriction enzymes (DdeI, BsaJI, NlaIV, Bsu36I and TaqI) were used to identify hemoglobins. Clinical and hematological data were obtained from children who attended scheduled medical visits. THE FOLLOWING ALPHA CHAIN VARIANTS WERE FOUND: seven children with hemoglobin Hasharon [alpha2 47(CE5) Asp>His, HbA2:c.142G>C], all associated with alpha-thalassemia, five with hemoglobin Ottawa [alpha1 15(A13) Gly>Arg, HBA1:c.46G>C], one with hemoglobin St Luke's [alpha1 95(G2) Pro>Arg, HBA1:c.287C>G] and another one with hemoglobin Etobicoke [alpha212 84(F5) Ser>Arg, HBA212:c.255C>G]. Two associations with hemoglobin S were found: one with hemoglobin Ottawa and one with hemoglobin St Luke's. The mutation underlying hemoglobin Etobicoke was located in a hybrid α212 allele in one child. There was no evidence of clinically relevant hemoglobins detected in this study. Apparently these are the first cases of hemoglobin Ottawa, St Luke's, Etobicoke and the α212 gene described in Brazil. The hemoglobins detected in this study may lead to false diagnosis of sickle cell trait or sickle cell disease when only isoelectric focusing is used in neonatal screening. Additional tests are necessary for the correct identification of hemoglobin variants.

Epitope mapping of the alpha-chain of the insulin-like growth factor I receptor using antipeptide antibodies.

PubMed

Delafontaine, P; Ku, L; Ververis, J J; Cohen, C; Runge, M S; Alexander, R W

1994-12-01

Insulin-like growth factor I (IGF I) is an important mitogen for vascular smooth muscle cells (VSMC). The IGF I receptor (IGF IR) is a heterotetramer composed of two cross-linked extracellular alpha-chains and two membrane-spanning beta-chains that contain a tyrosine-kinase domain. It has a high degree of sequence similarity to the insulin receptor (IR), and the putative ligand-specific binding site has been localized to a cysteine-rich region (CRR) of the alpha-chain. To obtain insights into antigenic determinants of the IGF IR, we raised a panel of site-specific polyclonal antibodies against short peptide sequences N-terminal to and within the CRR. Several antibodies raised against linear epitopes within the CRR bound to solubilized and native rat and human IGF IR by ELISA, did not cross-react with IR, but unexpectedly failed to inhibit 125I-IGF I binding. A polyclonal antibody directed against a 48-amino acid synthetic peptide, corresponding to a region of the CRR postulated to be essential for ligand binding, failed to react with either solubilized, reduced or intact IGF IR. Three antibodies specific for the N-terminus of the alpha-chain reacted with solubilized and native IGF IR. One of these, RAB 6, directed against amino acids 38-44 of the IGF IR, inhibited 125I-IGF I binding to rat aortic smooth muscle cells (RASM) and to IGF IR/3T3 cells (overexpressing human IGF IR) by up to 45%. Immunohistochemical analysis revealed strong IGF IR staining in the medial smooth muscle cell layer of rat aorta. These findings are consistent with a model wherein conformational epitopes within the CRR and linear epitopes within the N-terminus of the alpha-chain contribute to the IGF I binding pocket. These antibodies should provide a valuable tool to study structure-function relationships and in vivo regulation of the IGF IR.

Effect of interaction range on phonon relaxation in Fermi-Pasta-Ulam beta chain.

PubMed

Santhosh, G; Kumar, Deepak

2007-08-01

We study the effect of increasing the range of interactions on phonon relaxation in a chain of atoms with quartic anharmonicity. The study is motivated by recent numerical studies, showing that the value of the exponent alpha characterizing the divergence of conductivity with system size apparently depends on the presence of second neighbor couplings. We perform a quantum calculation of the wave-vector (q) dependent relaxation rate gamma(q) in the second order perturbation theory. The nonanalytic dependence of gamma(q) arises due to small-q singularity of the collision integral. We find that gamma(q) proportional to Aq(5/3) + Bq2. This gives rise to an asymptotic value alpha = 0.4, but the q2 terms lead to a higher apparent value of alpha at small sizes of the chain.

Sera of IgA nephropathy patients contain a heterogeneous population of relatively cationic alpha-heavy chains.

PubMed

Shuib, A S; Chua, C T; Hashim, O H

1998-01-01

Sera of IgA nephropathy (IgAN) patients and normal subjects were analysed by two-dimensional (2-D) gel electrophoresis. Densitometric analysis of the 2-D gels of IgAN patients and normal subjects revealed that their protein maps were comparable. There was no shift of pI values in the major alpha-heavy chain spots. However, the volume of the alpha-heavy chain bands were differently distributed. Distribution was significantly lower at the anionic region in IgAN patients (mean anionic:cationic ratio of 1.184 +/- 0.311) as compared to normal healthy controls (mean anionic:cationic ratio of 2.139 +/- 0.538). Our data are in support of the previously reported findings that IgA1 of IgAN patients were lacking in sialic acid residues.

alpha-Crystallin A sequences of Alligator mississippiensis and the lizard Tupinambis teguixin: molecular evolution and reptilian phylogeny.

PubMed

de Jong, W W; Zweers, A; Versteeg, M; Dessauer, H C; Goodman, M

1985-11-01

The amino acid sequences of the eye lens protein alpha-crystallin A from many mammalian and avian species, two frog species, and a dogfish have provided detailed information about the molecular evolution of this protein and allowed some useful inferences about phylogenetic relationships among these species. We now have isolated and sequenced the alpha-crystallins of the American alligator and the common tegu lizard. The reptilian alpha A chains appear to have evolved as slowly as those of other vertebrates, i.e., at two to three amino acid replacements per 100 residues in 100 Myr. The lack of charged replacements and the general types and distribution of replacements also are similar to those in other vertebrate alpha A chains. Maximum-parsimony analyses of the total data set of 67 vertebrate alpha A sequences support the monophyletic origin of alligator, tegu, and birds and favor the grouping of crocodilians and birds as surviving sister groups in the subclass Archosauria.

FcepsilonRI-alpha siRNA inhibits the antigen-induced activation of mast cells.

PubMed

Safaralizadeh, Reza; Soheili, Zahra-Soheila; Deezagi, Abdolkhaleg; Pourpak, Zahra; Samiei, Shahram; Moin, Mostafa

2009-12-01

FcepsilonRI, The high affinity receptor for IgE plays a critical role in triggering the allergic reactions. It is responsible for inducing mast cell degranulation and deliberation of allergy mediators when it is aggregated by allergen and IgE complexes. FcepsilonRI on the mast cells consists of three subunits; alpha chain directly binds IgE, beta chain and dimmer of gamma chains together mediate intracellular signaling. Cross-linking of IgE-bound FcepsilonRI on the surface of mast cells and basophils by the multivalent antigen induces release of chemical mediators. The present in vitro study was designed to investigate the effect of synthetic FcepsilonRI-alpha siRNA on the antigen-induced activation of MC/9 cells. MC/9 cells which are murine mast cells were transfected by FcepsilonRI-alpha siRNA and negative control siRNA. After 6 h, anti-DNP (Dinitrophenyl) IgE was used for the cells sensitization. Then the cells were challenged with Dinitrophenyl-Human Serum Albumin (DNP-HSA) for mast cell degranulation induction before collection of supernatants. The amount of mRNA and protein expression was measured by Real Time PCR and western blot analysis, respectively. Determination of the expression rate of FcepsilonRI-alpha on cell surface was achieved by flow cytometry. ELISA and spectrophotometry methods were used subsequently for measuring the effects of FcepsilonRI-alpha siRNA on antigen-induced histamine and beta-hexosaminidase release. FcepsilonRI-alpha siRNA treated cells showed significant decrease in FcepsilonRI-alpha mRNA and protein expression in comparison to control cells. FcepsilonRI-mediated mast cell release of beta-hexosaminidase and histamine were also inhibited. In this study it was shown that FcepsilonRI-alpha siRNA could suppress FcepsilonRI-alpha expression and inhibited degranulation and histamine release in antigen-stimulated MC/9 cells. In conclusion, knock-down of FcepsilonRI-alpha by siRNA could be a promising method for inhibition of the mast cell-mediated allergic reactions.

How main-chains of proteins explore the free-energy landscape in native states.

PubMed

Senet, Patrick; Maisuradze, Gia G; Foulie, Colette; Delarue, Patrice; Scheraga, Harold A

2008-12-16

Understanding how a single native protein diffuses on its free-energy landscape is essential to understand protein kinetics and function. The dynamics of a protein is complex, with multiple relaxation times reflecting a hierarchical free-energy landscape. Using all-atom molecular dynamics simulations of an alpha/beta protein (crambin) and a beta-sheet polypeptide (BS2) in their "native" states, we demonstrate that the mean-square displacement of dihedral angles, defined by 4 successive C(alpha) atoms, increases as a power law of time, t(alpha), with an exponent alpha between 0.08 and 0.39 (alpha = 1 corresponds to Brownian diffusion), at 300 K. Residues with low exponents are located mainly in well-defined secondary elements and adopt 1 conformational substate. Residues with high exponents are found in loops/turns and chain ends and exist in multiple conformational substates, i.e., they move on multiple-minima free-energy profiles.

Search for ternary fission of chromium-48

NASA Astrophysics Data System (ADS)

Dummer, Andrew K.

1999-07-01

Both alpha cluster model calculations and macroscopic energy calculations that allow for a double-neck shape of the compound nucleus suggest the possibility of a novel three 16O, chain-like configuration in 48 Cr. Such a configuration might lead to an enhanced cross section for three-16O breakup. To explore this possibility, the three-body exit channels for the 36Ar + 12C reaction at a beam energy of 210 MeV have been studied. The cross section for 16O + 16O + 16O breakup has been deduced and has been found to be in excess of what would be expected to result from a sequential binary fission process. However, the observation of a similarly enhanced 12C + 16O + 20Ne breakup cross section suggests that the observed 16O + 16O + 16O yields might still be associated with a statistical fission process. The results are discussed in the context of the fission of light nuclear systems and a simple cluster model calculation. This latter, ``Harvey model'' calculation suggests a possible inhibition of the formation of a three- 16O chain configuration from the 36Ar + 12C entrance channel. A further measurement using the 20Ne + 28Si-entrance channel is suggested.

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

Penning, T.D.

The enone, 2,2-diemthyl-3a..beta.., 6a..beta..-dihydro-4H-cyclopenta-1,3-dioxol-4-one, has been synthesized in six steps from cyclopentadiene, resolved using sulfoximine chemistry, and converted into (-)-prostaglandin E/sub 2/ methyl ester in three steps. Introduction of the optically pure omega side-chain using a conjugate addition of a stabilized organocopper reagent, followed by direct alkylation of the enolate with the ..cap alpha.. side-chain allylic iodide in the presence of hexamethylphosphoramide, afforded a trans, vicinally disubstituted cyclopentanone. Deprotection of the C-15 alcohol, followed by aluminum amalgam reduction of the C-10/oxygen bond, provided (-)-PGE/sub 2/ methyl ester in 47% overall yield from the enone. In an extension of previously describedmore » work, 2-chloro-3,4-dimethyl-5-phenyl-1,3,2-oxazaphospholidine 2-sulfide, prepared from l-ephedrine and thiophosphoryl chloride, was used to determine the enantiomeric excess of chiral alcohols in conjunction with /sup 31/P NMR. Chiral primary and secondary alcohols added quantitatively to the phospholidine to give diastereomers which could be analyzed by /sup 31/P NMR and HPLC. A number of other phosphorus heterocycles were also explored as potential chiral derivatizing reagents.« less

Muscular Dystrophy with Ribitol-Phosphate Deficiency: A Novel Post-Translational Mechanism in Dystroglycanopathy

PubMed Central

Kanagawa, Motoi; Toda, Tatsushi

2017-01-01

Muscular dystrophy is a group of genetic disorders characterized by progressive muscle weakness. In the early 2000s, a new classification of muscular dystrophy, dystroglycanopathy, was established. Dystroglycanopathy often associates with abnormalities in the central nervous system. Currently, at least eighteen genes have been identified that are responsible for dystroglycanopathy, and despite its genetic heterogeneity, its common biochemical feature is abnormal glycosylation of alpha-dystroglycan. Abnormal glycosylation of alpha-dystroglycan reduces its binding activities to ligand proteins, including laminins. In just the last few years, remarkable progress has been made in determining the sugar chain structures and gene functions associated with dystroglycanopathy. The normal sugar chain contains tandem structures of ribitol-phosphate, a pentose alcohol that was previously unknown in humans. The dystroglycanopathy genes fukutin, fukutin-related protein (FKRP), and isoprenoid synthase domain-containing protein (ISPD) encode essential enzymes for the synthesis of this structure: fukutin and FKRP transfer ribitol-phosphate onto sugar chains of alpha-dystroglycan, and ISPD synthesizes CDP-ribitol, a donor substrate for fukutin and FKRP. These findings resolved long-standing questions and established a disease subgroup that is ribitol-phosphate deficient, which describes a large population of dystroglycanopathy patients. Here, we review the history of dystroglycanopathy, the properties of the sugar chain structure of alpha-dystroglycan, dystroglycanopathy gene functions, and therapeutic strategies. PMID:29081423

Mechanism of insulin fibrillation: the structure of insulin under amyloidogenic conditions resembles a protein-folding intermediate.

PubMed

Hua, Qing-xin; Weiss, Michael A

2004-05-14

Insulin undergoes aggregation-coupled misfolding to form a cross-beta assembly. Such fibrillation has long complicated its manufacture and use in the therapy of diabetes mellitus. Of interest as a model for disease-associated amyloids, insulin fibrillation is proposed to occur via partial unfolding of a monomeric intermediate. Here, we describe the solution structure of human insulin under amyloidogenic conditions (pH 2.4 and 60 degrees C). Use of an enhanced sensitivity cryogenic probe at high magnetic field avoids onset of fibrillation during spectral acquisition. A novel partial fold is observed in which the N-terminal segments of the A- and B-chains detach from the core. Unfolding of the N-terminal alpha-helix of the A-chain exposes a hydrophobic surface formed by native-like packing of the remaining alpha-helices. The C-terminal segment of the B-chain, although not well ordered, remains tethered to this partial helical core. We propose that detachment of N-terminal segments makes possible aberrant protein-protein interactions in an amyloidogenic nucleus. Non-cooperative unfolding of the N-terminal A-chain alpha-helix resembles that observed in models of proinsulin folding intermediates and foreshadows the extensive alpha --> beta transition characteristic of mature fibrils.

Oligopeptides and copeptides of homochiral sequence, via beta-sheets, from mixtures of racemic alpha-amino acids, in a one-pot reaction in water; relevance to biochirogenesis.

PubMed

Illos, Roni A; Bisogno, Fabricio R; Clodic, Gilles; Bolbach, Gerard; Weissbuch, Isabelle; Lahav, Meir

2008-07-09

As part of our studies on the biochirogenesis of peptides of homochiral sequence during early evolution, the formation of oligopeptides composed of 14-24 residues of the same handedness in the polymerization of dl-leucine (Leu), dl-phenylalanine (Phe), and dl-valine (Val) in aqueous solutions, by activation with N, N'-carbonyldiimidazole and then initiation with a primary amine, in a one-pot reaction, was demonstrated by MALDI-TOF MS using deuterium enantio-labeled alpha-amino acids. The formation of long isotactic peptides is rationalized by the following steps occurring in tandem: (i) creation of a library of short diasteroisomeric oligopeptides containing isotactic peptides in excess in comparison to a binomial kinetics, as a result of an asymmetric induction exerted by the N-terminal residue of a given handedness; (ii) precipitation of the less soluble racemic isotactic penta- and hexapeptides in the form of beta-sheets that are delineated by homochiral rims; (iii) regio-enantiospecific chain elongation occurring heterogeneously at the beta-sheets/solution interface. Polymerization of l-Leu with l-isoleucine (Ile) or l-Phe with l- (1) N-Me-histidine yielded mixtures of copeptides containing both residues. In contrast, in the polymerization of the corresponding mixtures of l- + d-alpha-amino acids, the long oligopeptides were composed mainly from oligo- l-Leu and oligo- d-Ile in the first system and oligo- d-Phe in the second. Furthermore, in the polymerization of mixtures of hydrophobic racemic alpha-amino acids dl-Leu, dl-Val, and dl-Phe and with added racemic dl-alanine and dl-tyrosine, copeptides of homochiral sequences are most dominantly represented. Possible routes for a spontaneous "mirror-symmetry breaking" process of the racemic mixtures of homochiral peptides are presented.

Platelets from patients with the Quebec platelet disorder contain and secrete abnormal amounts of urokinase-type plasminogen activator.

PubMed

Kahr, W H; Zheng, S; Sheth, P M; Pai, M; Cowie, A; Bouchard, M; Podor, T J; Rivard, G E; Hayward, C P

2001-07-15

The Quebec platelet disorder (QPD) is an autosomal dominant platelet disorder associated with delayed bleeding and alpha-granule protein degradation. The degradation of alpha-granule, but not plasma, fibrinogen in patients with the QPD led to the investigation of their platelets for a protease defect. Unlike normal platelets, QPD platelets contained large amounts of fibrinolytic serine proteases that had properties of plasminogen activators. Western blot analysis, zymography, and immunodepletion experiments indicated this was because QPD platelets contained large amounts of urokinase-type plasminogen activator (u-PA) within a secretory compartment. u-PA antigen was not increased in all QPD plasmas, whereas it was increased more than 100-fold in QPD platelets (P <.00009), which contained increased u-PA messenger RNA. Although QPD platelets contained 2-fold more plasminogen activator inhibitor 1 (PAI-1) (P <.0008) and 100-fold greater u-PA-PAI-1 complexes (P <.0002) than normal platelets, they contained excess u-PA activity, predominantly in the form of two chain (tcu-PA), which required additional PAI-1 for full inhibition. There was associated proteolysis of plasminogen in QPD platelets, to forms that comigrated with plasmin. When similar amounts of tcu-PA were incubated with normal platelet secretory proteins, many alpha-granule proteins were proteolyzed to forms that resembled degraded QPD platelet proteins. These data implicate u-PA in the pathogenesis of alpha-granule protein degradation in the QPD. Although patients with the QPD have normal to increased u-PA levels in their plasma, without evidence of systemic fibrinogenolysis, their increased platelet u-PA could contribute to bleeding by accelerating fibrinolysis within the hemostatic plug. QPD is the only inherited bleeding disorder in humans known to be associated with increased u-PA.

Effect of chronic alcohol consumption on Hepatic SIRT1 and PGC-1{alpha} in rats

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

Lieber, Charles S.; Department of Medicine, Mount Sinai School of Medicine, New York, NY; Leo, Maria A.

2008-05-23

The nuclear genes, NAD-dependent deacetylase Sirtuis 1 (SIRT1) and the peroxisome proliferator-activated receptor-{gamma} coactivator1{alpha} (PGC-1{alpha}) are regulators of energy metabolism. Here, we studied the role of alcohol consumption in expression of these sensing molecules. Alcohol significantly reduced hepatic SIRT1 mRNA by 50% and PGC-1{alpha} mRNA by 46% and it significantly inhibited the protein expression of SIRT1 and PGC-1{alpha}, while the transcription factor PPAR-{gamma} remained unchanged. However, when the lipid composition of the alcohol diet was changed by replacing long-chain triglycerides (LCT) with medium chain triglycerides (MCT), SIRT1 and PGC-1{alpha} mRNA were restored to near control levels. This study demonstrates thatmore » alcohol reduces key energy sensing proteins and that replacement of LCT by MCT affects the transcription of these genes. Since there is a pathophysiological link between SIRT1 and PGC-1{alpha} and mitochondrial energy, the implication of the study is that mitochondrial dysfunction due to alcohol abuse can be treated by dietary modifications.« less

CPT1{alpha} over-expression increases long-chain fatty acid oxidation and reduces cell viability with incremental palmitic acid concentration in 293T cells

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

Jambor de Sousa, Ulrike L.; Koss, Michael D.; Fillies, Marion

2005-12-16

To test the cellular response to an increased fatty acid oxidation, we generated a vector for an inducible expression of the rate-limiting enzyme carnitine palmitoyl-transferase 1{alpha} (CPT1{alpha}). Human embryonic 293T kidney cells were transiently transfected and expression of the CPT1{alpha} transgene in the tet-on vector was activated with doxycycline. Fatty acid oxidation was measured by determining the conversion of supplemented, synthetic cis-10-heptadecenoic acid (C17:1n-7) to C15:ln-7. CPT1{alpha} over-expression increased mitochondrial long-chain fatty acid oxidation about 6-fold. Addition of palmitic acid (PA) decreased viability of CPT1{alpha} over-expressing cells in a concentration-dependent manner. Both, PA and CPT1{alpha} over-expression increased cell death. Interestingly,more » PA reduced total cell number only in cells over-expressing CPT1{alpha}, suggesting an effect on cell proliferation that requires PA translocation across the mitochondrial inner membrane. This inducible expression system should be well suited to study the roles of CPT1 and fatty acid oxidation in lipotoxicity and metabolism in vivo.« less

Formation and self-organization kinetics of alpha-CD/PEO-based pseudo-polyrotaxanes in water. A specific behavior at 30 degrees C.

PubMed

Travelet, Christophe; Schlatter, Guy; Hébraud, Pascal; Brochon, Cyril; Lapp, Alain; Hadziioannou, Georges

2009-08-04

alpha-Cyclodextrins (alpha-CDs) have the ability to form inclusion complexes with poly(ethylene oxide) (PEO) polymer chains. These pseudo-polyrotaxanes (PPRs) can be obtained by quenching an alpha-CD/PEO mixture in water from 70 degrees C down to a lower temperature (typically in the range from 5 to 30 degrees C) thanks to favorable interactions between alpha-CD cavities and PEO chains. Moreover, starting from a liquid alpha-CD/PEO mixture at a total mass fraction of 15% w/w at 70 degrees C, the formation of PPRs with time at a lower temperature induces a white physical gel with time, and phase separation is observed. We established that PPR molecules are exclusively found in the precipitated phase although unthreaded alpha-CD molecules and unthreaded PEO chains are in the liquid phase. At 30 degrees C, the physical gel formation is much slower than at 5 degrees C. At 30 degrees C, we established that, in a first step, alpha-CDs thread onto PEO chains, forming PPR molecules which are not in good solvent conditions in water. At a higher length scale, rapid aggregation of the PPR molecules occurs, and threaded alpha-CD-based nanocylinders form (cylinder length L = 5.7 nm and cylinder radius R = 4.7 nm). At a higher length scale, alpha-CD-based nanocylinders associate in a Gaussian way, engendering the formation of precipitated domains which are responsible for the high turbidity of the studied system. At the end of this first step (i.e., after 20 min), the system still remains liquid and the PPRs are totally formed. Then, in a second step (i.e., after 150 min), the system undergoes its reorganization characterized by a compacity increase of the precipitated domains and forms a physical gel. We found that PPRs are totally formed after 20 min at 30 degrees C and that the system stays in a nongel state up to 150 min. This opens new perspectives regarding the PPR chemical modification: between these two characteristic times, we can easily envisage an efficient chemical modification of the PPR molecules in water, as for instance an end-capping reaction leading to the synthesis of polyrotaxanes.

Interactions among the branched-chain amino acids and their effects on methionine utilization in growing pigs: effects on plasma amino- and keto-acid concentrations and branched-chain keto-acid dehydrogenase activity.

PubMed

Langer, S; Scislowski, P W; Brown, D S; Dewey, P; Fuller, M F

2000-01-01

The present experiment was designed to elucidate the mechanism of the methionine-sparing effect of excess branched-chain amino acids (BCAA) reported in the previous paper (Langer & Fuller, 2000). Twelve growing gilts (30-35 kg) were prepared with arterial catheters. After recovery, they received for 7 d a semipurified diet with a balanced amino acid pattern. On the 7th day blood samples were taken before (16 h postabsorptive) and after the morning meal (4 h postprandial). The animals were then divided into three groups and received for a further 7 d a methionine-limiting diet (80% of requirement) (1) without any amino acid excess; (2) with excess leucine (50% over requirement); or (3) with excesses of all three BCAA (leucine, isoleucine, valine, each 50% over the requirement). On the 7th day blood samples were taken as in the first period, after which the animals were killed and liver and muscle samples taken. Plasma amino acid and branched-chain keto acid (BCKA) concentrations in the blood and branched-chain keto-acid dehydrogenase (BCKDH; EC 1.2.4.4) activity in liver and muscle homogenates were determined. Compared with those on the balanced diet, pigs fed on methionine-limiting diets had significantly lower (P < 0.05) plasma methionine concentrations in the postprandial but not in the postabsorptive state. There was no effect of either leucine or a mixture of all three BCAA fed in excess on plasma methionine concentrations. Excess dietary leucine reduced (P < 0.05) the plasma concentrations of isoleucine and valine in both the postprandial and postabsorptive states. Plasma concentrations of the BCKA reflected the changes in the corresponding amino acids. Basal BCKDH activity in the liver and total BCKDH activity in the biceps femoris muscle were significantly (P < 0.05) increased by excesses of leucine or all BCAA.

Energy Contents of Frequently Ordered Restaurant Meals and Comparison with Human Energy Requirements and US Department of Agriculture Database Information: A Multisite Randomized Study

PubMed Central

Urban, Lorien E.; Weber, Judith L.; Heyman, Melvin B.; Schichtl, Rachel L.; Verstraete, Sofia; Lowery, Nina S.; Das, Sai Krupa; Schleicher, Molly M.; Rogers, Gail; Economos, Christina; Masters, William A.; Roberts, Susan B.

2017-01-01

Background Excess energy intake from meals consumed away from home is implicated as a major contributor to obesity, and ~50% of US restaurants are individual or small-chain (non–chain) establishments that do not provide nutrition information. Objective To measure the energy content of frequently ordered meals in non–chain restaurants in three US locations, and compare with the energy content of meals from large-chain restaurants, energy requirements, and food database information. Design A multisite random-sampling protocol was used to measure the energy contents of the most frequently ordered meals from the most popular cuisines in non–chain restaurants, together with equivalent meals from large-chain restaurants. Setting Meals were obtained from restaurants in San Francisco, CA; Boston, MA; and Little Rock, AR, between 2011 and 2014. Main outcome measures Meal energy content determined by bomb calorimetry. Statistical analysis performed Regional and cuisine differences were assessed using a mixed model with restaurant nested within region×cuisine as the random factor. Paired t tests were used to evaluate differences between non–chain and chain meals, human energy requirements, and food database values. Results Meals from non–chain restaurants contained 1,205±465 kcal/meal, amounts that were not significantly different from equivalent meals from large-chain restaurants (+5.1%; P=0.41). There was a significant effect of cuisine on non–chain meal energy, and three of the four most popular cuisines (American, Italian, and Chinese) had the highest mean energy (1,495 kcal/meal). Ninety-two percent of meals exceeded typical energy requirements for a single eating occasion. Conclusions Non–chain restaurants lacking nutrition information serve amounts of energy that are typically far in excess of human energy requirements for single eating occasions, and are equivalent to amounts served by the large-chain restaurants that have previously been criticized for providing excess energy. Restaurants in general, rather than specific categories of restaurant, expose patrons to excessive portions that induce overeating through established biological mechanisms. PMID:26803805

Energy Contents of Frequently Ordered Restaurant Meals and Comparison with Human Energy Requirements and U.S. Department of Agriculture Database Information: A Multisite Randomized Study.

PubMed

Urban, Lorien E; Weber, Judith L; Heyman, Melvin B; Schichtl, Rachel L; Verstraete, Sofia; Lowery, Nina S; Das, Sai Krupa; Schleicher, Molly M; Rogers, Gail; Economos, Christina; Masters, William A; Roberts, Susan B

2016-04-01

Excess energy intake from meals consumed away from home is implicated as a major contributor to obesity, and ∼50% of US restaurants are individual or small-chain (non-chain) establishments that do not provide nutrition information. To measure the energy content of frequently ordered meals in non-chain restaurants in three US locations, and compare with the energy content of meals from large-chain restaurants, energy requirements, and food database information. A multisite random-sampling protocol was used to measure the energy contents of the most frequently ordered meals from the most popular cuisines in non-chain restaurants, together with equivalent meals from large-chain restaurants. Meals were obtained from restaurants in San Francisco, CA; Boston, MA; and Little Rock, AR, between 2011 and 2014. Meal energy content determined by bomb calorimetry. Regional and cuisine differences were assessed using a mixed model with restaurant nested within region×cuisine as the random factor. Paired t tests were used to evaluate differences between non-chain and chain meals, human energy requirements, and food database values. Meals from non-chain restaurants contained 1,205±465 kcal/meal, amounts that were not significantly different from equivalent meals from large-chain restaurants (+5.1%; P=0.41). There was a significant effect of cuisine on non-chain meal energy, and three of the four most popular cuisines (American, Italian, and Chinese) had the highest mean energy (1,495 kcal/meal). Ninety-two percent of meals exceeded typical energy requirements for a single eating occasion. Non-chain restaurants lacking nutrition information serve amounts of energy that are typically far in excess of human energy requirements for single eating occasions, and are equivalent to amounts served by the large-chain restaurants that have previously been criticized for providing excess energy. Restaurants in general, rather than specific categories of restaurant, expose patrons to excessive portions that induce overeating through established biological mechanisms. Copyright © 2016 Academy of Nutrition and Dietetics. Published by Elsevier Inc. All rights reserved.

Optical and UV spectroscopy of the peculiar RS CVn system RT Lacertae

NASA Technical Reports Server (NTRS)

Huenemoerder, D. P.; Barden, S. C.

1986-01-01

H-alpha and H-beta spectra of the peculiar double-lined RS CVn binary RT Lacertae have been obtained using the IUE, together with a ground-based coude-feed telescope at KPNO. The ground-based spectra show an asymmetry related to the orbital phase in the H-alpha profile. H-beta profiles showed excess emission in one hemisphere and excess absorption in the other, with a broad Gaussian emission component superposed on the excess H-alpha line. A radial velocity curve was derived to estimate the mass ratio and geometry of the system. It is shown that the component of RT Lac fills 80-90 percent of the equilibrium Roche surface. Low-resolution ultraviolet data show that the supposed cooler component is bluer than its companion, suggesting evidence of a scattering shell or a cloud produced by the splash of a gas stream. The phase behavior of the low resolution ultraviolet data support the conclusion that RT Lac is a mass transfer system and that mass transfer is the primary cause of its activity.

Molecular basis of maple syrup urine disease: Novel mutations at the E1[alpha] locus that impair E1([alpha][sub 2][beta][sub 2]) assembly or decrease steady-state E1[alpha] mRNA levels of branched-chain [alpha]-keto acid dehydrogenase complex

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

Chuang, J.L.; Fisher, C.R.; Chuang, D.T.

1994-08-01

The authors report the occurrence of three novel mutations in the E1[alpha] (BCKDHA) locus of the branched-chain [alpha]-keto acid dehydrogenase (BCKAD) complex that cause maple syrup urine disease (MSUD). An 8-bp deletion in exon 7 is present in one allele of a compound-heterozygous patient (GM-649). A single C nucleotide insertion in exon 2 occurs in one allele of an intermediate-MSUD patient (Lo). The second allele of patient Lo carries an A-to-G transition in exon 9 of the E1[alpha] gene. This missense mutation changes Tyr-368 to Cys (Y368C) in the E1[alpha] subunit. Both the 8-bp deletion and the single C insertionmore » generate a downstream nonsense codon. Both mutations appear to be associated with a low abundance of the mutant E1[alpha] mRNA, as determined by allele-specific oligonucleotide probing. Transfection studies strongly suggest that the Y368C substitution in the E1[alpha] subunit impairs its proper assembly with the normal E1[beta]. Unassembled as well as misassembled E1[alpha] and E1[beta] subunits are degraded in the cell. 32 refs., 8 figs.« less

High-resolution protein design with backbone freedom.

PubMed

Harbury, P B; Plecs, J J; Tidor, B; Alber, T; Kim, P S

1998-11-20

Recent advances in computational techniques have allowed the design of precise side-chain packing in proteins with predetermined, naturally occurring backbone structures. Because these methods do not model protein main-chain flexibility, they lack the breadth to explore novel backbone conformations. Here the de novo design of a family of alpha-helical bundle proteins with a right-handed superhelical twist is described. In the design, the overall protein fold was specified by hydrophobic-polar residue patterning, whereas the bundle oligomerization state, detailed main-chain conformation, and interior side-chain rotamers were engineered by computational enumerations of packing in alternate backbone structures. Main-chain flexibility was incorporated through an algebraic parameterization of the backbone. The designed peptides form alpha-helical dimers, trimers, and tetramers in accord with the design goals. The crystal structure of the tetramer matches the designed structure in atomic detail.

Alpha-decay chains of superheavy nuclei 292-296118

NASA Astrophysics Data System (ADS)

Singh, U. K.; Kumawat, M.; Saxena, G.; Kaushik, M.; Jain, S. K.

2018-05-01

We have employed relativistic mean-field plus BCS (RMF+BCS) approach for the study of even-even superheavy nuclei with Z = 118 which is the last and recent observed element in the periodic chart so far. Our study includes binding energies, Qα values, alpha-decay half-lives and spontaneous decay half-lives along with comparison of available experimental data and the results of FRDM calculations. We find an excellent match with the only known decay chain of 294118 for Z = 118 so far and predict decay chain of 292118 and 296118 in consistency with known experimental decay chains and FRDM results. These results may provide a very helpful insight to conduct experiments for realizing the presence of nuclei with Z = 118.

Complete primary structure of rainbow trout type I collagen consisting of alpha1(I)alpha2(I)alpha3(I) heterotrimers.

PubMed

Saito, M; Takenouchi, Y; Kunisaki, N; Kimura, S

2001-05-01

The subunit compositions of skin and muscle type I collagens from rainbow trout were found to be alpha1(I)alpha2(I)alpha3(I) and [alpha1(I)](2)alpha2(I), respectively. The occurrence of alpha3(I) has been observed only for bonyfish. The skin collagen exhibited more susceptibility to both heat denaturation and MMP-13 digestion than the muscle counterpart; the former had a lower denaturation temperature by about 0.5 degrees C than the latter. The lower stability of skin collagen, however, is not due to the low levels of imino acids because the contents of Pro and Hyp were almost constant in both collagens. On the other hand, some cDNAs coding for the N-terminal and/or a part of triple-helical domains of proalpha(I) chains were cloned from the cDNA library of rainbow trout fibroblasts. These cDNAs together with the previously cloned collagen cDNAs gave information about the complete primary structure of type I procollagen. The main triple-helical domain of each proalpha(I) chain had 338 uninterrupted Gly-X-Y triplets consisting of 1014 amino acids and was unique in its high content of Gly-Gly doublets. In particular, the bonyfish-specific alpha(I) chain, proalpha3(I) was characterized by the small number of Gly-Pro-Pro triplets, 19, and the large number of Gly-Gly doublets, 38, in the triple-helical domain, compared to 23 and 22, respectively, for proalpha1(I). The small number of Gly-Pro-Pro and the large number of Gly-Gly in proalpha3(I) was assumed to partially loosen the triple-helical structure of skin collagen, leading to the lower stability of skin collagen mentioned above. Finally, phylogenetic analyses revealed that proalpha3(I) had diverged from proalpha1(I). This study is the first report of the complete primary structure of fish type I procollagen.

Effect of low electric fields on alpha scintillation light yield in liquid argon

NASA Astrophysics Data System (ADS)

Agnes, P.; Albuquerque, I. F. M.; Alexander, T.; Alton, A. K.; Asner, D. M.; Back, H. O.; Baldin, B.; Biery, K.; Bocci, V.; Bonfini, G.; Bonivento, W.; Bossa, M.; Bottino, B.; Brigatti, A.; Brodsky, J.; Budano, F.; Bussino, S.; Cadeddu, M.; Cadoni, M.; Calaprice, F.; Canci, N.; Candela, A.; Caravati, M.; Cariello, M.; Carlini, M.; Catalanotti, S.; Cavalcante, P.; Chepurnov, A.; Cicalò, C.; Cocco, A. G.; Covone, G.; D'Angelo, D.; D'Incecco, M.; Davini, S.; De Cecco, S.; De Deo, M.; De Vincenzi, M.; Derbin, A.; Devoto, A.; Di Eusanio, F.; Di Pietro, G.; Dionisi, C.; Edkins, E.; Empl, A.; Fan, A.; Fiorillo, G.; Fomenko, K.; Forster, G.; Franco, D.; Gabriele, F.; Galbiati, C.; Giagu, S.; Giganti, C.; Giovanetti, G. K.; Goretti, A. M.; Granato, F.; Gromov, M.; Guan, M.; Guardincerri, Y.; Hackett, B. R.; Herner, K.; Hughes, D.; Humble, P.; Hungerford, E. V.; Ianni, A.; James, I.; Johnson, T. N.; Jollet, C.; Keeter, K.; Kendziora, C. L.; Koh, G.; Korablev, D.; Korga, G.; Kubankin, A.; Li, X.; Lissia, M.; Loer, B.; Lombardi, P.; Longo, G.; Ma, Y.; Machulin, I. N.; Mandarano, A.; Mari, S. M.; Maricic, J.; Marini, L.; Martoff, C. J.; Meregaglia, A.; Meyers, P. D.; Milincic, R.; Miller, J. D.; Montanari, D.; Monte, A.; Mount, B. J.; Muratova, V. N.; Musico, P.; Napolitano, J.; Navrer Agasson, A.; Odrowski, S.; Oleinik, A.; Orsini, M.; Ortica, F.; Pagani, L.; Pallavicini, M.; Pantic, E.; Parmeggiano, S.; Pelczar, K.; Pelliccia, N.; Pocar, A.; Pordes, S.; Pugachev, D. A.; Qian, H.; Randle, K.; Ranucci, G.; Razeti, M.; Razeto, A.; Reinhold, B.; Renshaw, A. L.; Rescigno, M.; Riffard, Q.; Romani, A.; Rossi, B.; Rossi, N.; Rountree, D.; Sablone, D.; Saggese, P.; Sands, W.; Savarese, C.; Schlitzer, B.; Segreto, E.; Semenov, D. A.; Shields, E.; Singh, P. N.; Skorokhvatov, M. D.; Smirnov, O.; Sotnikov, A.; Stanford, C.; Suvorov, Y.; Tartaglia, R.; Tatarowicz, J.; Testera, G.; Tonazzo, A.; Trinchese, P.; Unzhakov, E. V.; Verducci, M.; Vishneva, A.; Vogelaar, B.; Wada, M.; Walker, S.; Wang, H.; Wang, Y.; Watson, A. W.; Westerdale, S.; Wilhelmi, J.; Wojcik, M. M.; Xiang, X.; Xiao, X.; Xu, J.; Yang, C.; Zhong, W.; Zhu, C.; Zuzel, G.

2017-01-01

Measurements were made of scintillation light yield of alpha particles from the 222Rn decay chain within the DarkSide-50 liquid argon time projection chamber. The light yield was found to increase as the applied electric field increased, with alphas in a 200 V/cm electric field exhibiting a ~2% increase in light yield compared to alphas in no field.

Functional CD1d and/or NKT cell invariant chain transcript in horse, pig, African elephant and guinea pig, but not in ruminants.

PubMed

Looringh van Beeck, Frank A; Reinink, Peter; Hermsen, Roel; Zajonc, Dirk M; Laven, Marielle J; Fun, Axel; Troskie, Milana; Schoemaker, Nico J; Morar, Darshana; Lenstra, Johannes A; Vervelde, Lonneke; Rutten, Victor P M G; van Eden, Willem; Van Rhijn, Ildiko

2009-04-01

CD1d-restricted invariant natural killer T cells (NKT cells) have been well characterized in humans and mice, but it is unknown whether they are present in other species. Here we describe the invariant TCR alpha chain and the full length CD1d transcript of pig and horse. Molecular modeling predicts that porcine (po) invariant TCR alpha chain/poCD1d/alpha-GalCer and equine (eq) invariant TCR alpha chain/eqCD1d/alpha-GalCer form complexes that are highly homologous to the human complex. Since a prerequisite for the presence of NKT cells is the expression of CD1d protein, we performed searches for CD1D genes and CD1d transcripts in multiple species. Previously, cattle and guinea pig have been suggested to lack CD1D genes. The CD1D genes of European taurine cattle (Bos taurus) are known to be pseudogenes because of disrupting mutations in the start codon and in the donor splice site of the first intron. Here we show that the same mutations are found in six other ruminants: African buffalo, sheep, bushbuck, bongo, N'Dama cattle, and roe deer. In contrast, intact CD1d transcripts were found in guinea pig, African elephant, horse, rabbit, and pig. Despite the discovery of a highly homologous NKT/CD1d system in pig and horse, our data suggest that functional CD1D and CD1d-restricted NKT cells are not universally present in mammals.

Calorimetric and spectroscopic studies of the thermotropic phase behavior of lipid bilayer model membranes composed of a homologous series of linear saturated phosphatidylserines.

PubMed Central

Lewis, R N; McElhaney, R N

2000-01-01

The thermotropic phase behavior of lipid bilayer model membranes composed of the even-numbered, N-saturated 1,2-diacyl phosphatidylserines was studied by differential scanning calorimetry and by Fourier-transform infrared and (31)P-nuclear magnetic resonance spectroscopy. At pH 7.0, 0.1 M NaCl and in the absence of divalent cations, aqueous dispersions of these lipids, which have not been incubated at low temperature, exhibit a single calorimetrically detectable phase transition that is fully reversible, highly cooperative, and relatively energetic, and the transition temperatures and enthalpies increase progressively with increases in hydrocarbon chain length. Our spectroscopic observations confirm that this thermal event is a lamellar gel (L(beta))-to-lamellar liquid crystalline (L(alpha)) phase transition. However, after low temperature incubation, the L(beta)/L(alpha) phase transition of dilauroyl phosphatidylserine is replaced by a higher temperature, more enthalpic, and less cooperative phase transition, and an additional lower temperature, less enthalpic, and less cooperative phase transition appears in the longer chain phosphatidylserines. Our spectroscopic results indicate that this change in thermotropic phase behavior when incubated at low temperatures results from the conversion of the L(beta) phase to a highly ordered lamellar crystalline (L(c)) phase. Upon heating, the L(c) phase of dilauroyl phosphatidylserine converts directly to the L(alpha) phase at a temperature slightly higher than that of its original L(beta)/L(alpha) phase transition. Calorimetrically, this process is manifested by a less cooperative but considerably more energetic, higher-temperature phase transition, which replaces the weaker L(beta)/L(alpha) phase transition alluded to above. However, with the longer chain compounds, the L(c) phase first converts to the L(beta) phase at temperatures some 10-25 degrees C below that at which the L(beta) phase converts to the L(alpha) phase. Our results also suggest that shorter chain homologues form L(c) phases that are structurally related to, but more ordered than, those formed by the longer chain homologues, but that these L(c) phases are less ordered than those formed by other phospholipids. These studies also suggest that polar/apolar interfaces of the phosphatidylserine bilayers are more hydrated than those of other glycerolipid bilayers, possibly because of interactions between the polar headgroup and carbonyl groups of the fatty acyl chains. PMID:11023908

Methods and Data Used to Investigate Polonium-210 as a Source of Excess Gross-Alpha Radioactivity in Ground Water, Churchill County, Nevada

USGS Publications Warehouse

Seiler, Ralph L.

2007-01-01

Ground water is the major source of drinking water in the Carson River Basin, California and Nevada. Previous studies have shown that uranium and gross-alpha radioactivities in ground water can be greater than U.S. Environmental Protection Agency Maximum Contaminant Levels, particularly in the Carson Desert, Churchill County, Nevada. Studies also have shown that the primary source of the gross-alpha radioactivity and alpha-emitting radionuclides in ground water is the dissolution of uranium-rich granitic rocks and basin-fill sediments that have their origins in the Sierra Nevada. However, ground water sampled from some wells in the Carson Desert had gross-alpha radioactivities greater than could be accounted for by the decay of dissolved uranium. The occurrence of polonium-210 (Po-210) was hypothesized to explain the higher than expected gross-alpha radioactivities. This report documents and describes the study design, field and analytical methods, and data used to determine whether Po-210 is the source of excess gross-alpha radioactivity in ground water underlying the Carson Desert in and around Fallon, Nevada. Specifically, this report presents: 1) gross alpha and uranium radioactivities for 100 wells sampled from June to September 2001; and 2) pH, dissolved oxygen, specific conductance, and Po-210 radioactivity for 25 wells sampled in April and June 2007. Results of quality-control samples for the 2007 dataset are also presented.

Surface structural conformations of fibrinogen polypeptides for improved biocompatibility.

PubMed

Yaseen, Mohammed; Zhao, Xiubo; Freund, Amy; Seifalian, Alexander M; Lu, Jian R

2010-05-01

This work reports on how incorporation of silica nanocages into poly(urethane) copolymers (PU) affects conformational orientations of adsorbed fibrinogen and how different surfaces subsequently influenced HeLa cell attachment and proliferation. Incorporation of 2 wt% silica nanocages into poly(urethane) (PU4) substantially altered the surface topography of the films and some 50% of the surface was covered with the nanocages due to their preferential exposure. AFM studies revealed the deposition of a dense protein network on the soft polymeric domains of PU4 and much reduced fibrinogen adsorption on the hard nanocage domains. As on the bare SiO(2) control surface, fibrinogen molecules adsorbed on top of the hard nanocages mainly took the dominant trinodular structures in monomeric and dimeric forms. In addition, net positively charged long alpha chains were prone to being hidden beneath the D domains whilst gamma chains predominantly remained exposed. Dynamic interfacial adsorption as probed by spectroscopic ellipsometry revealed fast changes in interfacial conformation induced by electrostatic interactions between different segments of fibrinogen and the surface, consistent with the AFM imaging. On the PU surfaces without nanocage incorporation (PUA), however, adsorbed fibrinogen molecules formed beads-like chain networks, consistent with the structure featured on the soft PU4 domains, showing very different effects of surface chemical nature. Monoclonal antibodies specific to the alpha and gamma chains showed reduced alpha but increased gamma chain binding at the silicon oxide control and PU4 surfaces, whilst on the PUA, C18 and amine surfaces (organic surface controls) the opposite binding trend was detected with alpha chain binding dominant, showing different fibrinogen conformations. Cell attachment studies revealed differences in cell attachment and proliferation, consistent with the different polypeptide conformations on the two types of surfaces, showing a strong preference to the extent of exposure of gamma chains. Crown Copyright 2010. Published by Elsevier Ltd. All rights reserved.

STELLAR MEMBERSHIP AND DUSTY DEBRIS DISKS IN THE {alpha} PERSEI CLUSTER

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

Zuckerman, B.; Melis, Carl; Rhee, Joseph H.

2012-06-10

Because of its proximity to the Galactic plane, reliable identification of members of the {alpha} Persei cluster is often problematic. Based primarily on membership evaluations contained in six published papers, we constructed a mostly complete list of high-fidelity members of spectral type G and earlier that lie within 3 arc degrees of the cluster center. {alpha} Persei was the one nearby, rich, young open cluster not surveyed with the Spitzer Space Telescope. We examined the first and final data releases of the Wide-field Infrared Survey Explorer and found 11, or perhaps 12, {alpha} Per cluster members that have excess mid-infraredmore » emission above the stellar photosphere attributable to an orbiting dusty debris disk. The most unusual of these is V488 Per, a K-type star with an excess IR luminosity 16% (or more) of the stellar luminosity; this is a larger excess fraction than that of any other known dusty main-sequence star. Much of the dust that orbits V488 Per is at a temperature of {approx}800 K; if these grains radiate like blackbodies, then they lie only {approx}0.06 AU from the star. The dust is probably the aftermath of a collision of two planetary embryos or planets with small semimajor axes; such orbital radii are similar to those of many of the transiting planets discovered by the Kepler satellite.« less

Light-induced yellowing of selectively 13C-enriched dehydrogenation polymers (DHPs). Part 1, Side-chain 13C-enriched DHP ([alpha], [beta], and [gamma]-13C)

Treesearch

Jim Parkas; Magnus Paulsson; Terashima Noritsugu; Ulla Westermark; Sally Ralph

2004-01-01

Light-induced yellowing has been studied using side-chain ([alpha], [beta], and [gamma]) 13C-enriched DHP (dehydrogenation polymer) and quantitative solution state 13C NMR spectroscopy. The DHP was formed from 13C-enriched coniferin using an enzymatic system consisting of [beta]-glucosidase, glucose oxidase, and peroxidase in a pH 6 buffer solution. The DHP was applied...

A new mechanism for DNA alterations induced by alpha particles such as those emitted by radon and radon progeny.

PubMed Central

Lehnert, B E; Goodwin, E H

1997-01-01

The mechanism(s) by which alpha (alpha) particles like those emitted from inhaled radon and radon progeny cause their carcinogenic effects in the lung remains unclear. Although direct nuclear traversals by alpha-particles may be involved in mediating these outcomes, increasing evidence indicates that a particles can cause alterations in DNA in the absence of direct hits to cell nuclei. Using the occurrence of excessive sister chromatid exchanges (SCE) as an index of DNA damage in human lung fibroblasts, we investigated the hypothesis that alpha-particles may induce DNA damage through the generation of extracellular factors. We have found that a relatively low dose of alpha-particles can result in the generation of extracellular factors, which, upon transfer to unexposed normal human cells, can cause excessive SCE to an extent equivalent to that observed when the cells are directly irradiated with the same irradiation dose. A short-lived, SCE-inducing factor(s) is generated in alpha-irradiated culture medium containing serum in the absence of cells. A more persistent SCE-inducing factor(s), which can survive freeze-thaw and is heat labile is produced by fibroblasts after exposure to the alpha-particles. These results indicate that the initiating target for alpha-particle-induced genetic changes can be larger than a cell's nucleus or even a whole cell. How transmissible factors like those observed here in vitro may extend to the in vivo condition in the context of a-particle-induced carcinogenesis in the respiratory tract remains to be determined. PMID:9400706

ISOLATED WOLF-RAYET STARS AND O SUPERGIANTS IN THE GALACTIC CENTER REGION IDENTIFIED VIA PASCHEN-{alpha} EXCESS

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

Mauerhan, J. C.; Stolovy, S. R.; Cotera, A.

We report the discovery of 19 hot, evolved, massive stars near the Galactic center region (GCR). These objects were selected for spectroscopy owing to their detection as strong sources of Paschen-{alpha} (P{alpha}) emission-line excess, following a narrowband imaging survey of the central 0.{sup 0}65 x 0.{sup 0}25 (l, b) around Sgr A* with the Hubble Space Telescope. Discoveries include six carbon-type (WC) and five nitrogen-type (WN) Wolf-Rayet stars, six O supergiants, and two B supergiants. Two of the O supergiants have X-ray counterparts having properties consistent with solitary O stars and colliding-wind binaries. The infrared photometry of 17 stars ismore » consistent with the Galactic center distance, but 2 of them are located in the foreground. Several WC stars exhibit a relatively large infrared excess, which is possibly thermal emission from hot dust. Most of the stars appear scattered throughout the GCR, with no relation to the three known massive young clusters; several others lie near the Arches and Quintuplet clusters and may have originated within one of these systems. The results of this work bring the total sample of Wolf-Rayet (WR) stars in the GCR to 88. All sources of strong P{alpha} excess have been identified in the area surveyed with HST, which implies that the sample of WN stars in this region is near completion, and is dominated by late (WNL) types. The current WC sample, although probably not complete, is almost exclusively dominated by late (WCL) types. The observed WR subtype distribution in the GCR is a reflection of the intrinsic rarity of early subtypes (WNE and WCE) in the inner Galaxy, an effect that is driven by metallicity.« less

alpha-Ketoglutarate application in hemodialysis patients improves amino acid metabolism.

PubMed

Riedel, E; Nündel, M; Hampl, H

1996-01-01

In hemodialysis patients, free amino acids and alpha-ketoacids in plasma were determined by fluorescence HPLC to assess the effect of alpha-ketoglutarate administration in combination with the phosphate binder calcium carbonate on the amino acid metabolism. During 1 year of therapy in parallel to inorganic phosphate, urea in plasma decreased significantly, histidine, arginine and proline as well as branched chain alpha-ketoacids, in particular alpha-ketoisocaproate, a regulator of protein metabolism, increased. Thus, administration of alpha-ketoglutarate with calcium carbonate effectively improves amino acid metabolism in hemodialysis patients as it decreases hyperphosphatemia.

Molecular analysis and association with clinical and laboratory manifestations in children with sickle cell anemia

PubMed Central

Camilo-Araújo, Roberta Faria; Amancio, Olga Maria Silverio; Figueiredo, Maria Stella; Cabanãs-Pedro, Ana Carolina; Braga, Josefina Aparecida Pellegrini

2014-01-01

Objectives To analyze the frequency of βS-globin haplotypes and alpha-thalassemia, and their influence on clinical manifestations and the hematological profile of children with sickle cell anemia. Method The frequency of βS-globin haplotypes and alpha-thalassemia and any association with clinical and laboratorial manifestations were determined in 117 sickle cell anemia children aged 3–71 months. The confirmation of hemoglobin SS and determination of the haplotypes were achieved by polymerase chain reaction-restriction fragment length polymorphism, and alpha-thalassemia genotyping was by multiplex polymerase chain reaction (single-tube multiplex-polymerase chain reaction). Results The genotype distribution of haplotypes was 43 (36.7%) Central African Republic/Benin, 41 (35.0%) Central African Republic/Central African Republic, 20 (17.0%) Rare/atypical, and 13 (11.1%) Benin/Benin. The frequency of the α3.7 deletion was 1.71% as homozygous (−α3.7/−α3.7) and 11.9% as heterozygous (−α3.7/αα). The only significant association in respect to haplotypes was related to the mean corpuscular volume. The presence of alpha-thalassemia was significantly associated to decreases in mean corpuscular volume, mean corpuscular hemoglobin and reticulocyte count and to an increase in the red blood cell count. There were no significant associations of βS-globin haplotypes and alpha-thalassemia with clinical manifestations. Conclusions In the study population, the frequency of alpha-thalassemia was similar to published data in Brazil with the Central African Republic haplotype being the most common, followed by the Benin haplotype. βS-globin haplotypes and interaction between alpha-thalassemia and sickle cell anemia did not influence fetal hemoglobin concentrations or the number of clinical manifestations. PMID:25305165

Diving behaviour and haemoglobin function: the primary structure of the alpha- and beta-chains of the sea turtle (Caretta caretta) and its functional implications.

PubMed

Petruzzelli, R; Aureli, G; Lania, A; Galtieri, A; Desideri, A; Giardina, B

1996-06-15

The amino acid sequence of the alpha- and beta-chains of haemoglobin (Hb) from the loggerhead sea turtle (Caretta caretta) has been determined. Comparison with that of human Hb shows differences in several residues involved in both alpha 1 beta 1 and alpha 1 beta 2 packing contacts. On the whole, in spite of the mutations, the essential characteristics of both interfaces seem to be maintained. The functional properties of the sea turtle Hb have been investigated at different temperatures and as a function of proton, chloride and organic phosphate concentrations. In addition to overall similarities shared with most of the vertebrate Hbs previously described, this molecule shows significant differences which could be related to the life behaviour of the turtle. In fact, while the shape of the Bohr-effect curve is well adapted for gas exchange during prolonged dives, the very small enthalpy change for O2 binding ensures that O2 delivery becomes essentially insensitive to the temperature changes of the environment. Moreover, and similarly to the case of emperor penguin Hb, the small alkaline Bohr effect appears to be only choride-linked, since the pH dependence of the O2 affinity is abolished in the absence of this ion. These functional characteristics are discussed on the basis of the primary structure of alpha- and beta-chains.

Pattern of cytokine receptors expressed by human dendritic cells migrated from dermal explants.

PubMed Central

Larregina, A T; Morelli, A E; Kolkowski, E; Sanjuan, N; Barboza, M E; Fainboim, L

1997-01-01

Different reasons account for the lack of information about the expression of cytokine receptors on human dendritic cells (DC): (a) DC are a trace population; (b) the proteolytic treatment used to isolate DC may alter enzyme-sensitive epitopes; and (c) low numbers of receptors per cell. In the present work the expression of cytokine receptors was analysed by flow cytometry on the population of dermal DC (DDC) that spontaneously migrate from short-term culture dermal explants. DDC obtained after dermal culture were CD1alow, CD1b+, CD1c+, human leucocyte antigen (HLA)-DR+, CD11chigh, CD11b+ and CD32+. The DC lineage was confirmed by ultrastructural analysis. DDC expressed interleukin (IL)-1R type 1 (monoclonal antibody (mAb) hIL-1R1-M1; and 6B5); IL-1R type 2 (mAb hIL-1R2-M22); IL-2R alpha chain (mAb anti-Tac; and hIL-2R-M1) and IL-2R gamma chain (mAb 3B5; and AG14C). DDC did not stain for IL-2R beta chain using four mAbs recognizing two different epitopes of IL-2R beta (mAb 2R-B; Mik-beta 1; and CF1; Mik-beta 3, respectively). DDC were also positive for the cytokine binding chains (alpha chains) of IL-3R (mAb 9F5); IL-4R (mAb hIL-4R-M57; and S456C9); and IL-7R (mAb hIL-7R-M20; and R3434). DDC showed low levels of IL-6R alpha chain (mAb B-F19; B-R6; and B-E23) and its signal transducer gp130 (mAb A2; and B1). DDC strongly expressed interferon-gamma receptor (IFN-gamma R) (mAb GIR-208) and were negative for IL-8R (mAb B-G20; and B-F25). All DDC were highly positive for granulocyte-macrophage colony-stimulating factor receptor (GM-CSFR) alpha chain (mAb hGM-CSFR-M1; SC06; SC04, and 8G6) and to a lesser extent for the common beta chain of GM-CSFR, IL-3R and IL-5R (mAb 3D7). On the other hand, reactivity was not found for granulocyte colony-stimulating factor receptor (G-CSFR) (mAb hGCSFR-M1) nor macrophage colony-stimulating factor receptor (M-CSFR) (mAb 7-7A3-17) confirming the DC lineage of DDC. As previously reported for lymphoid DC, DDC expressed tumour necrosis factor receptort (TNFR) 75000 MW (mAb utr-1; hTNFR-M1; and MR2-1) but lacked TNFR 55000 MW (mAb htr-9; MR1-1; and MR1-2). In summary, DDC express receptors for a broad panel of cytokines, even receptors for cytokines whose effects on DC are still unknown (i.e. IL-2R alpha gamma; IL-6R alpha/gp 130; IL-7R alpha gamma). Images Figure 1 PMID:9227332

Optical and UV spectroscopy of the peculiar RS CVn system, RT Lacertae

NASA Technical Reports Server (NTRS)

Huenemoerder, D. P.; Barden, S. C.

1985-01-01

Spectra in the H-alpha and H-beta regions of the peculiar double-lined RS CVn binary, RT Lacertae, were obtained in the fall of 1984. Limited International Ultraviolet Explorer (IUE) long wavelength low and high resolution spectra were obtained concurrently. The ground based spectra have shown an asymmetry with orbital phase in the H-alpha profile. The H-beta profiles were consistent with the same effect. One hemisphere showed excess emission and the other excess absorption, with a broad Gaussian emission component superposed upon the excess H-alpha line. An improved radial velocity curve, giving a better determined mass ratio and geometry was derived. This combined with the radii implied by the rotational broadening of the spectra, showed one component to be 80 to 90% filling the equilibrium Roche surface. The two-faced nature is, therfore, very likely due to mass transfer from the contact component impacting upon its companion. Low resolution ultraviolet data showed that the supposed cooler component is bluer than its companion. High resolution ultraviolet data taken during secondary eclipse showed Mg II emission strength which decreased more slowly than the area visible. The phase behavior of the low resolution data support the former situation, indicating traditional chromospheric activity.

Optical and UV spectroscopy of the peculiar RS CVn system, RT Lacertae

NASA Astrophysics Data System (ADS)

Huenemoerder, D. P.; Barden, S. C.

1985-11-01

Spectra in the H-alpha and H-beta regions of the peculiar double-lined RS CVn binary, RT Lacertae, were obtained in the fall of 1984. Limited International Ultraviolet Explorer (IUE) long wavelength low and high resolution spectra were obtained concurrently. The ground based spectra have shown an asymmetry with orbital phase in the H-alpha profile. The H-beta profiles were consistent with the same effect. One hemisphere showed excess emission and the other excess absorption, with a broad Gaussian emission component superposed upon the excess H-alpha line. An improved radial velocity curve, giving a better determined mass ratio and geometry was derived. This combined with the radii implied by the rotational broadening of the spectra, showed one component to be 80 to 90% filling the equilibrium Roche surface. The two-faced nature is, therfore, very likely due to mass transfer from the contact component impacting upon its companion. Low resolution ultraviolet data showed that the supposed cooler component is bluer than its companion. High resolution ultraviolet data taken during secondary eclipse showed Mg II emission strength which decreased more slowly than the area visible. The phase behavior of the low resolution data support the former situation, indicating traditional chromospheric activity.

Manifestation of the structure of heavy nuclei in their alpha decays

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

Adamian, G. G., E-mail: adamian@theor.jinr.ru; Antonenko, N. V.; Bezbakh, A. N.

2016-11-15

Low-lying one- and two-quasiparticle states of heavy nuclei are predicted. Alpha-decay chains, including those that proceed through isomeric states, are examined on the basis of the predicted properties of superheavy nuclei.

Structural characterization of novel L-galactose-containing oligosaccharide subunits of jojoba seed xyloglucans.

PubMed

Hantus, S; Pauly, M; Darvill, A G; Albersheim, P; York, W S

1997-10-28

Jojoba seed xyloglucan was shown to be a convenient source of biologically active xyloglucan oligosaccharides that contain both L- and D-galactosyl residues [E. Zablackis et al., Science, 272 (1996) 1808-1810]. Oligosaccharides were isolated by liquid chromatography of the mixture of oligosaccharides generated by treating jojoba seed xyloglucan with a beta-(1-->4)-endoglucanase. The purified oligosaccharides were reduced with NaBH4, converting them to oligoglycosyl alditol derivatives that were structurally characterized by a combination of mass spectrometry and 2-dimensional NMR spectroscopy. This analysis established that jojoba xyloglucan oligosaccharides contain the novel side-chain [alpha-L-Gal p-(1-->2)-beta-D-Galp-(1-->2)-alpha-D-Xyl p-(1-->6)-], which is structurally homologous to the fucose-containing side-chain [alpha-L-Fucp-(1-->2)-beta-D-Galp-(1-->2)-alpha-D-Xyl p-(1-->6)-] found in other biologically active xyloglucan oligosaccharides.

Molecular and isotopic analyses of the hydroxy acids, dicarboxylic acids, and hydroxydicarboxylic acids of the Murchison meteorite

NASA Astrophysics Data System (ADS)

Cronin, J. R.; Pizzarello, S.; Epstein, S.; Krishnamurthy, R. V.

1993-10-01

The hydroxymonocarboxylic acids, dicarboxylic acids, and hydroxydicarboxylic acids of the Murchison meteorite were analyzed as their tert-butyldimethylsilyl derivatives using combined gas chromatography-mass spectrometry. The hydroxydicarboxylic acids have not been found previously in meteorites. Each class of compounds is numerous with carbon chains up to C8 or C9 and many, if not all, chain and substitution position isomers represented at each carbon number. The alpha-hydroxycarboxylic acids and alpha-hydroxydicarboxylic acids correspond structurally to many of the known meteoritic alpha-aminocarboxylic acids and alpha-aminodicarboxylic acids, a fact that supports the proposal that a Strecker synthesis was involved in the formation of both classes of compounds. Isotopic analyses show these acids to be D-rich relative to terrestrial organic compounds, as expected; however, the hydroxy acids appear to be isotopically lighter than the amino acids with respect to both carbon and hydrogen.

Spectroscopic observations of X-ray selected late type stars

NASA Technical Reports Server (NTRS)

Takalo, L. O.

1988-01-01

A spectroscopic survey of nine X-ray selected late type stars was conducted. These stars are serendipitously discovered EINSTEIN X-ray sources, selected from two large x-ray surveys: the Columbia Astrophysical Laboratory survey (five stars) and the CFA Medium Sensitivity survey (four stars). Four of the Columbia survey stars were found to be short period binaries. The fifth was found to be an active single G dwarf. None of the Medium Sensitivity survey stars were found to be either binaries or active stars. Activity was measured by comparing the H-alpha and the CaII infrared triplet (8498, 8542) lines in these stars to the lines in inactive stars of similar spectral type. A correlation was found between the excess H-alpha lime emission and V sin(i) and between the excess H-alpha line emission and X-ray luminosity. No correlation was found between the infrared line emission and any other measured quantity.

Measurement of messenger RNA encoding the alpha-chain, polymeric immunoglobulin receptor, and J-chain in duodenal mucosa from dogs with and without chronic diarrhea by use of quantitative real-time reverse transcription-polymerase chain reaction assays.

PubMed

Peters, Iain R; Helps, Chris R; Calvert, Emma L; Hall, Edward J; Day, Michael J

2005-01-01

To examine the difference in expression of messenger RNA (mRNA) transcripts for polymeric immunoglobulin receptor (plgR), alpha-chain, and J-chain determined by use of quantitative real-time reverse transcription-polymerase chain reaction (QRT-PCR) assays in duodenal biopsy specimens obtained from dogs with and without chronic diarrhea. Biopsy specimens of the proximal portion of the duodenum were obtained endoscopically from 39 dogs evaluated because of chronic diarrhea (12 German Shepherd Dogs and 27 non-German Shepherd Dog breeds); specimens were also obtained from a control group of 7 dogs evaluated because of other gastrointestinal tract diseases and 2 dogs that were euthanatized as a result of nongastrointestinal tract disease. Dogs were anesthetized, and multiple mucosal biopsy specimens were obtained endoscopically at the level of the caudal duodenal flexure by use of biopsy forceps; in 2 control dogs, samples were obtained from the descending duodenum within 5 minutes of euthanasia. One-step QRT-PCR was used to quantify the level of expression of transcripts for the housekeeper gene glyceraldehyde-3-phosphate dehydrogenase, plgR, alpha-chain, and J-chain in duodenal mucosal tissue. There was no significant difference in the level of expression of any transcript among non-German Shepherd Dog breeds without diarrhea (control group), non-German Shepherd Dog breeds with chronic diarrhea, and German Shepherd Dogs with chronic diarrhea. Conclusions and Clinical Relevance-Results indicated that the susceptibility of German Shepherd Dogs to chronic diarrhea is not a result of simple failure of transcription of the key genes that encode molecules involved in mucosal IgA secretion.

Molecular basis for defect in Alix-binding by alternatively spliced isoform of ALG-2 (ALG-2DeltaGF122) and structural roles of F122 in target recognition.

PubMed

Inuzuka, Tatsutoshi; Suzuki, Hironori; Kawasaki, Masato; Shibata, Hideki; Wakatsuki, Soichi; Maki, Masatoshi

2010-08-06

ALG-2 (a gene product of PDCD6) belongs to the penta-EF-hand (PEF) protein family and Ca2+-dependently interacts with various intracellular proteins including mammalian Alix, an adaptor protein in the ESCRT system. Our previous X-ray crystal structural analyses revealed that binding of Ca2+ to EF3 enables the side chain of R125 to move enough to make a primary hydrophobic pocket (Pocket 1) accessible to a short fragment of Alix. The side chain of F122, facing a secondary hydrophobic pocket (Pocket 2), interacts with the Alix peptide. An alternatively spliced shorter isoform, designated ALG-2DeltaGF122, lacks Gly121Phe122 and does not bind Alix, but the structural basis of the incompetence has remained to be elucidated. We solved the X-ray crystal structure of the PEF domain of ALG-2DeltaGF122 in the Ca2+-bound form and compared it with that of ALG-2. Deletion of the two residues shortened alpha-helix 5 (alpha5) and changed the configuration of the R125 side chain so that it partially blocked Pocket 1. A wall created by the main chain of 121-GFG-123 and facing the two pockets was destroyed. Surprisingly, however, substitution of F122 with Ala or Gly, but not with Trp, increased the Alix-binding capacity in binding assays. The F122 substitutions exhibited different effects on binding of ALG-2 to other known interacting proteins, including TSG101 (Tumor susceptibility gene 101) and annexin A11. The X-ray crystal structure of the F122A mutant revealed that removal of the bulky F122 side chain not only created an additional open space in Pocket 2 but also abolished inter-helix interactions with W95 and V98 (present in alpha4) and that alpha5 inclined away from alpha4 to expand Pocket 2, suggesting acquirement of more appropriate positioning of the interacting residues to accept Alix. We found that the inability of the two-residue shorter ALG-2 isoform to bind Alix is not due to the absence of bulky side chain of F122 but due to deformation of a main-chain wall facing pockets 1 and 2. Moreover, a residue at the position of F122 contributes to target specificity and a smaller side chain is preferable for Alix binding but not favored to bind annexin A11.

Effect of low electric fields on alpha scintillation light yield in liquid argon

DOE PAGES

Agnes, P.; Albuquerque, I. F. M.; Alexander, T.; ...

2017-01-24

Measurements were made of scintillation light yield of alpha particles from themore » $$^{222}$$Rn decay chain within the DarkSide-50 liquid argon time projection chamber. Furthermore, the light yield was found to increase as the applied electric field increased, with alphas in a 200 V/cm electric field exhibiting a 2% increase in light yield compared to alphas in no field.« less

Lysine hydroxylation of collagen in a fibroblast cell culture system

NASA Technical Reports Server (NTRS)

Uzawa, Katsuhiro; Yeowell, Heather N.; Yamamoto, Kazushi; Mochida, Yoshiyuki; Tanzawa, Hideki; Yamauchi, Mitsuo

2003-01-01

The lysine (Lys) hydroxylation pattern of type I collagen produced by human fibroblasts in culture was analyzed and compared. Fibroblasts were cultured from normal human skin (NSF), keloid (KDF), fetal skin (FDF), and skin tissues of Ehlers-Danlos syndrome type VIA and VIB patients (EDS-VIA and -VIB). The type I collagen alpha chains with or without non-helical telopeptides were purified from the insoluble matrix and analyzed. In comparison with NSFs, KDF and FDF showed significantly higher Lys hydroxylation, particularly in the telopeptide domains of both alpha chains. Both EDS-VIA and -VIB showed markedly lower Lys hydroxylation in the helical domains of both alpha chains whereas that in the telopeptides was comparable with those of NSFs. A similar profile was observed in the tissue sample of the EDS-VIB patient. These results demonstrate that the Lys hydroxylation pattern is domain-specific within the collagen molecule and that this method is useful to characterize the cell phenotypes in normal/pathological connective tissues.

Alpha-beta T cells provide protection against lethal encephalitis in the murine model of VEEV infection

PubMed Central

Paessler, Slobodan; Yun, Nadezhda E.; Judy, Barbara M.; Dziuba, Natallia; Zacks, Michele A.; Grund, Anna H.; Frolov, Ilya; Campbell, Gerald A.; Weaver, Scott C.; Estes, D. Mark

2007-01-01

We evaluated the safety and immunogenicity of a chimeric alphavirus vaccine candidate in mice with selective immunodeficiencies. This vaccine candidate was highly attenuated in mice with deficiencies in the B and T cell compartments, as well as in mice with deficient gamma-interferon responsiveness. However, the level of protection varied among the strains tested. Wild type mice were protected against lethal VEEV challenge. In contrast, alpha/beta (αβ) TCR-deficient mice developed lethal encephalitis following VEEV challenge, while mice deficient in gamma/delta ( γδ) T cells were protected. Surprisingly, the vaccine potency was diminished by 50% in animals lacking interferon-gamma receptor alpha chain (R1)-chain and a minority of vaccinated immunoglobulin heavy chain-deficient (μMT) mice survived challenge, which suggests that neutralizing antibody may not be absolutely required for protection. Prolonged replication of encephalitic VEEV in the brain of pre-immunized mice is not lethal and adoptive transfer experiments indicate that CD3+ T cells are required for protection. PMID:17610927

A study of mass loss from the mid-ultraviolet spectrum of Alpha Cygni /A2 Ia/, Beta Orionis /B8 Ia/, and Eta Leonis /A0 Ib/

NASA Technical Reports Server (NTRS)

Lamers, H. J. G. L. M.; Stalio, R.; Kondo, Y.

1978-01-01

Results are presented for a study of mass loss from A and late-B supergiants based on high-resolution mid-UV spectra obtained with the echelle spectrograph of the Balloon-borne Ultraviolet Stellar Spectrometer. Spectra of Alpha Cyg, Beta Ori, Eta Leo, and Alpha Lyr are compared in selected wavelength regions; particular attention is given to previous observations of each star, the Mg II and Fe II resonance lines, lines due to other ions, and evidence for mass ejection. The results indicate that mass loss from late-B and A supergiants is variable, that a considerable fraction of envelope material is ejected in 'puffs', and that the puffs may be due to photospheric instabilities. A mass-loss rate of about 1 hundred-millionth of a solar mass per year is derived for Alpha Cyg and shown to be two orders of magnitude smaller than the value determined from the observed IR excess. This discrepancy is attributed to excess ionization in the envelope.

Linkage of interactions in sickle hemoglobin fiber assembly: inhibitory effect emanating from mutations in the AB region of the alpha-chain is annulled by a mutation at its EF corner.

PubMed

Sudha, Rajamani; Anantharaman, Lavanya; Sivaram, Mylavarapu V S; Mirsamadi, Neda; Choudhury, Devapriya; Lohiya, Nirmal K; Gupta, Rasik B; Roy, Rajendra P

2004-05-07

The AB and GH regions of the alpha-chain are located in spatial proximity and contain a cluster of intermolecular contact residues of the sickle hemoglobin (HbS) fiber. We have examined the role of dynamics of AB/GH region on HbS polymerization through simultaneous replacement of non-contact Ala(19) and Ala(21) of the AB corner with more flexible Gly or rigid alpha-aminoisobutyric acid (Aib) residues. The polymerization behavior of HbS with Aib substitutions was similar to the native HbS. In contrast, Gly substitutions inhibited HbS polymerization. Molecular dynamics simulation studies of alpha-chains indicated that coordinated motion of AB and GH region residues present in native (Ala) as well as in Aib mutant was disrupted in the Gly mutant. The inhibitory effect due to Gly substitutions was further explored in triple mutants that included mutation of an inter-doublestrand contact (alphaAsn(78) --> His or Gln) at the EF corner. Although the inhibitory effect of Gly substitutions in the triple mutant was unaffected in the presence of alphaGln(78), His at this site almost abrogated its inhibitory potential. The polymerization studies of point mutants (alphaGln(78) --> His) indicated that the inhibitory effect due to Gly substitutions in the triple mutant was synergistically compensated for by the polymerization-enhancing activity of His(78). Similar synergistic coupling, between alphaHis(78) and an intra-double-strand contact point (alpha16) mutation located in the AB region, was also observed. Thus, two conclusions are made: (i) Gly mutations at the AB corner inhibit HbS polymerization by perturbing the dynamics of the AB/GH region, and (ii) perturbations of AB region (through changes in dynamics of the AB/GH region or abolition of a specific fiber contact site) that influence HbS polymerization do so in concert with alpha78 site at the EF corner. The overall results provide insights about the interaction-linkage between distant regions of the HbS tetramer in fiber assembly.

Collagen type IV at the fetal-maternal interface.

PubMed

Oefner, C M; Sharkey, A; Gardner, L; Critchley, H; Oyen, M; Moffett, A

2015-01-01

Extracellular matrix proteins play a crucial role in influencing the invasion of trophoblast cells. However the role of collagens and collagen type IV (col-IV) in particular at the implantation site is not clear. Immunohistochemistry was used to determine the distribution of collagen types I, III, IV and VI in endometrium and decidua during the menstrual cycle and the first trimester of pregnancy. Expression of col-IV alpha chains during the reproductive cycle was determined by qPCR and protein localisation by immunohistochemistry. The structure of col-IV in placenta was examined using transmission electron microscopy. Finally, the expression of col-IV alpha chain NC1 domains and collagen receptors was localised by immunohistochemistry. Col-IV alpha chains were selectively up-regulated during the menstrual cycle and decidualisation. Primary extravillous trophoblast cells express collagen receptors and secrete col-IV in vitro and in vivo, resulting in the increased levels found in decidua basalis compared to decidua parietalis. A novel expression pattern of col-IV in the mesenchyme of placental villi, as a three-dimensional network, was found. NC1 domains of col-IV alpha chains are known to regulate tumour cell migration and the selective expression of these domains in decidua basalis compared to decidua parietalis was determined. Col-IV is expressed as novel forms in the placenta. These findings suggest that col-IV not only represents a structural protein providing tissue integrity but also influences the invasive behaviour of trophoblast cells at the implantation site. Copyright © 2014 Elsevier Ltd. All rights reserved.

The ability of lens alpha crystallin to protect against heat-induced aggregation is age-dependent

NASA Technical Reports Server (NTRS)

Horwitz, J.; Emmons, T.; Takemoto, L.; Spooner, B. S. (Principal Investigator)

1992-01-01

Alpha crystallin was prepared from newborn and aged bovine lenses. SDS-PAGE and tryptic peptide mapping demonstrated that both preparations contained only the alpha-A and alpha-B chains, with no significant contamination of other crystallins. Compared with alpha crystallin from the aged lens, alpha crystallin from the newborn lens was much more effective in the inhibition of beta L crystallin denaturation and precipitation induced in vitro by heat. Together, these results demonstrate that during the aging process, the alpha crystallins lose their ability to protect against protein denaturation, consistent with the hypothesis that the alpha crystallins play an important role in the maintenance of protein native structure in the intact lens.

Synthesis of 3 alpha,7 alpha,12 alpha,25-tetrahydroxy-5 beta-cholestan-24-one, an intermediate in the 25-hydroxylation pathway of cholic acid biosynthesis from cholesterol.

PubMed

Dayal, B; Tint, G S; Batta, A K; Shefer, S; Salen, G; Bose, A K; Pramanik, B N

1983-02-01

This paper describes the chemical synthesis of 3 alpha,7 alpha,12 alpha,25-tetrahydroxy-5 beta-cholestan-24-one via selective oxidation of 5 beta-cholestane-3 alpha,7 alpha,12 alpha, 24 xi,25-pentol with silver carbonate on celite. The structure of this 24-keto bile alcohol was confirmed by gas-liquid chromatography and mass spectrometry. Synthesis of this compound via pyridinium chlorochromate oxidation of the triacetoxy derivative of 5 beta-cholestane-3 alpha,7 alpha,12 alpha,24 xi,25-pentol followed by saponification further established its structure. 3 alpha,7 alpha,12 alpha,25-Tetrahydroxy-5 beta-cholestan-24-one was required for the in vivo and in vitro studies of side-chain oxidation and cleavage in the 25-hydroxylation pathway of cholic acid biosynthesis.

CD90-positive cells, an additional cell population, produce laminin {alpha}2 upon transplantation to dy{sup 3k}/dy{sup 3k} mice

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

Fukada, So-ichiro; Yamamoto, Yukiko; Segawa, Masashi

2008-01-01

Laminin {alpha}2 is a component of skeletal and cardiac muscle basal lamina. A defect of the laminin {alpha}2 chain leads to severe congenital muscular dystrophy (MDC1A) in humans and dy/dy mice. Myogenic cells including myoblasts, myotubes, and myofibers in skeletal muscle are a possible source of the laminin {alpha}2 chain, and myogenic cells are thus proposed as a cell source for congenital muscular dystrophy therapy. However, we observed production of laminin {alpha}2 in non-myogenic cells of normal mice, and we could enrich these laminin {alpha}2-producing cells in CD90{sup +} cell fractions. Intriguingly, the number of CD90{sup +} cells increased dramaticallymore » during skeletal muscle regeneration in mice. This fraction did not include myogenic cells but exhibited a fibroblast-like phenotype. Moreover, these cells were resident in skeletal muscle, not derived from bone marrow. Finally, the production of laminin {alpha}2 in CD90{sup +} cells was not dependent on fusion with myogenic cells. Thus, CD90{sup +} cells are a newly identified additional cell fraction that increased during skeletal muscle regeneration in vivo and could be another cell source for therapy for lama2-deficient muscular dystrophy.« less

T-cell receptor V sub. alpha. and C sub. alpha. alleles associated with multiple sclerosis and myasthenia gravis

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

Oksenberg, J.R.; Cavalli-Sforza, L.L.; Steinman, L.

1989-02-01

Polymorphic markers in genes encoding the {alpha} chain of the human T-cell receptor (TcR) have been detected by Southern blot analysis in Pss I digests. Polymorphic bands were observed at 6.3 and 2.0 kilobases (kb) with frequencies of 0.30 and 0.44, respectively, in the general population. Using the polymerase chain reaction (PCR) method, the authors amplified selected sequences derived from the full-length TcR {alpha} cDNA probe. These PcR products were used as specific probes to demonstrate that the 6.3-kb polymorphic fragment hybridizes to the variable (V)-region probe and the 2.0-kb fragment hybridizes to the constant (C)-region probe. Segregation of themore » polymorphic bands was analyzed in family studies. To look for associations between these markers and autoimmune diseases, the authors have studied the restriction fragment length polymorphism distribution of the Pss I markers in patients with multiple sclerosis, myasthenia gravis, and Graves disease. Significant differences in the frequency of the polymorphic V{sub {alpha}} and C{sub {alpha}} markers were identified between patients and healthy individuals.« less

Ultraviolet observations of alpha Aurigae from Copernicus

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

Dupree, A. K.

1975-08-01

Emission lines of L$alpha$ (1215.67 A) and O vi (1031.94 A) were detected in the spectroscopic binary $alpha$ Aur (Capella) with the Princeton experiment on Copernicus. Temperatures of the emitting regions are inferred to be in excess of 3times10$sup 5$ K. The temperature and emission measure are consistent with atmosphere is expanding with velocities approx.20 to 100 km s$sup -1$. Such expansion can lead to material within the binary system. The density of interstellar hydrogen inferred from absorption of stellar L$alpha$ appears to be approx.0.01 hydrogen atoms cm$sup -3$.

[Application of the polymerase chain reaction (PCR) in the diagnosis of Hb S-beta(+)-thalassemia].

PubMed

Harano, K; Harano, T; Kushida, Y; Ueda, S

1991-08-01

Isoelectric focusing of the hemolysate prepared from a two-year-old American black boy with microcytic hypochromia showed the presence of a high percentage (63.3%) of such Hb variant as Hb S, while the levels of Hb A, Hb F and Hb A2 were 20.0%, 12.7%, and 4.0%, respectively. The ratio of the non-alpha-chain to the alpha-chain of the biosynthesized globin chains was 0.49. The variant was identified as Hb S by amino acid analysis of the abnormal peptide (beta T-1) and digestion of DNA amplified by the polymerase chain reaction with enzyme Eco 81 I. This was further confirmed by DNA sequencing. DNA sequencing of a beta-gene without the beta s-mutation revealed a nucleotide change of T to C in the polyadenylation signal sequence AATAAA 3' to the beta-gene, resulting in beta(+)-thalassemia. These results are consistent with the existence of a beta s-gene and a beta(+)-thalassemia gene in trans.

Assembly of the epithelial Na+ channel evaluated using sucrose gradient sedimentation analysis.

PubMed

Cheng, C; Prince, L S; Snyder, P M; Welsh, M J

1998-08-28

Three subunits, alpha, beta, and gamma, contribute to the formation of the epithelial Na+ channel. To investigate the oligomeric assembly of the channel complex, we used sucrose gradient sedimentation analysis to determine the sedimentation properties of individual subunits and heteromultimers comprised of multiple subunits. When the alpha subunit was expressed alone, it first formed an oligomeric complex with a sedimentation coefficient of 11 S, and then generated a higher order multimer of 25 S. In contrast, individual beta and gamma subunits predominately assembled into 11 S complexes. We obtained similar results with expression in cells and in vitro. When we co-expressed beta with alpha or with alpha plus gamma, the beta subunit assembled into a 25 S complex. Glycosylation of the alpha subunit was not required for assembly into a 25 S complex. We found that the alpha subunit formed intra-chain disulfide bonds. Although such bonds were not required to generate an oligomeric complex, under nonreducing conditions the alpha subunit formed a complex that migrated more homogeneously at 25 S. This suggests that intra-chain disulfide bonds may stabilize the complex. These data suggest that the epithelial Na+ channel subunits form high order oligomeric complexes and that the alpha subunit contains the information that facilitates such formation. Interestingly, the ability of the alpha, but not the beta or gamma, subunit to assemble into a 25 S homomeric complex correlates with the ability of these subunits to generate functional channels when expressed alone.

Reversible addition-fragmentation chain transfer polymerization of N-isopropylacrylamide: a comparison between a conventional and a fast initiator.

PubMed

Bouchékif, Hassen; Narain, Ravin

2007-09-27

The reversible addition-fragmentation chain transfer (RAFT) polymerization of N-isopropylacrylamide (NIPAM) was studied to determine the reasons for deviation of experimental molecular weights to lower molecular weight at high monomer conversion when S-1-dodecyl-S-(alpha,alpha'-dimethyl-alpha' '-acetic acid)trithiocarbonate (CTAm) and S,S-bis(alpha,alpha'-dimethyl-alpha' '-acetic acid)trithiocarbonate (CTAd) were used as RAFT agents at 65 degrees C. For this purpose, experiments were performed in N,N'-dimethylformamide (DMF) at the NIPAM/CTA ratio of 200 with initiators capable of yielding fast and slow initiation, respectively by photochemical and thermal process, either at ambient temperature or at 65 degrees C. When the polymerization of NIPAM was conducted under these conditions with Irgacure-2959 (IRGC) as photoinitiator, a continuing supply of primary radicals by incremental initiator addition was required to achieve reasonably high conversion. This effect was also apparent by the loss of linearity of the first-order kinetic plot with a conventional initiator (4,4-azobis(4-cyanovaleric acid) (ACVA) as azo-initiator, 10h (t1/2) decomposition at 65 degrees C) indicating that steady-state concentration of the macroradical decreases significantly with the initiator consumption. Nevertheless, polymers with predictable number-average molecular weight Mn (i.e., based on [monomer]/([CTA] + [initiator]) ratio) and narrow polydispersities were obtained (PDIs < 1.2) with CTAm indicating that the process of chain growth was controlled. When CTAd was used, instead of CTAm, the polymers obtained were characterized by a larger polydispersity (1.2 < PDIs < 1.3). The so-called "living steady-state concentration" in chain equilibration together with the linear dependence of Mn vs conversion was observed only when the 200/1 NIPAM/CTA mixture in DMF was subjected to a permanent photoirradiation at 65 degrees C. With ACVA, the deviation of the experimentally measured molecular weights at high conversion was accounted for by the simultaneous self-initiated polymerization of NIPAM with the controlled process in the presence of CTA at 65 degrees C. Similar drift from the linear dependence Mn vs conversion was also observed at 65 degrees C when a significant number of low molecular weight polymer chains were generated intentionally by photodecomposition of IRGC.

Isolation and characterization at cholinergic nicotinic receptors of a neurotoxin from the venom of the Acanthophis sp. Seram death adder.

PubMed

Wickramaratna, Janith C; Fry, Bryan G; Loiacono, Richard E; Aguilar, Marie-Isabel; Alewood, Paul F; Hodgson, Wayne C

2004-07-15

The present study describes the isolation of the first neurotoxin (acantoxin IVa) from Acanthophis sp. Seram death adder venom and an examination of its activity at nicotinic acetylcholine receptor (nAChR) subtypes. Acantoxin IVa (MW 6815; 0.1-1.0 microM) caused concentration-dependent inhibition of indirect twitches (0.1 Hz, 0.2 ms, supramaximal V) and inhibited contractile responses to exogenous nicotinic agonists in the chick biventer cervicis nerve-muscle, confirming that this toxin is a postsynaptic neurotoxin. Acantoxin IVa (1-10 nM) caused pseudo-irreversible antagonism at skeletal muscle nAChR with an estimated pA2 of 8.36+/-0.17. Acantoxin IVa was approximately two-fold less potent than the long-chain (Type II) neurotoxin, alpha-bungarotoxin. With a pKi value of 4.48, acantoxin IVa was approximately 25,000 times less potent than alpha-bungarotoxin at alpha7-type neuronal nAChR. However, in contrast to alpha-bungarotoxin, acantoxin IVa completely inhibited specific [3H]-methyllycaconitine (MLA) binding in rat hippocampus homogenate. Acantoxin IVa had no activity at ganglionic nAChR, alpha4beta2 subtype neuronal nAChR or cytisine-resistant [3H]-epibatidine binding sites. While long-chain neurotoxin resistant [3H]-MLA binding in hippocampus homogenate requires further investigation, we have shown that a short-chain (Type I) neurotoxin is capable of fully inhibiting specific [3H]-MLA binding.

Hydrophobic core malleability of a de novo designed three-helix bundle protein.

PubMed

Walsh, S T; Sukharev, V I; Betz, S F; Vekshin, N L; DeGrado, W F

2001-01-12

De novo protein design provides a tool for testing the principles that stabilize the structures of proteins. Recently, we described the design and structure determination of alpha(3)D, a three-helix bundle protein with a well-packed hydrophobic core. Here, we test the malleability and adaptability of this protein's structure by mutating a small, Ala residue (A60) in its core to larger, hydrophobic side-chains, Leu and Ile. Such changes introduce strain into the structures of natural proteins, and therefore generally destabilize the native state. By contrast, these mutations were slightly stabilizing ( approximately 1.5 kcal mol(-1)) to the tertiary structure of alpha(3)D. The value of DeltaC(p) for unfolding of these mutants was not greatly affected relative to wild-type, indicating that the change in solvent accessibility for unfolding was similar. However, two-dimensional heteronuclear single quantum coherence spectra indicate that the protein adjusts to the introduction of steric bulk in different ways. A60L-alpha(3)D showed serious erosion in the dispersion of both the amide backbone as well as the side-chain methyl chemical shifts. By contrast, A60I-alpha(3)D showed excellent dispersion of the backbone resonances, and selective changes in dispersion of the aliphatic side-chains proximal to the site of mutation. Together, these data suggest that alpha(3)D, although folded into a unique three-dimensional structure, is nevertheless more malleable and flexible than most natural, native proteins. Copyright 2001 Academic Press.

From Beamline to Scanner with 225Ac

NASA Astrophysics Data System (ADS)

Robertson, Andrew K. H.; Ramogida, Caterina F.; Kunz, Peter; Rodriguez-Rodriguez, Cristina; Schaffer, Paul; Sossi, Vesna

2016-09-01

Due to the high linear energy transfer and short range of alpha-radiation, targeted radiation therapy using alpha-emitting pharmaceuticals that successfully target small disease clusters will kill target cells with limited harm to healthy tissue, potentially treating the most aggressive forms of cancer. As the parent of a decay chain with four alpha- and two beta-decays, 225Ac is a promising candidate for such a treatment. However, this requires retention of the entire decay chain at the target site, preventing the creation of freely circulating alpha-emitters that reduce therapeutic effect and increase toxicity to non-target tissues. Two major challenges to 225Ac pharmaceutical development exist: insufficient global supply, and the difficulty of preventing toxicity by retaining the entire decay chain at the target site. While TRIUMF works towards large-scale (C i amounts) production of 225Ac, we already use our Isotope Separation On-Line facility to provide small (< 1 mCi) quantities for in-house chemistry and imaging research that aims to improve and assess 225Ac radiopharmaceutical targeting. This presentation provides an overview of this research program and the journey of 225Ac from the beamline to the scanner. This research is funded by the Natural Sciences and Engineering Research Council of Canada.

Serum retinol, alpha-tocopherol, and beta-carotene levels are not altered by excess ingestion of diacylglycerol-containing plant sterol esters.

PubMed

Saito, Shinichiro; Tomonobu, Kazuichi; Kudo, Naoto; Shiiba, Daisuke; Hase, Tadashi; Tokimitsu, Ichiro

2006-01-01

Diacylglycerol (DAG) suppresses the postprandial increase in serum triglycerides, and has antiobesity effects. On the other hand, plant sterol esters (PSE) lower serum cholesterol levels in hypercholesterolemia. Thus, DAG-containing PSE (PSE/DAG) would be expected to maintain an appropriate serum cholesterol level and decrease the risk of arteriosclerotic disorders. Several recent studies, however, report negative effects of PSE on serum fat-soluble (pro)vitamin levels. The objective of this study was to investigate the effect of PSE/DAG on serum retinol, beta-carotene, and alpha-tocopherol levels using a threefold excess of the effective dose obtained in our previous study. A randomized placebo-controlled double-blind parallel study was performed in healthy and mildly hypercholesterolemic subjects, in which the subjects ingested 1.2 g PSE/30 g DAG for 2 weeks in the form of mayonnaise-type products. Triacylglycerol (TAG) mayonnaise was used as a control. There were no subjective adverse effects or changes in serum retinol, alpha-tocopherol, and beta-carotene levels, abdominal symptoms, hematologic values, or blood biochemical values. Ingestion of a threefold excess of PSE/DAG for 2 weeks had no adverse effects compared to ingestion of conventional TAG mayonnaise. Copyright 2006 S. Karger AG, Basel.

Herschel Discovery of a New Class of Cold, Faint Debris Discs

NASA Technical Reports Server (NTRS)

Eiroal, C.; Marshall, J. P.; Mora, A.; Krivov, A. V.; Montesinos, B.; Absil, O.; Ardila, D.; Arevalo, M.; Augereau, J.-Ch.; Bayo, A.;

Editing an α-globin enhancer in primary human hematopoietic stem cells as a treatment for β-thalassemia.

PubMed

Mettananda, Sachith; Fisher, Chris A; Hay, Deborah; Badat, Mohsin; Quek, Lynn; Clark, Kevin; Hublitz, Philip; Downes, Damien; Kerry, Jon; Gosden, Matthew; Telenius, Jelena; Sloane-Stanley, Jackie A; Faustino, Paula; Coelho, Andreia; Doondeea, Jessica; Usukhbayar, Batchimeg; Sopp, Paul; Sharpe, Jacqueline A; Hughes, Jim R; Vyas, Paresh; Gibbons, Richard J; Higgs, Douglas R

2017-09-04

β-Thalassemia is one of the most common inherited anemias, with no effective cure for most patients. The pathophysiology reflects an imbalance between α- and β-globin chains with an excess of free α-globin chains causing ineffective erythropoiesis and hemolysis. When α-thalassemia is co-inherited with β-thalassemia, excess free α-globin chains are reduced significantly ameliorating the clinical severity. Here we demonstrate the use of CRISPR/Cas9 genome editing of primary human hematopoietic stem/progenitor (CD34+) cells to emulate a natural mutation, which deletes the MCS-R2 α-globin enhancer and causes α-thalassemia. When edited CD34+ cells are differentiated into erythroid cells, we observe the expected reduction in α-globin expression and a correction of the pathologic globin chain imbalance in cells from patients with β-thalassemia. Xenograft assays show that a proportion of the edited CD34+ cells are long-term repopulating hematopoietic stem cells, demonstrating the potential of this approach for translation into a therapy for β-thalassemia.β-thalassemia is characterised by the presence of an excess of α-globin chains, which contribute to erythrocyte pathology. Here the authors use CRISP/Cas9 to reduce α-globin expression in hematopoietic precursors, and show effectiveness in xenograft assays in mice.

New acceptor-bridge-donor strategy for enhancing NLO response with long-range excess electron transfer from the NH2...M/M3O donor (M = Li, Na, K) to inside the electron hole cage C20F19 acceptor through the unusual σ chain bridge (CH2)4.

PubMed

Bai, Yang; Zhou, Zhong-Jun; Wang, Jia-Jun; Li, Ying; Wu, Di; Chen, Wei; Li, Zhi-Ru; Sun, Chia-Chung

2013-04-04

Using the strong electron hole cage C20F19 acceptor, the NH2...M/M3O (M = Li, Na, and K) complicated donors with excess electron, and the unusual σ chain (CH2)4 bridge, we construct a new kind of electride molecular salt e(-)@C20F19-(CH2)4-NH2...M(+)/M3O(+) (M = Li, Na, and K) with excess electron anion inside the hole cage (to be encapsulated excess electron-hole pair) serving as a new A-B-D strategy for enhancing nonlinear optical (NLO) response. An interesting push-pull mechanism of excess electron generation and its long-range transfer is exhibited. The excess electron is pushed out from the (super)alkali atom M/M3O by the lone pair of NH2 in the donor and further pulled inside the hole cage C20F19 acceptor through the efficient long σ chain (CH2)4 bridge. Owing to the long-range electron transfer, the new designed electride molecular salts with the excess electron-hole pair exhibit large NLO response. For the e(-)@C20F19-(CH2)4-NH2...Na(+), its large first hyperpolarizability (β0) reaches up to 9.5 × 10(6) au, which is about 2.4 × 10(4) times the 400 au for the relative e(-)@C20F20...Na(+) without the extended chain (CH2)4-NH2. It is shown that the new strategy is considerably efficient in enhancing the NLO response for the salts. In addition, the effects of different bridges and alkali atomic number on β0 are also exhibited. Further, three modulating factors are found for enhancing NLO response. They are the σ chain bridge, bridge-end group with lone pair, and (super)alkali atom. The new knowledge may be significant for designing new NLO materials and electronic devices with electrons inside the cages. They may also be the basis of establishing potential organic chemistry with electron-hole pair.

Definition of the mutation responsible for maple syrup urine disease in Poll Shorthorns and genotyping Poll Shorthorns and Poll Herefords for maple syrup urine disease alleles.

PubMed

Dennis, J A; Healy, P J

1999-08-01

The organisation of the E1alpha subunit of bovine branched-chain alpha-keto acid dehydrogenase gene was established. c DNA was cloned from Poll Shorthorn x Poll Hereford calves affected with Maple Syrup Urine Disease to identify the mutation responsible for the disease in Poll Shorthorns. Clones containing the c DNA sequences inherited from the Poll Shorthorn sire of the affected calves were identified. Paternal clones were sequenced and a cytidine to thymidine transition was found at nucleotide 1380. The mutation is predicted to substitute leucine in place of a highly conserved proline at codon 372. A polymerase chain reaction procedure was developed for detection of the 1380C-->T mutation in genomic DNA. Three Poll Shorthorn parents of affected calves and three affected Poll Shorthorn x Poll Hereford calves were heterozygous and an affected Poll Shorthorn calf was homozygous for this mutation. An improved polymerase chain reaction procedure was also devised to genotype Poll Herefords for the 248C-->T mutation. The procedures will facilitate disease prevention programs and assist in differential diagnosis of conditions in new-born calves that present with a rapid onset of progressive neurological disease and are characterised histologically by 'status spongiosus'. Maple Syrup Urine Disease (MSUD) is an autosomal recessive defect reported in humans (Danner and Elsas 1989), and in Poll Hereford (PH) and Poll Shorthorn (PS) calves (Harper et al 1986, Healy et al 1992). The clinical, biochemical and pathological manifestations of the disease are identical in the two breeds of cattle, and are characterised by the rapid onset of progressive neurological disease, leading to death within a few days of birth. The disease is caused by a deficiency of activity of the mitochondrial enzyme branched-chain alpha-keto acid dehydrogenase (BCKADH). This deficiency leads to elevated concentrations, in blood and tissues, of branched chain alpha-keto acids and their precursors, the branched chain amino acids, valine, leucine and isoleucine. BCKADH consists of four subunits E1alpha, E1beta, E2 and E3 that are encoded by separate genes, and MSUD may result from deficiency of any of the subunits. In PH s, the disease in caused by premature termination of translation, of the E1alpha subunit, that is induced by a cytidine to thymidine transition exon 2 (248C-->T), that converts the glutamine codon -6 to a stop codon (Q-6ST; Zhang et al 1990). We have shown that MSUD -affected PSxPH calves are heterozygous at the PH locus, illustrating molecular heterogeneity exists for bovine MSUD (Healy and Dennis 1994a). The fact that these crossbred calves are affected, indicates the PS, like the PH mutation, resides in the E1alpha subunit. Copyright 1999 Harcourt Publishers Ltd.

Cykotine mRNA expression in mouse retina after laser injury by reverse transcriptase-polymerase chain reaction (RT-PCR)

NASA Astrophysics Data System (ADS)

Schuschereba, Steven T.; Bowman, Phillip D.; Ujimore, Veronica; Hoxie, Stephen W.; Pizarro, Jose M.; Cross, Michael E.; Lund, David J.

1996-04-01

The purpose of this study was to identify cytokines produced by the retina after laser injury. With the aid of a scanning laser ophthalmoscope (SLO), right eyes of mice received lesions from a continuous wave argon laser. Left eyes served as unirradiated controls. At 2, 4, 6, 12, 24, and 48 hr after laser irradiation groups of 3 mice were euthanized and retinas fixed for histology or isolated for RNA. Messenger RNA (mRNA) was reverse-transcribed into complementary DNA (cDNA) and subjected to polymerase chain reaction for the following cytokines: tumor necrosis factor-(alpha) (TNF-(alpha) ), interleukin-1(alpha) /(Beta) (IL- 1(alpha) /(Beta) ), interleukin-6 (IL-6), transforming growth factor-(Beta) 1 (TGF- (Beta) 1), macrophage colony stimulating factor (M-CSF), inducible nitric oxide synthase (iNOS), and glyceraldehyde 3-phosphate dehydrogenase (G3PDH). Histologically, lesions were confined to the photoreceptors, retinal pigment epithelium, and choroid. In laser-injured retinas, mRNA levels were elevated for IL-1(alpha) , TGF-(Beta) 1, iNOS, and G3PDH, but not TNF-(alpha) , IL-1(Beta) , or IL-6. It appears that the retina, in response to laser injury, upregulates a select number of cytokines in a time-course dependent fashion.

Fusion protein of CDR mimetic peptide with Fc inhibit TNF-alpha induced cytotoxicity.

PubMed

Qin, Weisong; Feng, Jiannan; Li, Yan; Lin, Zhou; Shen, Beifen

2006-02-01

The variable regions of antibodies play central roles in the binding with antigens. Based on the model of a tumour necrosis factor-alpha (TNF-alpha) neutralizing monoclonal antibody (named as Z12) with TNF-alpha, heavy chain CDR2 (HCDR2) and light chain CDR3 (LCDR3) of Z12 were found to be the most responsible to bind with TNF-alpha. A mimetic peptide (PT) was designed based on the sequence derived from HCDR2 and LCDR3. Fusion protein PT-Fc was constructed by linking PT with Fc of human IgG1 through a flexible linker (GGGGGS). The primary structural characteristics of Fc and PT-Fc were analyzed, including the flexibility, hydrophilicity and epitopes. It was demonstrated that PT and Fc in the fusion protein possessed bio-function properly and non-interfering with each other. Furthermore, PT-Fc was expressed in Escherichia coli by fusion with thioredoxin (Trx). After trx-PT-Fc was cleaved with recombinant enterokinase, PT-Fc was obtained. The results of in vitro cytotoxic assays showed that both PT and PT-Fc could efficiently inhibit TNF-alpha induced apoptosis on L929 cells. At the same micromole concentration, the inhibition activity of PT-Fc was significantly higher than PT.

Integrins (alpha7beta1) in muscle function and survival. Disrupted expression in merosin-deficient congenital muscular dystrophy.

PubMed Central

Vachon, P H; Xu, H; Liu, L; Loechel, F; Hayashi, Y; Arahata, K; Reed, J C; Wewer, U M; Engvall, E

1997-01-01

Mutations in genes coding for dystrophin, for alpha, beta, gamma, and delta-sarcoglycans, or for the alpha2 chain of the basement membrane component merosin (laminin-2/4) cause various forms of muscular dystrophy. Analyses of integrins showed an abnormal expression and localization of alpha7beta1 isoforms in myofibers of merosin-deficient human patients and mice, but not in dystrophin-deficient or sarcoglycan-deficient humans and animals. It was shown previously that skeletal muscle fibers require merosin for survival and function (Vachon, P.H., F. Loechel, H. Xu, U.M. Wewer, and E. Engvall. 1996. J. Cell Biol. 134:1483-1497). Correction of merosin deficiency in vitro through cell transfection with the merosin alpha2 chain restored the normal localization of alpha7beta1D integrins as well as myotube survival. Overexpression of the apoptosis-suppressing molecule Bcl-2 also promoted the survival of merosin-deficient myotubes, but did not restore a normal expression of alpha7beta1D integrins. Blocking of beta1 integrins in normal myotubes induced apoptosis and severely reduced their survival. These findings (a) identify alpha7beta1D integrins as the de facto receptors for merosin in skeletal muscle; (b) indicate a merosin dependence for the accurate expression and membrane localization of alpha7beta1D integrins in myofibers; (c) provide a molecular basis for the critical role of merosin in myofiber survival; and (d) add new insights to the pathogenesis of neuromuscular disorders. PMID:9312189

Smad, but not MAPK, pathway mediates the expression of type I collagen in radiation induced fibrosis

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

Yano, Hiroyuki; Division of Radioisotope Research, Department of Research Support, Research Promotion Project, Oita University, 1-1 Idaigaoka Hasama-machi, Yufu, Oita 879-5593; Hamanaka, Ryoji

Highlights: Black-Right-Pointing-Pointer We examine how radiation affects the expression level and signal pathway of collagen. Black-Right-Pointing-Pointer TGF-{beta}1 mRNA is elevated earlier than those of collagen genes after irradiation. Black-Right-Pointing-Pointer Smad pathway mediates the expression of collagen in radiation induced fibrosis. Black-Right-Pointing-Pointer MAPK pathways are not affected in the expression of collagen after irradiation. -- Abstract: Radiation induced fibrosis occurs following a therapeutic or accidental radiation exposure in normal tissues. Tissue fibrosis is the excessive accumulation of collagen and other extracellular matrix components. This study investigated how ionizing radiation affects the expression level and signal pathway of type I collagen. Realmore » time RT-RCR showed that both {alpha}1and {alpha}2 chain of type I collagen mRNA were elevated from 48 h after irradiation with 10 Gy in NIH3T3 cells. The relative luciferase activities of both genes and type I collagen marker were elevated at 72 h. TGF-{beta}1 mRNA was elevated earlier than those of type I collagen genes. A Western blot analysis showed the elevation of Smad phosphorylation at 72 h. Conversely, treatment with TGF-{beta} receptor inhibitor inhibited the mRNA and relative luciferase activity of type I collagen. The phosphorylation of Smad was repressed with the inhibitor, and the luciferase activity was cancelled using a mutant construct of Smad binding site of {alpha}2(I) collagen gene. However, the MAPK pathways, p38, ERK1/2 and JNK, were not affected with specific inhibitors or siRNA. The data showed that the Smad pathway mediated the expression of type I collagen in radiation induced fibrosis.« less

Production of reactive oxygen species in brain mitochondria: contribution by electron transport chain and non-electron transport chain sources.

PubMed

Adam-Vizi, Vera

2005-01-01

Overwhelming evidence has accumulated indicating that oxidative stress is a crucial factor in the pathogenesis of neurodegenerative diseases. The major site of production of superoxide, the primary reactive oxygen species (ROS), is considered to be the respiratory chain in the mitochondria, but the exact mechanism and the precise location of the physiologically relevant ROS generation within the respiratory chain have not been disclosed as yet. Studies performed with isolated mitochondria have located ROS generation on complex I and complex III, respectively, depending on the substrates or inhibitors used to fuel or inhibit respiration. A more "physiological" approach is to address ROS generation of in situ mitochondria, which are present in their normal cytosolic environment. Hydrogen peroxide formation in mitochondria in situ in isolated nerve terminals is enhanced when complex I, complex III, or complex IV is inhibited. However, to induce a significant increase in ROS production, complex III and complex IV have to be inhibited by >70%, which raises doubts as to the physiological importance of ROS generation by these complexes. In contrast, complex I inhibition to a small degree is sufficient to enhance ROS generation, indicating that inhibition of complex I by approximately 25-30% observed in postmortem samples of substantia nigra from patients suffering from Parkinson's disease could be important in inducing oxidative stress. Recently, it has been described that a key Krebs cycle enzyme, alpha-ketoglutarate dehydrogenase (alpha-KGDH), is also able to produce ROS. ROS formation by alpha-KGDH is regulated by the NADH/NAD+ ratio, suggesting that this enzyme could substantially contribute to generation of oxidative stress due to inhibition of complex I. As alpha-KGDH is not only a generator but also a target of ROS, it is proposed that alpha-KGDH is a key factor in a vicious cycle by which oxidative stress is induced and promoted in nerve terminals.

Role of microsomal retinol/sterol dehydrogenase-like short-chain dehydrogenases/reductases in the oxidation and epimerization of 3alpha-hydroxysteroids in human tissues.

PubMed

Belyaeva, Olga V; Chetyrkin, Sergei V; Clark, Amy L; Kostereva, Natalia V; SantaCruz, Karen S; Chronwall, Bibie M; Kedishvili, Natalia Y

2007-05-01

Allopregnanolone (ALLO) and androsterone (ADT) are naturally occurring 3alpha-hydroxysteroids that act as positive allosteric regulators of gamma-aminobutyric acid type A receptors. In addition, ADT activates nuclear farnesoid X receptor and ALLO activates pregnane X receptor. At least with respect to gamma-aminobutyric acid type A receptors, the biological activity of ALLO and ADT depends on the 3alpha-hydroxyl group and is lost upon its conversion to either 3-ketosteroid or 3beta-hydroxyl epimer. Such strict structure-activity relationships suggest that the oxidation or epimerization of 3alpha-hydroxysteroids may serve as physiologically relevant mechanisms for the control of the local concentrations of bioactive 3alpha-hydroxysteroids. The exact enzymes responsible for the oxidation and epimerization of 3alpha-hydroxysteroids in vivo have not yet been identified, but our previous studies showed that microsomal nicotinamide adenine dinucleotide-dependent short-chain dehydrogenases/reductases (SDRs) with dual retinol/sterol dehydrogenase substrate specificity (RoDH-like group of SDRs) can oxidize and epimerize 3alpha-hydroxysteroids in vitro. Here, we present the first evidence that microsomal nicotinamide adenine dinucleotide-dependent 3alpha-hydroxysteroid dehydrogenase/epimerase activities are widely distributed in human tissues with the highest activity levels found in liver and testis and lower levels in lung, spleen, brain, kidney, and ovary. We demonstrate that RoDH-like SDRs contribute to the oxidation and epimerization of ALLO and ADT in living cells, and show that RoDH enzymes are expressed in tissues that have microsomal 3alpha-hydroxysteroid dehydrogenase/epimerase activities. Together, these results provide further support for the role of RoDH-like SDRs in human metabolism of 3alpha-hydroxysteroids and offer a new insight into the enzymology of ALLO and ADT inactivation.

A new stable alpha chain variant: Hb Basel [alpha14(A12)Trp-->Leu (alpha1)].

PubMed

Hergersberg, Martin; Brunner-Agten, Saskia; Kühne, Thomas; Paulussen, Michael; Huber, Andreas R

2010-06-01

We describe a heterozygosity for a new missense mutation on the alpha1-globin gene of an 18-year-old woman of Portuguese ancestry with severe hypochromic anemia and iron deficiency. Hemoglobin (Hb) analysis by high performance liquid chromatography (HPLC) found a prominent peak constituting about 12% of total Hb. Sequencing of the globin genes of the index patient found the mutation alpha14(A12)Trp-->Leu (alpha1), HBA1:c.44G

Serum Free Light Chains in Neoplastic Monoclonal Gammopathies: Relative Under-Detection of Lambda Dominant Kappa/Lambda Ratio, and Underproduction of Free Lambda Light Chains, as Compared to Kappa Light Chains, in Patients With Neoplastic Monoclonal Gammopathies.

PubMed

Lee, Won Sok; Singh, Gurmukh

2018-07-01

Quantitative evaluation of serum free light chains is recommended for the work up of monoclonal gammopathies. Immunoglobulin light chains are generally produced in excess of heavy chains. In patients with monoclonal gammopathy, κ/λ ratio is abnormal less frequently with lambda chain lesions. This study was undertaken to ascertain if the levels of overproduction of the two light chain types and their detection rates are different in patients with neoplastic monoclonal gammopathies. Results of serum protein electrophoresis (SPEP), serum protein immunofixation electrophoresis (SIFE), urine protein electrophoresis (UPEP), urine protein immunofixation electrophoresis (UIFE), and serum free light chain assay (SFLCA) in patients with monoclonal gammopathies were examined retrospectively. The κ/λ ratios were appropriately abnormal more often in kappa chain lesions. Ratios of κ/λ were normal in about 25% of patients with lambda chain lesions in whom free homogenous lambda light chains were detectable in urine. An illustrative case suggests underproduction of free lambda light chains, in some instances. The lower prevalence of lambda dominant κ/λ ratio in lesions with lambda light chains is estimated to be due to relative under-detection of lambda dominant κ/λ ratio in about 25% of the patients and because lambda chains are not produced in as much excess of heavy chains as are kappa chains, in about 5% of the patients. The results question the medical necessity and clinical usefulness of the serum free light chain assay. UPEP/UIFE is under-utilized.

Primary structure of the hemoglobins from Sphenodon (Sphenodon punctatus, Tuatara, Rynchocephalia). Evidence for the expression of alpha D-gene.

PubMed

Abbasi, A; Wells, R M; Brittain, T; Braunitzer, G

1988-08-01

Sphenodon is the sole representative of the "beakhead" reptiles which were widely distributed during the Triassic period before the spectacular rise of dinosaurs. Sphenodon punctatus is the only survivor ("living fossil") of this period. The morphological features of Sphenodon are remarkably conservative and differ little from reptiles living 200 million years ago. In the present paper the determination of the primary structure of the tetrameric hemoglobins is described: three components are identified: hemoglobin A' (alpha A2 beta II2), hemoglobin A (alpha A2 beta I2) and hemoglobin D (alpha D2 beta II2). The components were characterized electrophoretically, the four different peptide chains were characterized by Triton electrophoresis as well as by high-performance liquid chromatography. The hemoglobins and--under dissociating conditions--also the chains, were isolated on columns of cellulose ion exchangers. Sequence determination was carried out after cleavage of the individual chains with trypsin and after a specific chemical cleavage of the Asp-Pro bond. For sequence determination the film technique and gas-phase method were employed. The data are compared with the sequence of the human hemoglobin, and interpretations of the amino-acid sequences are given. Particularly notable is the evidence of hemoglobin D: this hemoglobin (alpha D2 beta II2) is found only in birds, and in two cases in turtles. However, this component is not found in other reptiles. The results make possible an interpretation of the relatively high oxygen affinity and explain the lack of cooperativity (myoglobin properties) of these tetrameric hemoglobins.

Inhibitory properties of the antibothropic complex from the South American opossum (Didelphis marsupialis) serum.

PubMed

Neves-Ferreira, A G; Perales, J; Ovadia, M; Moussatché, H; Domont, G B

1997-06-01

The South American opossum Didelphis marsupialis is known to be highly resistant to snake envenomation. In this paper it is shown that the opossum serum inhibits haemorrhage induced by both Crotalinae and Viperinae venoms. Tested against Bothrops jararaca (jararaca) venom, the antibothropic complex (ABC) isolated from the opossum serum was at least six times more antihaemorrhagic than the commercial antivenom. ABC showed no proteolytic activity by itself and was not hydrolysed by the venom. It inhibited the hydrolysis of casein by B. jararaca venom, but did not inhibit its hydrolytic activities upon N alpha-benzoyl-L-arginine ethyl ester (BAEE) and N alpha-benzoyl-DL-arginine p-nitroanilide (BAPNA). The inhibitor did not interfere with trypsin and bacterial collagenase activities on BAPNA and N-(3-[2-furyl]acryloyl)-Leu-Gly-Pro-Ala (FALGPA), respectively. It reduced chymotrypsin hydrolysis of N-acetyl-L-tyrosine ethyl ester (ATEE) because ABC is also a substrate for this enzyme. By sodium dodecyl sulfate-polyacrylamide gel electrophoresis, B. jararaca venom preferentially degraded fibrinogen A alpha-chain and fibrin alpha-chain. Tested on extracellular matrix proteins, the venom hydrolysed collagen IV, gelatins I and V, laminin and fibronectin, besides depolimerizing collagen I alpha-chain dimers. Fibrillar collagen V was not digested. These hydrolyses were inhibited by ABC and by EDTA. Our results show that the antibothropic complex is a venom metalloproteinase inhibitor, which could, at least partially, account for its antihaemorrhagic activity. Electrophoretic evidence indicated non-covalent complex formation between the antihaemorrhagic factor and component(s) of B. jararaca venom.

Different functional modulation by heterotropic ligands (2,3-diphosphoglycerate and chlorides) of the two haemoglobins from fallow-deer (Dama dama).

PubMed

Angeletti, M; Pucciarelli, S; Priori, A M; Canofeni, S; Barra, D; Fioretti, E; Coletta, M

2001-02-01

Two haemoglobin components have been identified and purified from fallow-deer (Dama dama) erythrocytes. They are present in similar amounts and the two tetrameric molecules share the same alpha chain, while two different beta chains are detected in the two components. The beta chains differ by 14 residues, even though they both have 145 amino-acid residues, which account for a molecular mass of 16,023 and 16,064 Da, respectively, while alpha chain has 141 residues, yielding a molecular mass of 15,142 Da. Compared with human Hb, the N-terminal region of both beta chains shows deletion of Val beta 1 and the replacement of His beta 2 by a methionyl residue, a modification which is common to most ruminant haemoglobins. Although both isolated components show a low intrinsic affinity for oxygen, meaningful differences between the two haemoglobins have been found with respect to the effect of heterotropic effectors, such as 2,3-diphosphoglycerate and chloride ions. In view of the high sequence homology between the two components, the different effect of heterotropic ligands has been tentatively correlated to possible localized structural variations between beta chains of the two haemoglobin components.

Mass spectral analysis of long chain alkyl aromatic compounds synthesized from alpha-olefin alkylation

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

Cheng, M.T.; Hudson, J.D.

1994-12-31

Long chain alkyl aromatic compounds are important petrochemicals with many applications. They are generally synthesized by alkylating the corresponding aromatic nucleus. In this report, the authors will describe the mass spectral fragmentation of alkylphenols and alkylsalicylates.

Role of Fetuin-A in Breast Tumor Cell Growth

DTIC Science & Technology

2009-03-01

UniRef100_Q0IIK2 289 390 1.4 Transferrin UniRef100_Q9GLY6 18 24 1.3 Inter- alpha -trypsin inhibitor heavy chain2 UniRef100_Q3SZ57 25 33 1.3 Alpha - fetoprotein ...the first time that cell attachment is mediated only by fetuin-A and not its major contaminant which is alpha -2-macroglobulin. We also show both...fetuin-A and alpha -2-macroglobulin can activate PI3 kinase/Akt. 15. SUBJECT TERMS Mammary tumors;fetuin-A;Polyoma middle T antigen; genotyping

Tumour necrosis factor-alpha polymorphism as one of the complex inherited factors in pemphigus.

PubMed Central

Torzecka, Jolanta Dorota; Narbutt, Joanna; Sysa-Jedrzejowska, Anna; Borowiec, Maciej; Ptasinska, Anetta; Woszczek, Grzegorz; Kowalski, Marek L

2003-01-01

The aim of our study was to analyse a significance of tumour necrosis factor (TNF)-alpha promoter gene polymorphisms in relation to the HLA-DR locus in genetic predisposition to pemphigus. TNF-alpha gene polymorphisms in position -238 and -308 were identified using a modified polymerase chain reaction-restriction fragment length polymorphism method in 53 patients with pemphigus (38 with pemphigus vulgaris, 15 with pemphigus foliaceus) and 87 healthy controls. The HLA-DRB1 locus was typed using the polymerase chain reaction SSO method in all the patients and 152 population controls. Carriers of the TNF-alpha polymorphic -308 A allele were found to be more frequent in the pemphigus foliaceus group in comparison with the control group (odds ratio (OR) = 8.12; p = 0.0005). A significant association between HLA-DRB1*04 (OR = 3.86; pcor = 0.0001) and DRB1*14 (OR = 8.4; pcor = 0.0001) and pemphigus vulgaris was found. In this group of patients a decreased frequency of HLA-DRB1*07 (OR = 0.08; pcor = 0.006) was also identified. We have shown for the first time a positive association of TNF-alpha polymorphism in position -308 with pemphigus foliaceus. PMID:14760938

Tumour necrosis factor-alpha polymorphism as one of the complex inherited factors in pemphigus.

PubMed

Torzecka, Jolanta Dorota; Narbutt, Joanna; Sysa-Jedrzejowska, Anna; Borowiec, Maciej; Ptasinska, Anetta; Woszczek, Grzegorz; Kowalski, Marek L

2003-10-01

The aim of our study was to analyse a significance of tumour necrosis factor (TNF)-alpha promoter gene polymorphisms in relation to the HLA-DR locus in genetic predisposition to pemphigus. TNF-alpha gene polymorphisms in position -238 and -308 were identified using a modified polymerase chain reaction-restriction fragment length polymorphism method in 53 patients with pemphigus (38 with pemphigus vulgaris, 15 with pemphigus foliaceus) and 87 healthy controls. The HLA-DRB1 locus was typed using the polymerase chain reaction SSO method in all the patients and 152 population controls. Carriers of the TNF-alpha polymorphic -308 A allele were found to be more frequent in the pemphigus foliaceus group in comparison with the control group (odds ratio (OR) = 8.12; p = 0.0005). A significant association between HLA-DRB1*04 (OR = 3.86; pcor = 0.0001) and DRB1*14 (OR = 8.4; pcor = 0.0001) and pemphigus vulgaris was found. In this group of patients a decreased frequency of HLA-DRB1*07 (OR = 0.08; pcor = 0.006) was also identified. We have shown for the first time a positive association of TNF-alpha polymorphism in position -308 with pemphigus foliaceus.

Murine Ia-associated invariant chain's processing to complex oligosaccharide forms and its dissociation from the I-Ak complex.

PubMed

Holt, G D; Swiedler, S J; Freed, J H; Hart, G W

1985-07-01

The processing of murine invariant chain (Ii) to a cell surface form bearing complex N-linked oligosaccharides has been demonstrated in the B cell lymphoma, AKTB-1b. In addition, the rate of processing of pulse-labeled Ii has been determined relative to its rate of dissociation from the alpha/beta complex of I-Ak. Ii, alpha-, and beta-chains were immunoprecipitated with anti-I-Ak or anti-Ii monoclonal antibodies. The heretofore uncharacterized complex oligosaccharide form of Ii (Ii-c) was identified in gel-purified immunoprecipitates by peptide mapping with reverse-phase HPLC. Ii-c is resistant to deglycosylation by Endo H, which is specific for high-mannose N-linkages, but can be digested with Endo F, a glycosidase capable of cleaving both complex and high-mannose N-linked oligosaccharides. Immunoprecipitation of surface iodinated cells indicates that Ii-c is expressed on the plasma membrane. Pulse-chase metabolic labeling data show that the processing of Ii to Ii-c occurs with a t1/2 of about 120 min. In contrast, the processing of both alpha- and beta-chains of I-Ak to complex forms occurs with a t1/2 of 15 to 20 min. Our data show that Ii-hm begins to dissociate rapidly from the I-Ak complex after 100 to 120 min of chase. Only a small amount (less than 5% on a per mole basis) of Ii-c was found associated with the I-Ak complexes after 300 min of continuous metabolic labeling. These results are consistent with Ii serving as a carrier for Ia antigens as they are transported to the cell surface. In addition, they suggest that the processing of Ii to Ii-c, or a late processing event of the alpha- and beta-chains, such as their sialylation, may be a possible mechanism for inducing the dissociation of Ii from the I-Ak complex.

In situ catalyst improvement in the proline-mediated alpha-amination of aldehydes.

PubMed

Iwamura, Hiroshi; Mathew, Suju P; Blackmond, Donna G

2004-09-29

Kinetic investigations show that the proline-mediated alpha-amination of aldehydes exhibits autoinductive rate behavior and amplification of product enantiomeric excess. Further experiments highlight the role of product, offering suggestions for the design of catalysts of improved efficiency for such transformations. The unusual characteristics exhibited by these reactions implicate amino acid catalysis in rationalizations of the origin of biological homochirality.

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

Suhara, Tadahiro; Kanada-En'yo, Yoshiko

Structures of excited states in {sup 14}C are investigated with a method of {beta}-{gamma} constraint antisymmetrized molecular dynamics in combination with the generator coordinate method. Various excited states with the developed 3{alpha}-cluster core structures are suggested in positive- and negative-parity states. In the positive-parity states, triaxial deformed and linear-chain structures are found to construct excited bands. Interestingly, {sup 10}Be+{alpha} correlation is found in the cluster states above the {sup 10}Be+{alpha} threshold energy.

Heteronuclear NMR assignments and secondary structure of the coiled coil trimerization domain from cartilage matrix protein in oxidized and reduced forms.

PubMed Central

Wiltscheck, R.; Kammerer, R. A.; Dames, S. A.; Schulthess, T.; Blommers, M. J.; Engel, J.; Alexandrescu, A. T.

1997-01-01

The C-terminal oligomerization domain of chicken cartilage matrix protein is a trimeric coiled coil comprised of three identical 43-residue chains. NMR spectra of the protein show equivalent magnetic environments for each monomer, indicating a parallel coiled coil structure with complete threefold symmetry. Sequence-specific assignments for 1H-, 15N-, and 13C-NMR resonances have been obtained from 2D 1H NOESY and TOCSY spectra, and from 3D HNCA, 15N NOESY-HSQC, and HCCH-TOCSY spectra. A stretch of alpha-helix encompassing five heptad repeats (35 residues) has been identified from intra-chain HN-HN and HN-H alpha NOE connectivities. 3JHNH alpha coupling constants, and chemical shift indices. The alpha-helix begins immediately downstream of inter-chain disulfide bonds between residues Cys 5 and Cys 7, and extends to near the C-terminus of the molecule. The threefold symmetry of the molecule is maintained when the inter-chain disulfide bonds that flank the N-terminus of the coiled coil are reduced. Residues Ile 21 through Glu 36 show conserved chemical shifts and NOE connectivities, as well as strong protection from solvent exchange in the oxidized and reduced forms of the protein. By contrast, residues Ile 10 through Val 17 show pronounced chemical shift differences between the oxidized and reduced protein. Strong chemical exchange NOEs between HN resonances and water indicate solvent exchange on time scales faster than 10 s, and suggests a dynamic fraying of the N-terminus of the coiled coil upon reduction of the disulfide bonds. Possible roles for the disulfide crosslinks of the oligomerization domain in the function of cartilage matrix protein are proposed. PMID:9260286

Distinct membrane properties are differentially influenced by cardiolipin content and acyl chain composition in biomimetic membranes.

PubMed

Pennington, Edward Ross; Fix, Amy; Sullivan, E Madison; Brown, David A; Kennedy, Anthony; Shaikh, Saame Raza

2017-02-01

Cardiolipin (CL) has a critical role in maintaining mitochondrial inner membrane structure. In several conditions such as heart failure and aging, there is loss of CL content and remodeling of CL acyl chains, which are hypothesized to impair mitochondrial inner membrane biophysical organization. Therefore, this study discriminated how CL content and acyl chain composition influenced select properties of simple and complex mitochondrial mimicking model membranes. We focused on monolayer excess area/molecule (a measure of lipid miscibility), bilayer phase transitions, and microdomain organization. In monolayer compression studies, loss of tetralinoleoyl [(18:2) 4 ] CL content decreased the excess area/molecule. Replacement of (18:2) 4 CL acyl chains with tetraoleoyl [(18:1) 4 ] CL or tetradocosahexaenoyl [(22:6) 4 ] CL generally had little influence on monolayer excess area/molecule; in contrast, replacement of (18:2) 4 CL acyl chains with tetramyristoyl [(14:0) 4 ] CL increased monolayer excess area/molecule. In bilayers, calorimetric studies showed that substitution of (18:2) 4 CL with (18:1) 4 CL or (22:6) 4 CL lowered the phase transition temperature of phosphatidylcholine vesicles whereas (14:0) 4 CL had no effect. Finally, quantitative imaging of giant unilamellar vesicles revealed differential effects of CL content and acyl chain composition on microdomain organization, visualized with the fluorescent probe Texas Red DHPE. Notably, microdomain areas were decreased by differing magnitudes upon lowering of (18:2) 4 CL content and substitution of (18:2) 4 CL with (14:0) 4 CL or (22:6) 4 CL. Conversely, exchanging (18:2) 4 CL with (18:1) 4 CL increased microdomain area. Altogether, these data demonstrate that CL content and fatty acyl composition differentially target membrane physical properties, which has implications for understanding how CL regulates mitochondrial activity and the design of CL-specific therapeutics. Copyright © 2016 Elsevier B.V. All rights reserved.

Four methods for determining the composition of trace radioactive surface contamination of low-radioactivity metal

NASA Astrophysics Data System (ADS)

O'Keeffe, H. M.; Burritt, T. H.; Cleveland, B. T.; Doucas, G.; Gagnon, N.; Jelley, N. A.; Kraus, C.; Lawson, I. T.; Majerus, S.; McGee, S. R.; Myers, A. W.; Poon, A. W. P.; Rielage, K.; Robertson, R. G. H.; Rosten, R. C.; Stonehill, L. C.; VanDevender, B. A.; Van Wechel, T. D.

2011-12-01

Four methods for determining the composition of low-level uranium- and thorium-chain surface contamination are presented. One method is the observation of Cherenkov light production in water. In two additional methods a position-sensitive proportional counter surrounding the surface is used to make both a measurement of the energy spectrum of alpha particle emissions and also coincidence measurements to derive the thorium-chain content based on the presence of short-lived isotopes in that decay chain. The fourth method is a radiochemical technique in which the surface is eluted with a weak acid, the eluate is concentrated, added to liquid scintillator and assayed by recording beta-alpha coincidences. These methods were used to characterize two 'hotspots' on the outer surface of one of the 3He proportional counters in the Neutral Current Detection array of the Sudbury Neutrino Observatory experiment. The methods have similar sensitivities, of order tens of ng, to both thorium- and uranium-chain contamination.

Polysialic acid in human milk. CD36 is a new member of mammalian polysialic acid-containing glycoprotein.

PubMed

Yabe, Uichiro; Sato, Chihiro; Matsuda, Tsukasa; Kitajima, Ken

2003-04-18

The neural cell adhesion molecule and the voltage-sensitive sodium channel alpha-subunit are the only two molecules in mammals known to be modified by alpha-2,8-linked polysialic acid (polySia). We found a new polySia-containing glycoprotein in human milk and identified it as CD36, a member of the B class of the scavenger receptor superfamily. The polySia-containing glycan chain(s) were removed by alkaline treatment but not by peptide:N-glycanase F digestion, indicating that milk CD36 contained polySia on O-linked glycan chain(s). Polysialylation of CD36 occurs not only in human milk but also in mouse milk. However, CD36 in human platelets is not polysialylated. PolySia CD36 is secreted in milk at any lactation stage and reaches peak level at 1 month after parturition. Thus, it is suggested that polySia of milk CD36 is significant for neonatal development in terms of protection and nutrition.

TNF{alpha} release from peripheral blood leukocytes depends on a CRM1-mediated nuclear export

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

Miskolci, Veronika; Department of Pediatrics, Feinstein Institute for Medical Research at the North Shore-Long Island Jewish Health System, New Hyde Park, NY 11040; Ghosh, Chandra C.

2006-12-15

Tumor necrosis factor-{alpha} (TNF{alpha}) is a potent pro-inflammatory cytokine that plays a major role in the pathogenesis of acute and chronic inflammatory disorders such as septic shock and arthritis, respectively. Leukocytes stimulated with inflammatory signals such as lipopolysaccharide (LPS) are the predominant producers of TNF{alpha}, and thus control of TNF{alpha} release from stimulated leukocytes represents a potential therapeutic target. Here, we report that leptomycin B (LMB), a specific inhibitor of CRM1-dependent nuclear protein export, inhibits TNF{alpha} release from LPS-stimulated human peripheral blood neutrophils and mononuclear cells. In addition, immunofluorescence confocal microscopy and immunoblotting analysis indicate that TNF{alpha} is localized inmore » the nucleus of human neutrophils and mononuclear cells. This study demonstrates that the cellular release of TNF{alpha} from stimulated leukocytes is mediated by the CRM1-dependent nuclear export mechanism. Inhibition of CRM1-dependent cellular release of TNF{alpha} could thus provide a novel therapeutic approach for disorders involving excessive TNF{alpha} release.« less

Surfactant-controlled polymerization of semiconductor clusters to quantum dots through competing step-growth and living chain-growth mechanisms.

PubMed

Evans, Christopher M; Love, Alyssa M; Weiss, Emily A

2012-10-17

This article reports control of the competition between step-growth and living chain-growth polymerization mechanisms in the formation of cadmium chalcogenide colloidal quantum dots (QDs) from CdSe(S) clusters by varying the concentration of anionic surfactant in the synthetic reaction mixture. The growth of the particles proceeds by step-addition from initially nucleated clusters in the absence of excess phosphinic or carboxylic acids, which adsorb as their anionic conjugate bases, and proceeds indirectly by dissolution of clusters, and subsequent chain-addition of monomers to stable clusters (Ostwald ripening) in the presence of excess phosphinic or carboxylic acid. Fusion of clusters by step-growth polymerization is an explanation for the consistent observation of so-called "magic-sized" clusters in QD growth reactions. Living chain-addition (chain addition with no explicit termination step) produces QDs over a larger range of sizes with better size dispersity than step-addition. Tuning the molar ratio of surfactant to Se(2-)(S(2-)), the limiting ionic reagent, within the living chain-addition polymerization allows for stoichiometric control of QD radius without relying on reaction time.

Monte Carlo simulation and equation of state for flexible charged hard-sphere chain fluids: polyampholyte and polyelectrolyte solutions.

PubMed

Jiang, Hao; Adidharma, Hertanto

2014-11-07

The thermodynamic modeling of flexible charged hard-sphere chains representing polyampholyte or polyelectrolyte molecules in solution is considered. The excess Helmholtz energy and osmotic coefficients of solutions containing short polyampholyte and the osmotic coefficients of solutions containing short polyelectrolytes are determined by performing canonical and isobaric-isothermal Monte Carlo simulations. A new equation of state based on the thermodynamic perturbation theory is also proposed for flexible charged hard-sphere chains. For the modeling of such chains, the use of solely the structure information of monomer fluid for calculating the chain contribution is found to be insufficient and more detailed structure information must therefore be considered. Two approaches, i.e., the dimer and dimer-monomer approaches, are explored to obtain the contribution of the chain formation to the Helmholtz energy. By comparing with the simulation results, the equation of state with either the dimer or dimer-monomer approach accurately predicts the excess Helmholtz energy and osmotic coefficients of polyampholyte and polyelectrolyte solutions except at very low density. It also well captures the effect of temperature on the thermodynamic properties of these solutions.

Measurements and analysis of alpha-induced reactions of importance for nuclear astrophysics

NASA Astrophysics Data System (ADS)

de Messieres, Genevieve Escande

2011-11-01

Reactions during stellar helium burning are of primary importance for understanding nucleosynthesis. A detailed understanding of the critical reaction chain 4He(2alpha, gamma)12C( alpha, gamma)16O(alpha, gamma) 20Ne is necessary both because it is the primary energy source and because it determines the ratio of 12C to 16O produced, which in turn significantly effects subsequent nucleosynthesis. Also during Helium burning, the reactions 22Ne(alpha, n)25Mg and 22Ne(alpha, gamma )26Mg are crucial in determining the amount of neutrons available for the astrophysical s-process. This thesis presents new experimental results concerning the 16O(alpha, gamma) 20Ne, 22Ne(alpha, n)25Mg, and 22Ne(alpha, gamma)26Mg reaction rates. These results are then applied to the calculation of the associated stellar reaction rates in order to achieve better accuracy.

Realizing the potential of the Actinium-225 radionuclide generator in targeted alpha-particle therapy applications

PubMed Central

Miederer, Matthias; Scheinberg, David A.; McDevitt, Michael R.

2013-01-01

Alpha particle-emitting isotopes have been proposed as novel cytotoxic agents for augmenting targeted therapy. Properties of alpha particle radiation such as their limited range in tissue of a few cell diameters and their high linear energy transfer leading to dense radiation damage along each alpha track are promising in the treatment of cancer, especially when single cells or clusters of tumor cells are targeted. Actinium-225 (225Ac) is an alpha particle-emitting radionuclide that generates 4 net alpha particle isotopes in a short decay chain to stable 209Bi, and as such can be described as an alpha particle nanogenerator. This article reviews the literature pertaining to the research, development, and utilization of targeted 225Ac to potently and specifically affect cancer. PMID:18514364

Developmental programming of polycystic ovary syndrome (PCOS): prenatal androgens establish pancreatic islet α/β cell ratio and subsequent insulin secretion.

PubMed

Ramaswamy, S; Grace, C; Mattei, A A; Siemienowicz, K; Brownlee, W; MacCallum, J; McNeilly, A S; Duncan, W C; Rae, M T

2016-06-06

Exogenous androgenic steroids applied to pregnant sheep programmes a PCOS-like phenotype in female offspring. Via ultrasound guidance we applied steroids directly to ovine fetuses at d62 and d82 of gestation, and examined fetal (day 90 gestation) and postnatal (11 months old) pancreatic structure and function. Of three classes of steroid agonists applied (androgen - Testosterone propionate (TP), estrogen - Diethystilbesterol (DES) and glucocorticoid - Dexamethasone (DEX)), only androgens (TP) caused altered pancreatic development. Beta cell numbers were significantly elevated in prenatally androgenised female fetuses (P = 0.03) (to approximately the higher numbers found in male fetuses), whereas alpha cell counts were unaffected, precipitating decreased alpha:beta cell ratios in the developing fetal pancreas (P = 0.001), sustained into adolescence (P = 0.0004). In adolescence basal insulin secretion was significantly higher in female offspring from androgen-excess pregnancies (P = 0.045), and an exaggerated, hyperinsulinaemic response to glucose challenge (P = 0.0007) observed, whereas prenatal DES or DEX treatment had no effects upon insulin secretion. Postnatal insulin secretion correlated with beta cell numbers (P = 0.03). We conclude that the pancreas is a primary locus of androgenic stimulation during development, giving rise to postnatal offspring whose pancreas secreted excess insulin due to excess beta cells in the presence of a normal number of alpha cells.

Developmental programming of polycystic ovary syndrome (PCOS): prenatal androgens establish pancreatic islet α/β cell ratio and subsequent insulin secretion

PubMed Central

Ramaswamy, S.; Grace, C.; Mattei, A. A.; Siemienowicz, K.; Brownlee, W.; MacCallum, J.; McNeilly, A. S.; Duncan, W. C.; Rae, M. T.

2016-01-01

Exogenous androgenic steroids applied to pregnant sheep programmes a PCOS-like phenotype in female offspring. Via ultrasound guidance we applied steroids directly to ovine fetuses at d62 and d82 of gestation, and examined fetal (day 90 gestation) and postnatal (11 months old) pancreatic structure and function. Of three classes of steroid agonists applied (androgen - Testosterone propionate (TP), estrogen - Diethystilbesterol (DES) and glucocorticoid - Dexamethasone (DEX)), only androgens (TP) caused altered pancreatic development. Beta cell numbers were significantly elevated in prenatally androgenised female fetuses (P = 0.03) (to approximately the higher numbers found in male fetuses), whereas alpha cell counts were unaffected, precipitating decreased alpha:beta cell ratios in the developing fetal pancreas (P = 0.001), sustained into adolescence (P = 0.0004). In adolescence basal insulin secretion was significantly higher in female offspring from androgen-excess pregnancies (P = 0.045), and an exaggerated, hyperinsulinaemic response to glucose challenge (P = 0.0007) observed, whereas prenatal DES or DEX treatment had no effects upon insulin secretion. Postnatal insulin secretion correlated with beta cell numbers (P = 0.03). We conclude that the pancreas is a primary locus of androgenic stimulation during development, giving rise to postnatal offspring whose pancreas secreted excess insulin due to excess beta cells in the presence of a normal number of alpha cells. PMID:27265420

The stars with H-alpha missing

NASA Technical Reports Server (NTRS)

Downes, Ronald A.

1987-01-01

During the course of a survey for ultraviolet-excess objects, a group of objects were found that spectroscopically resemble the subdwarf B stars, except for variable strength H-alpha absorption. The H-alpha line can have the nominal strength predicted from the other Balmer lines, or it can be completely absent. Observations reveal significant changes in the H-alpha profile on both a night-to-night and month-to-month basis. IUE observations of three stars reveal few features in the ultraviolet region and, when combined with optical data, allow a rough determination of photospheric temperatures. A binary model is proposed for these systems, and the ramifications of the objects found here and in the Palomar-Green survey are discussed.

Ultraviolet observations of alpha Aurigae from Copernicus

NASA Technical Reports Server (NTRS)

Dupree, A. K.

1975-01-01

Emission lines of L-alpha (1215.67 A) and O VI (1031.94 A) were detected in the spectroscopic binary alpha Aur (Capella) with the Princeton experiment on Copernicus. Temperatures of the emitting regions are inferred to be in excess of 300,000 K. The temperature and emission measure are consistent with a variable source of soft X-rays. If the emission is attributed to the primary star (G5 III), the atmosphere is expanding with velocities of about 20-100 km/s. Such expansion can lead to material within the binary system. The density of interstellar hydrogen inferred from absorption of stellar L-alpha appears to be approximately 0.01 hydrogen atoms per cu cm.

A reinterpretation of millimeter observations of nearby IRAS excess stars

NASA Technical Reports Server (NTRS)

Weintraub, David A.; Stern, S. Alan

1994-01-01

We analyze new and previously published 1300, 870, and 800 micrometers, single-element bolometer observations of Vega (alpha Lyr), Formalhaut (alpha PsA), epsilon Eri, tau(sup1) Eri, and Beta Leo. We show that these data are consistent with models in which the dust disks around these stars are larger than the radio telescope beams with which they were observed; thus, these disks may be many hundreds of AU in radius or larger. Our interpretation of submillimeter/millimeter measurements of these stars also indicates that for some Infrared Astronomy Satellite (IRAS) IR excess stars the assumption that there is no astrophysical flux in the OFF beams, when using the standard ON-minus-OFF chopping technique and the chop distance is less than 1000AU, may be incorrect. Therefore, for IRAS IR excess stars within approximately 20 pc, virtually all submillimeter/millimeter chopping observations with chop throws less than 100" may have subtracted away some or most of the flux associated with their circumstellar disks. Finally, we present new 1300 micrometers continuum observations of Vega made with chop throws of 500 and 1000 AU. These data are consistent with an interpretation in which Vega has a disk that is at least 1000AU in radius; this disk could have a region with much less material per beam area near 500 AU than at both 100 AU and 1000 AU, corresponding to a gap at the orbital distance of alpha Lyr B. The observations of Vega are also consistent with the assumption that the circumVega dust structure is unresolved. Thus, these new data very effectively illustrate that the 'standard' model of small, unresolved, dust structures around main-sequence IRAS IR excess stars is not unique. Maps of IRAS IR excess stars, which soon will be available from submillimeter/millimeter array detectors, will determine whether the paradigm shift we propose will occur.

Three outer arm dynein heavy chains of Chlamydomonas reinhardtii operate in a coordinated fashion both in vitro and in vivo.

PubMed

Takazaki, Hiroko; Liu, Zhongmei; Jin, Mingyue; Kamiya, Ritsu; Yasunaga, Takuo

2010-07-01

Outer arm dynein (OAD) in cilia and flagella contains two to three nonidentical heavy chains (HCs) that possess motor activity. In Chlamydomonas, flagellar OAD contains three HCs, alpha-, beta-, and gamma-HCs, each appearing to have a distinct role. To determine the precise molecular mechanism of their function, cross-sectional electron micrographs of wild-type and single HC-disruption mutants were compared and statistically analyzed. While the alpha-HC mutant displayed an OAD of lower density, which was attributed to a lack of alpha-HC, the OAD of beta- and gamma-HC mutants not only lacked the corresponding HC, but was also significantly affected in its structure, particularly with respect to the localization of alpha-HC. The lack of beta-HC induced mislocalization of alpha-HC, while a disruption of the gamma-HC gene resulted in the synchronized movement of alpha-HC and beta-HC in the manners for stacking. Interestingly, using cryo-electron microscopy, purified OADs were typically observed consisting of two stacked heads and an independent single head, which presumably corresponded to gamma-HC. This conformation is different from previous reports in which the three HCs displayed a stacked form in flagella observed by cryo-electron tomography and a bouquet structure on mica in deep-etch replica images. These results suggest that gamma-HC supports the tight stacking arrangement of inter or intra alpha-/beta-HC to facilitate the proper functioning of OAD. 2010 Wiley-Liss, Inc.

The Val192Leu mutation in the alpha-subunit of beta-hexosaminidase A is not associated with the B1-variant form of Tay-Sachs disease.

PubMed Central

Hou, Y.; Vavougios, G.; Hinek, A.; Wu, K. K.; Hechtman, P.; Kaplan, F.; Mahuran, D. J.

1996-01-01

Substitution mutations adversely affecting the alpha-subunit of beta-hexosaminidase A (alphabeta) (EC 3.2.1.52) result in Tay-Sachs disease. The majority affect the initial folding of the pro-alpha chain in the endoplasmic reticulum, resulting in its retention and degradation. A much less common occurrence is a mutation that specifically affects an "active-site" residue necessary for substrate binding and/or catalysis. In this case, hexosaminidase A is present in the lysosome, but it lacks all alpha-specific activity. This biochemical phenotype is referred to as the "B1-variant form" of Tay-Sachs disease. Kinetic analysis of suspected B1-variant mutations is complex because hexosaminidase A is heterodimeric and both subunits possess similar active sites. In this report, we examine a previously identified B1-variant mutation, alpha-Val192Leu. Chinese hamster ovary cells were permanently cotransfected with an alpha-cDNA-construct encoding the substitution and a mutant beta-cDNA (beta-Arg211Lys), encoding a beta-subunit that is inactive but normal in all other respects. We were surprised to find that the Val192Leu substitution, produced a pro-alpha chain that did not form alpha-beta dimers and was not transported to the lysosome. Finally, we reexamined the hexosaminidase activity and protein levels in the fibroblasts from the original patient. These data were also not consistent with the biochemical phenotype of the B1 variant of Tay-Sachs disease previously reported to be present. Thus, we conclude that the Val192Leu substitution does not specifically affect the alpha-active site. Images Figure 1 Figure 2 Figure 3 Figure 4 PMID:8659543

The Val192Leu mutation in the alpha-subunit of beta-hexosaminidase A is not associated with the B1-variant form of Tay-Sachs disease.

PubMed

Hou, Y; Vavougios, G; Hinek, A; Wu, K K; Hechtman, P; Kaplan, F; Mahuran, D J

1996-07-01

Substitution mutations adversely affecting the alpha-subunit of beta-hexosaminidase A (alphabeta) (EC 3.2.1.52) result in Tay-Sachs disease. The majority affect the initial folding of the pro-alpha chain in the endoplasmic reticulum, resulting in its retention and degradation. A much less common occurrence is a mutation that specifically affects an "active-site" residue necessary for substrate binding and/or catalysis. In this case, hexosaminidase A is present in the lysosome, but it lacks all alpha-specific activity. This biochemical phenotype is referred to as the "B1-variant form" of Tay-Sachs disease. Kinetic analysis of suspected B1-variant mutations is complex because hexosaminidase A is heterodimeric and both subunits possess similar active sites. In this report, we examine a previously identified B1-variant mutation, alpha-Val192Leu. Chinese hamster ovary cells were permanently cotransfected with an alpha-cDNA-construct encoding the substitution and a mutant beta-cDNA (beta-Arg211Lys), encoding a beta-subunit that is inactive but normal in all other respects. We were surprised to find that the Val192Leu substitution, produced a pro-alpha chain that did not form alpha-beta dimers and was not transported to the lysosome. Finally, we reexamined the hexosaminidase activity and protein levels in the fibroblasts from the original patient. These data were also not consistent with the biochemical phenotype of the B1 variant of Tay-Sachs disease previously reported to be present. Thus, we conclude that the Val192Leu substitution does not specifically affect the alpha-active site.

Structural reorganization of the interleukin-7 signaling complex

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

McElroy, Craig A.; Holland, Paul J.; Zhao, Peng

2012-06-29

We report here an unliganded receptor structure in the common gamma-chain ({gamma}{sub c}) family of receptors and cytokines. The crystal structure of the unliganded form of the interleukin-7 alpha receptor (IL-7R{alpha}) extracellular domain (ECD) at 2.15 {angstrom} resolution reveals a homodimer forming an 'X' geometry looking down onto the cell surface with the C termini of the two chains separated by 110 {angstrom} and the dimer interface comprising residues critical for IL-7 binding. Further biophysical studies indicate a weak association of the IL-7R{alpha} ECDs but a stronger association between the {gamma}{sub c}/IL-7R{alpha} ECDs, similar to previous studies of the full-lengthmore » receptors on CD4{sup +} T cells. Based on these and previous results, we propose a molecular mechanism detailing the progression from the inactive IL-7R{alpha} homodimer and IL-7R{alpha}-{gamma}{sub c} heterodimer to the active IL-7-IL-7R{alpha}-{gamma}{sub c} ternary complex whereby the two receptors undergo at least a 90{sup o} rotation away from the cell surface, moving the C termini of IL-7R{alpha} and {gamma}{sub c} from a distance of 110 {angstrom} to less than 30 {angstrom} at the cell surface. This molecular mechanism can be used to explain recently discovered IL-7- and {gamma}{sub c}-independent gain-of-function mutations in IL-7R{alpha} from B- and T-cell acute lymphoblastic leukemia patients. The mechanism may also be applicable to other {gamma}{sub c} receptors that form inactive homodimers and heterodimers independent of their cytokines.« less

Results from the First {sup 249}Cf + {sup 48}Ca Experiment

DOE R&D Accomplishments Database

Oganessian, Y. T.; Utyonkov, V. K.; Lobanov, Y. V.; Abdullin, F. S.; Polyakov, A. N.; Shirokovsky, I. V.; Tsyganov, Y. S.; Mezentsev, A. N.; Iliev, S.; Subbotin, V. G.; Sukhov, A. M.; Ivanov, O. V.; Voinov, A. A.; Subotic, K.; Zagrebaev, V. I.; Itkis, M. G.; Moody, K. J.; Wild, J. F.; Stoyer, M. A.; Stoyer, N. J.; Laue, C. A.; Shaughnessy, D. A.; Patin, J. B.; Lougheed, R. W.

2003-02-03

The present paper reports the results of an attempt aimed at the synthesis of element 118 in the reaction {sup 249}Cf({sup 48}Ca,3n){sup 294}118. The experiment was performed employing the Dubna Gas-filled Recoil Separator and the U400 heavy-ion cyclotron at FLNR, JINR, Dubna. In the course of a 2300-hour irradiation of an enriched {sup 249}Cf target (0.23 mg/cm{sup 2}) with a beam of 245-MeV {sup 48}Ca ions, we accumulated a total beam dose of 2.5 x 10{sup 19} ions. We detected two events that may be attributed to the formation and decay of nuclei with Z=118. For one event, we observed a decay chain of two correlated {alpha}-decays with corresponding energies and correlation times of E{sub {alpha}1} = 11.65 {+-} 0.06 MeV, t{sub {alpha}1} = 2.55 ms and E{sub {alpha}2} = 10.71 {+-} 0.17 MeV, t{sub {alpha}2} = 42.1 ms and, finally, a spontaneous fission with the sum of the kinetic energies of the fission fragments E{sub tot} = 207 MeV (TKE {approx} 230 MeV) and t{sub SF} = 0.52 s. In the second event chain, the recoil nucleus decayed into two fission fragments with E{sub tot} = 223 MeV (TKE {approx} 245 MeV) 3.16 ms later, without intervening {alpha} decays. The probabilities that these events were caused by the chance correlations of unrelated signals are negligible. Both events were observed at an excitation energy of the compound nucleus {sup 297}118 of E* = 30.0 {+-} 2.4 MeV, close to the expected maximum of the 3n-evaporation channel. The relationship between the decay energy Q{sub {alpha}} and decay period T{sub {alpha}} shows that sequential {alpha}-transitions in the first event correspond to the decay chain with Z = 118{_}116{_}114. Decay characteristics of the newly observed nuclides are compared with radioactive decay properties of the even-even isotopes with Z = 116, 114 and 112 previously produced in the reactions {sup 244}Pu, {sup 248}Cm + {sup 48}Ca and calculations made in various nuclear models.

Logistical and Analytical Approach to a Failure Aboard the International Space Station

NASA Technical Reports Server (NTRS)

McDanels, Seve; Wright, M. Clara; Salazar, Victoria; Lubas, David; Tucker, Bryan

2009-01-01

The starboard Solar Alpha Rotary Joint (SARJ) from the International Space Station (ISS) began exhibiting off-nominal electrical demands and vibration. Examination by spacewalking astronauts revealed metallic debris contaminating the system and damage to the outboard race of the SARJ. Samples of the contamination were returned to Earth and analyzed. Excessive friction caused the nitride region of the 15-5 PH stainless steel race to spall, generating the debris and damaging the race surface. Excessive vibration and excess power was required to operate the system as a result.

Three-Dimensional Polypeptide Architectures Through Tandem Catalysis and Click Chemistry

NASA Astrophysics Data System (ADS)

Rhodes, Allison Jane

Rapid renal clearance, liver accumulation, proteolytic degradation and non-specificity are challenges small molecule drugs, peptides, proteins and nucleic acid therapeutics encounter en route to their intended destination within the body. Nanocarriers (i.e. dendritric polymers, vesicles, and micelles) of approximately 100 nm in diameter, shuttle small molecule drugs to their desired location through passive (EPR effect) and active (ligand-mediated) targeting, maximizing therapeutic efficiency. Polypeptide-based polymers are water-soluble, biocompatible, non-toxic and are therefore excellent candidates for nanocarriers. Dendritic polymers, including dendrimers, cylindrical brushes, and star polymers, are the newest class of nanomedicine drug delivery vehicles. The synthesis and characterization of dendritic polymers is challenging, with tedious and costly procedures. Dendritic polymers possess peripheral pendent functional groups that can potentially be used in ligand-mediated drug delivery vehicles and bioimaging applications. More specifically, cylindrical brushes are dendritic polymers where a single linear polymer (primary chain) has polymer chains (secondary chains) grafted to it. Recently, research groups have shown that cylindrical brush polymers are capable of nanoparticle and supramolecular structure self-assembly. The facile preparation of high-density brush copolypeptides by the "grafting from" approach will be discussed. This approach utilizes a novel, tandem catalytic methodology where alloc-alpha-aminoamide groups are installed within the side-chains of the alpha-amino-N-carboxyanhydride (NCA) monomer serving as masked initiators. These groups are inert during cobalt initiated NCA polymerization, and give alloc-alpha-aminoamide substituted polypeptide main-chains. The alloc-alpha-aminoamide groups are activated in situ using nickel to generate initiators for growth of side-chain brush segments. This method proves to be efficient, yielding well-defined, high-density brushes for applications in drug delivery and imaging. Here, we also report a method for the synthesis of soluble, well-defined, azido functionalized polypeptides in a straightforward, 3-step synthesis. Homo and diblock azidopolypeptides were prepared with controlled segment lengths via living polymerization using Co(PMe3)4 initiator. Through copper azide alkyne click chemistry (CuAAC) in organic solvent, azidopolypeptides were regioselectively and quantitatively modified with carboxylic acid (pH-responsive), amino acid and sugar functional groups. Finally, the advances towards well-defined hyperbranched polypeptides through alpha-amino-acid-N-thiocarboxyanhydrides (NTAs) will be discussed. Within the past 10 years, controlled NCA (alpha-amino acid-N-carboxyanhydride) ring-opening polymerization (ROP) has emerged, expanding the application of copolypeptide polymers in various drug delivery and tissue engineering motifs. Modification of NCA monomers to the corresponding alpha-amino-acid-N-thiocarboxyanhydride (NTA) will diversify ROP reactions, leading to more complex polypeptides (such as hyperbranched polymers), in addition to the possibility of performing these polymerizations under ambient conditions, which would greatly expand their potential utility. The project focuses on the preparation of hyperbranched polypeptides with well-defined architectures and controlled branching density in a one-pot reaction. This will be accomplished by taking advantage of the different selectivities of Co(PMe3)4 and depeNi(COD) polymerization initiators, and by exploiting the reactivity difference between NCA and the more stable NTA monomers.

Serum Free Light Chains in Neoplastic Monoclonal Gammopathies: Relative Under-Detection of Lambda Dominant Kappa/Lambda Ratio, and Underproduction of Free Lambda Light Chains, as Compared to Kappa Light Chains, in Patients With Neoplastic Monoclonal Gammopathies

PubMed Central

Lee, Won Sok; Singh, Gurmukh

2018-01-01

Background Quantitative evaluation of serum free light chains is recommended for the work up of monoclonal gammopathies. Immunoglobulin light chains are generally produced in excess of heavy chains. In patients with monoclonal gammopathy, κ/λ ratio is abnormal less frequently with lambda chain lesions. This study was undertaken to ascertain if the levels of overproduction of the two light chain types and their detection rates are different in patients with neoplastic monoclonal gammopathies. Methods Results of serum protein electrophoresis (SPEP), serum protein immunofixation electrophoresis (SIFE), urine protein electrophoresis (UPEP), urine protein immunofixation electrophoresis (UIFE), and serum free light chain assay (SFLCA) in patients with monoclonal gammopathies were examined retrospectively. Results The κ/λ ratios were appropriately abnormal more often in kappa chain lesions. Ratios of κ/λ were normal in about 25% of patients with lambda chain lesions in whom free homogenous lambda light chains were detectable in urine. An illustrative case suggests underproduction of free lambda light chains, in some instances. Conclusions The lower prevalence of lambda dominant κ/λ ratio in lesions with lambda light chains is estimated to be due to relative under-detection of lambda dominant κ/λ ratio in about 25% of the patients and because lambda chains are not produced in as much excess of heavy chains as are kappa chains, in about 5% of the patients. The results question the medical necessity and clinical usefulness of the serum free light chain assay. UPEP/UIFE is under-utilized. PMID:29904440

Efficient asymmetric alpha-oxyamination of aldehydes by resin-supported peptide catalyst in aqueous media.

PubMed

Akagawa, Kengo; Fujiwara, Takuma; Sakamoto, Seiji; Kudo, Kazuaki

2010-04-16

The resin-supported peptide catalyst having the terminal five-residue Pro-d-Pro-Aib-Trp-Trp combined with polyleucine successfully catalyzed the asymmetric alpha-oxyamination of aldehydes in aqueous media. The secondary structure and the chirality sense of the hydrophobic polyleucine chain significantly affected both reactivity and enantioselectivity.

Alpha-beta T cells provide protection against lethal encephalitis in the murine model of VEEV infection

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

Paessler, Slobodan; Yun, Nadezhda E.; Judy, Barbara M.

2007-10-25

We evaluated the safety and immunogenicity of a chimeric alphavirus vaccine candidate in mice with selective immunodeficiencies. This vaccine candidate was highly attenuated in mice with deficiencies in the B and T cell compartments, as well as in mice with deficient gamma-interferon responsiveness. However, the level of protection varied among the strains tested. Wild type mice were protected against lethal VEEV challenge. In contrast, alpha/beta ({alpha}{beta}) TCR-deficient mice developed lethal encephalitis following VEEV challenge, while mice deficient in gamma/delta ({gamma}{delta}) T cells were protected. Surprisingly, the vaccine potency was diminished by 50% in animals lacking interferon-gamma receptor alpha chain (R1)-chainmore » and a minority of vaccinated immunoglobulin heavy chain-deficient ({mu}MT) mice survived challenge, which suggests that neutralizing antibody may not be absolutely required for protection. Prolonged replication of encephalitic VEEV in the brain of pre-immunized mice is not lethal and adoptive transfer experiments indicate that CD3{sup +} T cells are required for protection.« less

Structural determination of the acidic exopolysaccharide produced by a Pseudomonas sp. strain 1.15.

PubMed

Cescutti, P; Toffanin, R; Pollesello, P; Sutherland, I W

1999-01-31

Pseudomonas strain 1.15 was isolated from a freshwater biofilm and shown to produce considerable amounts of an acidic polysaccharide which was investigated by methylation analysis, NMR spectroscopy and ionspray mass spectrometry (ISMS). The polysaccharide was depolymerised by a bacteriophage-associated endoglucosidase and by autohydrolysis, and the resulting oligosaccharides were investigated by NMR spectroscopy and mass spectrometry. The resulting data showed that the parent repeating unit of the 1.15 exopolysaccharide (EPS) is a branched hexasaccharide. The main chain is constituted of the trisaccharide -->4)-alpha-L-Fucp-(1-->4)-alpha-L-Fucp-(1-->3)-beta-D-Glcp- (1--> and the side chain alpha-D-Galp-(1-->4)-beta-D-GlcAp-(1-->3)-alpha-D-Galp-(1-->is linked to O-3 of the first Fuc residue. The terminal non-reducing Gal carries a 1-carboxyethylidene acetal in the R configuration at the positions 4 and 6. Of the four different O-acetyl groups present in non-stoichiometric amounts, two were established to be on O-2 of the 3-linked Gal and on O-2 of the 4-linked Fuc.

Ketone EC50 values in the Microtox test.

PubMed

Chen, H F; Hee, S S

1995-03-01

The Microtox EC50 values for the following ketones are reported in the following homologous series: straight chain methyl ketones (acetone, 2-butanone, 2-pentanone, 2-hepatonone, 2-octanone, 2-decanone, and 2-tridecanone); methyl ketones substituted at one alpha carbon (3-methyl-2-butanone; 3,3-dimethyl-2-butanone); methyl substituted at two alpha carbons (2,4-dimethyl-3-pentanone; 2,2,4,4-tetramethyl-3-pentanone); phenyl groups replacing methyl in acetone (acetophenone; benzophenone); methyl groups substituted at the alpha carbons of cyclohexanone; and 2,3- 2,4-, and 2,5-hexanediones, most for the first time. While there were linear relationships between log EC50 and MW for the straight chain methyl ketones, and for methyl substitution at the alpha carbon for methyl ketones, there were no other linear relationships. As molecular weight increased, the EC50 values of soluble ketones decreased; as distance between two carbonyl groups decreased so too did EC50 values. Thus, for the ketones the geometry around the carbonyl group is an important determinant of toxicity as well as MW, water solubility, and octanol/water coefficient.

Solution structure for Pandinus toxin K-alpha (PiTX-K alpha), a selective blocker of A-type potassium channels.

PubMed

Tenenholz, T C; Rogowski, R S; Collins, J H; Blaustein, M P; Weber, D J

1997-03-11

PiTX-K alpha, a 35-residue peptide recently isolated from the venom of Pandinus imperator, blocks the rapidly inactivating (A-type) K+ channel(s) in rat brain synaptosomes and the cloned Kv 1.2 potassium channel at very low toxin concentrations (6 nM and 32 pM, respectively) [Rogowski, R. S., Collins, J. H., O'Neil, T. J., Gustafson, T. A., Werkman, T. A., Rogawski, M. A., Tenenholz, T. C., Weber, D. J., & Blaustein, M. P. (1996) Mol. Pharmacol. 50, 1167-1177]. The three-dimensional structure of PiTX-K alpha was determined using NMR spectroscopy in order to understand its selectivity and affinity toward K+ channels. PiTX-K alpha was found to have an alpha-helix from residues 10 to 21 and two beta-strands (betaI, 26-28; betaII, 33-35) connected by a type II beta-turn to form a small antiparallel beta-sheet. Three disulfide bonds, which are conserved in all members of the charybdotoxin family (alpha-K toxins), anchor one face of the alpha-helix to the beta-sheet. The N-terminal portion of PiTX-K alpha has three fewer residues than other alpha-K toxins such as charybdotoxin. Rather than forming a third beta-strand as found for other alpha-K toxins, the N-terminal region of PiTX-K alpha adopts an extended conformation. This structural difference in PiTX-K alpha together with differences in sequence at Pro-10, Tyr-14, and Asn-25 (versus Ser-10, Trp-14, and Arg-25 in CTX) may explain why PiTX-K alpha does not block maxi-K+ channels. Differences in three-dimensional structure between PiTX-K alpha and charybdotoxin are also observed in both the tight turn and the loop that connects the first beta-strand to the alpha-helix. As a result, side chains of two residues (Tyr-23 and Arg-31) are in regions of PiTX-K alpha that probably interact with rapidly inactivating A-type K+ channels. The analogous residues in charybdotoxin are positioned differently on the toxin surface. Thus, the locations of Tyr-23 and Arg-31 side chains in PiTX-K alpha could explain why this toxin blocks A-type channels at much lower concentrations than does charybdotoxin.

The effects of branched-chain amino acid granules on the accumulation of tissue triglycerides and uncoupling proteins in diet-induced obese mice.

PubMed

Arakawa, Mie; Masaki, Takayuki; Nishimura, Junko; Seike, Masataka; Yoshimatsu, Hironobu

2011-01-01

It has been demonstrated the involvement of branched-chain amino acids (BCAA) on obesity and related metabolic disorder. We investigated the effects of branched-chain amino acids (BCAA) on obesity and on glucose/fat homeostasis in mice fed on a high-fat (45%) diet. BCAA was dissolved in 0.5% methylcellulose and added to the drinking water (BCAA-treated group). A high-fat diet was provided for 6 weeks and BCAA was given for 2 weeks. The BCAA-treated group gained almost 7% less body weight and had less epididymal adipose tissue (WAT) mass than the control group (p<0.05). BCAA supplementation also reduced the hepatic and skeletal muscle triglyceride (TG) concentrations (p<0.05). The hepatic levels of PPAR-alpha and uncoupling protein (UCP) 2, and the level of PPAR-alpha and UCP3 in the skeletal muscle were greater in the BCAA-treated group than in the control mice (p<0.05). These results demonstrate that the liver and muscle TG concentration are less in BCAA-treated group. BCAA affects PPAR-alpha and UCP expression in muscle and liver tissue.

The role of the carbohydrate chains in complement (C3) fixation by solid-phase-bound human IgA.

PubMed Central

Nikolova, E B; Tomana, M; Russell, M W

1994-01-01

In contrast to antigen-antibody complexes containing native human IgA, solid-phase-deposited IgA activates the alternative complement pathway and binds C3b. To investigate the role of carbohydrate chains in this, various human IgA preparations were treated with neuraminidase alone or together with N-glycanase or O-glycanase, or with mixed glycosidases from the oral bacterium, Streptococcus mitis. Depletion of oligosaccharides was determined by carbohydrate analysis. Removal of sialic acid and N-linked glycan chains greatly increased the C3b-fixing properties of normal serum IgA1 and IgA2. Myeloma IgA1 and IgA2 proteins and secretory IgA had higher C3b-binding activity than normal serum IgA, and this was further increased by removal of sialic acid and N-linked glycans. Fc alpha and Fc alpha-SC fragments of myeloma and secretory IgA1, respectively, but not Fab alpha fragments, obtained by cleavage with bacterial IgA1 proteases and also free secretory component, fixed C3b by the alternative pathway. Images Figure 4 PMID:7927504

Jacalin: a jackfruit (Artocarpus heterophyllus) seed-derived lectin of versatile applications in immunobiological research.

PubMed

Kabir, S

1998-03-15

Jacalin, the major protein from the jackfruit (Artocarpus heterophyllus) seeds, is a tetrameric two-chain lectin (molecular mass 65 kDa) combining a heavy alpha chain of 133 amino acid residues with a light beta chain of 20-21 amino acid residues. It is highly specific for the alpha-O-glycoside of the disaccharide Thomsen-Friedenreich antigen (Gal beta1-3GalNAc), even in its sialylated form. This property has made jacalin suitable for studying various O-linked glycoproteins, particularly human IgA1. Jacalin's uniqueness in being strongly mitogenic for human CD4+ T lymphocytes has made it a useful tool for the evaluation of the immune status of patients infected with human immunodeficiency virus (HIV)-1. The abundance of source material for the production of jacalin, its ease of purification, yield and stability have made it an attractive cost-effective lectin. It has found applications in diverse areas such as the isolation of human plasma glycoproteins (IgA1, C1-inhibitor, hemopexin, alpha2-HSG), the investigation of IgA-nephropathy, the analysis of O-linked glycoproteins and the detection of tumours.

Predictions on the modes of decay of even Z superheavy isotopes within the range 104 ≤ Z ≤ 136

NASA Astrophysics Data System (ADS)

Santhosh, K. P.; Nithya, C.

2018-01-01

The decay modes and half lives of all the even Z isotopes of superheavy elements within the range 104 ≤ Z ≤ 136 have been predicted by comparing the alpha decay half-lives with the spontaneous fission half-lives. The Coulomb and proximity potential model for deformed nuclei (CPPMDN) and the shell-effect-dependent formula of Santhosh et al. are used to calculate the alpha half-lives and spontaneous fission half-lives respectively. For theoretical comparison the alpha decay half-lives are also calculated using Coulomb and proximity potential model (CPPM), the Viola-Seaborg-Sobiczewski semi-empirical (VSS) relation, the universal (UNIV) curve of Poenaru et al., the analytical formula of Royer and the universal decay law (UDL) of Qi et al. Another tool used for the evaluation of spontaneous fission half-lives is the semi-empirical formula of Xu et al. The nuclei with alpha decay half-lives less than spontaneous fission half-lives will survive fission and hence decay through alpha emission. The predicted half lives and decay modes are compared with the available experimental results. The one-proton and two-proton separation energies of all the isotopes are calculated to find nuclei which lie beyond the proton drip line. Among 1119 even Z nuclei within the range 104 ≤ Z ≤ 136, 164 nuclei show sequential alpha emission followed by subsequent spontaneous fission. Since the isotopes decay through alpha decay chain and the half-lives are in measurable range, these isotopes are predicted to be synthesized and detected in laboratory via alpha decay. 2 nuclei will decay by alpha decay followed by proton emission, 54 nuclei show full alpha chains, 642 nuclei will decay through spontaneous fission, 166 nuclei exhibit proton decay and 91 isotopes are found to be stable against alpha decay. All the isotopes are tabulated according to their decay modes. The study is intended to enhance further experimental investigations in superheavy region.

Investigation of isovaline enantiomeric excesses in CM meteorites using liquid chromatography time of flight mass spectrometry

NASA Technical Reports Server (NTRS)

Glavin, Daniel P.; Dworkin, Jason P.

2003-01-01

The enantiomeric abundances of the alpha-dialkyl amino acid isovaline were measured in the CM2 meteorites Murchison and LEW 90500 using a new liquid chromatography-time of flight-mass spectrometry (LC-ToF-MS) technique coupled with OPA/NAC derivatization and UV fluorescence detection. Previous analyses of Murchison have shown that L-enantiomeric excesses of isovaline range from 0 to 15.2% with significant variation between meteorite fragments [1]. For this study, hot water extracts of interior fragments (> 2 cm from fusion crust) of the Murchison (USNM 6650.2, mass 6 g) and LEW 90500 (split 69, parent 1, mass 5 g) carbonaceous meteorites were analyzed. Enantiomeric excesses were measured using the single ion LC-ToF-MS trace for the OPA/NAC derivative of isovaline at d z 393.15 (Fig. 1). L-isovaline excesses in these meteorite samples ranged from 18.9 to 20.5% for Murchison and -0.5 to 3.0% for LEW 90500. The measured values for Murchison are the largest enantiomeric excesses for isovaline reported to date. The enantiomeric excesses of L-isovaline cannot be the result of interference from other C5 amino acid isomers present in the meteorites or terrestrial contamination from the landing site environments. The L-isovaline excesses in Murchison are inconsistent with the synthesis of all of the isovaline by the Strecker-cyanohydrin pathway on the CM meteorite parent body. The mechanism(s) for the formation of the enantiomeric asymmetry in isovaline in Murchison are currently unknown and it is not clear how the asymmetry of alpha-dialkyl amino acids could be transferred to the a-hydrogen protein amino acids common in all life on Earth today.

Spark Plasma Sintering for Nanostructured Smart Materials

DTIC Science & Technology

2009-03-02

polyester) with excess isocyanate to form a prepolymer , followed by the addition of a short chain diol that acts as a chain extender to link the... prepolymers together. Due to the thermodynamic imicisibility of segments of PU, phase separation into a flexible soft segment (long chain diol) and a...other reactions of the isocyanate groups with the other functional groups in the chain. [Hepburn, 1992] However, during the initial prepolymer

Thermal conductivity of the Lennard-Jones chain fluid model.

PubMed

Galliero, Guillaume; Boned, Christian

2009-12-01

Nonequilibrium molecular dynamics simulations have been performed to estimate, analyze, and correlate the thermal conductivity of a fluid composed of short Lennard-Jones chains (up to 16 segments) over a large range of thermodynamic conditions. It is shown that the dilute gas contribution to the thermal conductivity decreases when the chain length increases for a given temperature. In dense states, simulation results indicate that the residual thermal conductivity of the monomer increases strongly with density, but is weakly dependent on the temperature. Compared to the monomer value, it has been noted that the residual thermal conductivity of the chain was slightly decreasing with its length. Using these results, an empirical relation, including a contribution due to the critical enhancement, is proposed to provide an accurate estimation of the thermal conductivity of the Lennard-Jones chain fluid model (up to 16 segments) over the domain 0.8

Pathophysiologic roles of the fibrinogen gamma chain.

PubMed

Farrell, David H

2004-05-01

Fibrinogen binds through its gamma chains to cell surface receptors, growth factors, and coagulation factors to perform its key roles in fibrin clot formation, platelet aggregation, and wound healing. However, these binding interactions can also contribute to pathophysiologic processes, including inflammation and thrombosis. This review summarizes the latest findings on the role of the fibrinogen gamma chain in these processes, and illustrates the potential for therapeutic intervention. Novel gamma chain epitopes that bind platelet integrin alpha IIbbeta3 and leukocyte integrin alphaMbeta2 have been characterized, leading to the revision of former dogma regarding the processes of platelet aggregation, clot retraction, inflammation, and thrombosis. A series of studies has shown that the gamma chain serves as a depot for fibroblast growth factor-2 (FGF-2), which is likely to play an important role in wound healing. Inhibition of gamma chain function with the monoclonal antibody 7E9 has been shown to interfere with multiple fibrinogen activities, including factor XIIIa crosslinking, platelet adhesion, and platelet-mediated clot retraction. The role of the enigmatic variant fibrinogen gamma chain has also become clearer. Studies have shown that gamma chain binding to thrombin and factor XIII results in clots that are mechanically stiffer and resistant to fibrinolysis, which may explain the association between gammaA/gamma' fibrinogen levels and cardiovascular disease. The identification of new interactions with gamma chains has revealed novel targets for the treatment of inflammation and thrombosis. In addition, several exciting studies have shown new functions for the variant gamma chain that may contribute to cardiovascular disease.

Product ion tandem mass spectrometric differentiation of regioisomeric side-chain groups in cathinone derivatives.

PubMed

Abiedalla, Younis; DeRuiter, Jack; Clark, C Randall

2016-07-30

Precursor materials are available to prepare aminoketone drugs containing regioisomeric propyl and isopropyl side-chain groups related to the drug alpha-pyrrovalerone (Flakka) and MDPV (3,4-methylenedioxypyrrovalerone). These compounds yield equivalent regioisomeric iminium cation base peaks in electron ionization mass spectrometry (EI-MS). The propyl and isopropyl side-chain groups related to alpha-pyrrovalerone and MDPV were prepared and evaluated in EI-MS and tandem mass spectrometry (MS/MS) product ion experiments. Deuterium labeling in both the pyrrolidine and alkyl side-chain groups allowed for the confirmation of the structures for the major product ions formed from the regioisomeric EI-MS iminium cation base peaks. These iminium cation base peaks show characteristic product ion spectra which allow differentiation of the side-chain propyl and isopropyl groups in the structure. The n-propyl side chain containing iminium cation base peak (m/z 126) in the EI-MS spectrum yields a major product ion at m/z 84 while the regioisomeric m/z 126 base peak for the isopropyl side chain yields a characteristic product ion at m/z 70. Deuterium labeling in both the pyrrolidine ring and the alkyl side chain confirmed the process for the formation of these major product ions. Product ion fragmentation provides useful data for differentiation of n-propyl and isopropyl side-chain iminium cations from cathinone derivative drugs of abuse. Regioisomeric n-propyl and isopropyl iminium cations of equal mass yield characteristic product ions identifying the alkyl side-chain regioisomers in the pyrrolidine cathinone derivatives. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

Mixed isotype class II antigen expression. A novel class II molecule is expressed on a murine B cell lymphoma

PubMed Central

1989-01-01

The structures of Ia molecules expressed by two BALB/c B cell lymphoma lines, A20-1.11 (A20) and 2PK3, were analyzed in an effort to explain the differences in antigen-presenting capacity displayed by these cells. Alloreactive T cell hybridomas specific for I-Ad and antigen- specific, I-Ad-restricted T cells responded well to A20 as the APC. The same alloreactive T cell hybridomas responded weakly or not at all to 2PK3 and the responses of the antigen-specific, I-Ad-restricted T cells were consistently lower to antigen presented by 2PK3 as compared with A20. T cells restricted to I-Ed responded equally well to either A20 or 2PK3 as APC. Additionally 2PK3, but not A20, stimulated a strong syngeneic mixed lymphocyte response. Structural analyses of the Ia antigens revealed that I-A and I-E molecules were expressed by A20, whereas an I-E and a novel I-A-like molecule were expressed by 2PK3. The novel class II molecule was affinity purified from 2PK3 cells using an mAb specific for Ad beta (MK-D6), and this molecule was subsequently shown by an RIA to react with an E alpha-specific mAb (14-4-4S) as well. Chain-specific polyclonal antisera raised against I-A and I-E alpha and beta chains indicated that the 2PK3 "I-A" alpha chain reacted in immunoblot with E alpha-specific and not A alpha-specific antisera, whereas the beta chain reacted with A beta- and not E beta-specific antisera. Peptide map and partial amino acid sequence analyses indicated that the "I-A" molecule expressed by 2PK3 represented a mixed isotype structure resulting from the pairing of Ed alpha with Ad beta. By immunofluorescence staining analysis, 2PK3 did not react with an mAb specific for Ad alpha. 2PK3 was capable of limited antigen presentation through the mixed isotype molecule to I-Ad-restricted OVA-specific T cell hybridomas, although the responses induced were low compared with presentation through I-A on A20. Previous descriptions of the expression of mixed isotype class II molecules in the mouse have resulted primarily from DNA-mediated gene transfer experiments. The results presented indicate that a mixed isotype class II molecule can be expressed naturally. PMID:2647893

Problem-Solving Test: The Mechanism of Protein Synthesis

ERIC Educational Resources Information Center

Szeberenyi, Jozsef

2009-01-01

Terms to be familiar with before you start to solve the test: protein synthesis, ribosomes, amino acids, peptides, peptide bond, polypeptide chain, N- and C-terminus, hemoglobin, [alpha]- and [beta]-globin chains, radioactive labeling, [[to the third power]H] and [[to the fourteenth power]C]leucine, cytosol, differential centrifugation, density…

Enhancement of fruit shelf life by suppressing N-glycan processing enzymes.

PubMed

Meli, Vijaykumar S; Ghosh, Sumit; Prabha, T N; Chakraborty, Niranjan; Chakraborty, Subhra; Datta, Asis

2010-02-09

In a globalized economy, the control of fruit ripening is of strategic importance because excessive softening limits shelf life. Efforts have been made to reduce fruit softening in transgenic tomato through the suppression of genes encoding cell wall-degrading proteins. However, these have met with very limited success. N-glycans are reported to play an important role during fruit ripening, although the role of any particular enzyme is yet unknown. We have identified and targeted two ripening-specific N-glycoprotein modifying enzymes, alpha-mannosidase (alpha-Man) and beta-D-N-acetylhexosaminidase (beta-Hex). We show that their suppression enhances fruit shelf life, owing to the reduced rate of softening. Analysis of transgenic tomatoes revealed approximately 2.5- and approximately 2-fold firmer fruits in the alpha-Man and beta-Hex RNAi lines, respectively, and approximately 30 days of enhanced shelf life. Overexpression of alpha-Man or beta-Hex resulted in excessive fruit softening. Expression of alpha-Man and beta-Hex is induced by the ripening hormone ethylene and is modulated by a regulator of ripening, rin (ripening inhibitor). Furthermore, transcriptomic comparative studies demonstrate the down-regulation of cell wall degradation- and ripening-related genes in RNAi fruits. It is evident from these results that N-glycan processing is involved in ripening-associated fruit softening. Genetic manipulation of N-glycan processing can be of strategic importance to enhance fruit shelf life, without any negative effect on phenotype, including yield.

Structural characterization of alpha-terminal group of natural rubber. 2. Decomposition of branch-points by phospholipase and chemical treatments.

PubMed

Tarachiwin, Lucksanaporn; Sakdapipanich, Jitladda; Ute, Koichi; Kitayama, Tatsuki; Tanaka, Yasuyuki

2005-01-01

The treatment of deproteinized natural rubber (DPNR) latex with phospholipases A(2), B, C, and D decreased significantly the long-chain fatty acid ester contents in DPNR and also the molecular weight and Higgins' k' constant, except for phospholipase D treatment. This indicates the presence of phospholipid molecules in NR, which combine rubber molecules together. Transesterification of DPNR resulted in the decomposition of the functional group at the terminal chain-end (alpha-terminal), including phospholipids and formed linear rubber molecules. The addition of small amounts of ethanol into the DPNR solution reduced the molecular weight and shifted the molecular weight distribution (MWD) comparable to that of transesterified DPNR (TE-DPNR). The addition of diammonium hydrogen phosphate into DPNR-latex in order to remove Mg2+ ions yielded a slight decrease in molecular weight and a slight shift in MWD. The phospholipids are expected to link with mono- and diphosphate groups at the alpha-terminal by hydrogen bonding and/or ionic linkages. The decrease in the molecular weight and Huggins' k' constant of DPNR demonstrates the formation of linear molecules after decomposition of branch-points by this treatment, showing that phospholipids participate in the branching formation of NR. The branch-points formed at the alpha-terminus are postulated to originate predominantly by the association of phospholipids via micelle formation of long-chain fatty acid esters and hydrogen bonding between polar headgroups of phospholipids.

Significance of detecting circulating hepatocellular carcinoma cells in peripheral blood of hepatocellular carcinoma patients by nested reverse transcription-polymerase chain reaction and its clinical value: a retrospective study.

PubMed

Liu, Yang; Wang, Yue-ru; Wang, Long; Song, Rui-mei; Zhou, Bo; Song, Zhen-shun

2014-01-01

Circulating hepatocellular carcinoma cells may be detected by reverse transcription-polymerase chain reaction. We investigated the relationship between circulating hepatocellular carcinoma cells and hepatoma patient survival after different managements and survival periods. Peripheral vein blood (5 ml) samples were obtained from 113 patients with hepatocellular carcinoma and from 33 control subjects (9 with liver cirrhosis after hepatitis B, 14 with chronic hepatitis B, 10 healthy individuals) between January 1, 2009, and December 31, 2013. To detect circulating hepatocellular carcinoma cells in peripheral blood, alpha-fetoprotein messenger RNA was amplified from total RNA extracted from whole blood by reverse transcription-polymerase chain reaction. Alpha-fetoprotein messenger RNA was detected in 59 blood samples from the hepatocellular carcinoma patients (59/113, 52.2%). In contrast, there were no clinical control subjects whose samples showed detectable alpha-fetoprotein messenger RNA. The presence of alpha-fetoprotein messenger RNA in blood seemed to be correlated with the stage (by TNM classification) of hepatocellular carcinoma, serum alpha-fetoprotein value, and the presence of intrahepatic metastasis, portal vein thrombosis, tumor diameter and/or distant metastasis. In addition, alpha-fetoprotein messenger RNA was detected in the blood of 25 patients showing distant metastasis at extrahepatic organs (100%), in contrast to 32 of 88 cases without metastasis (36.4%). All the patients with hepatocellular carcinoma were followed. Seventeen patients with resection of a T 2 stage hepatocellular carcinoma had a survival of 3.2 years after surgical management, 38 cases with resection of a T3 stage hepatocellular carcinoma had a 1.3-year survival, and only 37 cases with T4 stage disease after different treatments except surgery survived for 0.6 years (P <0.01). The presence of alpha-fetoprotein messenger RNA in peripheral blood may be an indicator of circulating hepatocellular carcinoma cells, which might predict hematogenous spreading metastasis in patients with hepatocellular carcinoma and may be a poor prognostic factor for hepatocellular carcinoma patients.

Serial EEG findings in anti-NMDA receptor encephalitis: correlation between clinical course and EEG.

PubMed

Ueda, Jun; Kawamoto, Michi; Hikiami, Ryota; Ishii, Junko; Yoshimura, Hajime; Matsumoto, Riki; Kohara, Nobuo

2017-12-01

Anti-NMDA receptor encephalitis is a paraneoplastic encephalitis characterised by psychiatric features, involuntary movement, and autonomic instability. Various EEG findings in patients with anti-NMDA receptor encephalitis have been reported, however, the correlation between the EEG findings and clinical course of anti-NMDA receptor encephalitis remains unclear. We describe a patient with anti-NMDA receptor encephalitis with a focus on EEG findings, which included: status epilepticus, generalised rhythmic delta activity, excess beta activity, extreme delta brush, and paroxysmal alpha activity upon arousal from sleep, which we term"arousal alpha pattern". Initially, status epilepticus was observed on the EEG when the patient was comatose with conjugate deviation. The EEG then indicated excess beta activity, followed by the emergence of continuous slow activity, including generalised rhythmic delta activity and extreme delta brush, in the most severe phase. Slow activity gradually faded in parallel with clinical amelioration. Excess beta activity persisted, even after the patient became almost independent in daily activities, and finally disappeared with full recovery. In summary, our patient with anti-NMDA receptor encephalitis demonstrated slow activity on the EEG, including extreme delta brush during the most severe phase, which gradually faded in parallel with clinical amelioration, with excess beta activity persisting into the recovery phase.

Spatial and Time Coincidence Detection of the Decay Chain of Short-Lived Radioactive Nuclei

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

Granja, Carlos; Jakubek, Jan; Platkevic, Michal

The quantum counting position sensitive pixel detector Timepix with per-pixel energy and time resolution enables to detect radioactive ions and register the consecutive decay chain by simultaneous position-and time-correlation. This spatial and timing coincidence technique in the same sensor is demonstrated by the registration of the decay chain {sup 8}He{yields}{sup {beta} 8}Li and {sup 8}Li{yields}{sup {beta}-} {sup 8}Be{yields}{alpha}+{alpha} and by the measurement of the {beta} decay half-lives. Radioactive ions, selectively obtained from the Lohengrin fission fragment spectrometer installed at the High Flux Reactor of the ILL Grenoble, are delivered to the Timepix silicon sensor where decays of the implanted ionsmore » and daughter nuclei are registered and visualized. We measure decay lifetimes in the range {>=}{mu}s with precision limited just by counting statistics.« less

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

Downes, R.A.

Galactic plane ultraviolet-excess (uv-excess) objects covering about 1000 square degrees of sky were surveyed. Photographic plates were obtained with both uv and blue filters, to select the uv-excess candidates, which were then observed spectroscopically to determine their classification. Most of the objects selected were nearby early-type stars with low interstellar reddening; however, a collection of hot white dwarfs, subdwarf O (sdO) stars, subdwarf B (sdB) stars, and cataclysmic variables was also found. Photoelectric photometry was obtained for these stars and a statistical analysis was performed to determine the space densities and scale heights for the four classes of objects. Severalmore » interesting objects (or class of objects) were discovered, and data for some of these stars are presented. Among the peculiar objects found are an emission-line white dwarf similar to the pulsating PG 1159 stars, a Population II Wolf-Rayet star, a previously catalogued object with a strong Fe II emission-line spectrum, and a new class of object, resembling the sdB stars, that shows variable strength H..cap alpha.. absorption, with the H..cap alpha.. line sometimes completely filled in.« less

Motion of spin label side chains in cellular retinol-binding protein: correlation with structure and nearest-neighbor interactions in an antiparallel beta-sheet.

PubMed

Lietzow, Michael A; Hubbell, Wayne L

2004-03-23

A goal in the development of site-directed spin labeling in proteins is to correlate the motion of a nitroxide side chain with local structure, interactions, and dynamics. Significant progress toward this goal has been made using alpha-helical proteins of known structure, and the present study is the first step in a similar exploration of a beta-sheet protein, cellular retinol-binding protein (CRBP). Nitroxide side chains were introduced along both interior and edge strands. At sites in interior strands, the side-chain motion is strongly influenced by interactions with side chains of neighboring strands, giving rise to a rich variety of dynamic modes (weakly ordered, strongly ordered, immobilized) and complex electron paramagnetic resonance spectra that are modulated by strand twist. The interactions giving rise to the dynamic modes are explored using mutagenesis, and the results demonstrate the particular importance of the non-hydrogen-bonded neighbor residue in giving rise to highly ordered states. Along edge strands of the beta-sheet, the motion of the side chain is simple and weakly ordered, resembling that at solvent-exposed surfaces of an alpha-helix. A simple working model is proposed that can account for the wide variety of dynamic modes encountered. Collectively, the results suggest that the nitroxide side chain is an effective probe of side-chain interactions, and that site-directed spin labeling should be a powerful means of monitoring conformational changes that involve changes in beta-sheet topology.

Verification survey report of the south waste tank farm training/test tower and hazardous waste storage lockers at the West Valley demonstration project, West Valley, New York

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

Weaver, Phyllis C.

A team from ORAU's Independent Environmental Assessment and Verification Program performed verification survey activities on the South Test Tower and four Hazardous Waste Storage Lockers. Scan data collected by ORAU determined that both the alpha and alpha-plus-beta activity was representative of radiological background conditions. The count rate distribution showed no outliers that would be indicative of alpha or alpha-plus-beta count rates in excess of background. It is the opinion of ORAU that independent verification data collected support the site?s conclusions that the South Tower and Lockers sufficiently meet the site criteria for release to recycle and reuse.

Alport syndrome and thin glomerular basement membrane nephropathy: a practical approach to diagnosis.

PubMed

Haas, Mark

2009-02-01

Alport syndrome and thin glomerular basement membrane nephropathy (TBMN) are genetically heterogeneous conditions characterized by structural abnormalities in the glomerular basement membrane and an initial presentation that usually involves hematuria. Approximately 40% of patients with TBMN are heterozygous carriers for autosomal recessive Alport syndrome, with mutations at the genetic locus encoding type IV collagen alpha(3) [alpha(3)(IV)] and alpha(4) chains. However, although the clinical course of TBMN is usually benign, Alport syndrome, particularly the X-linked form with mutations in the locus encoding the alpha(5) chain of type IV collagen [alpha(5)(IV)], typically results in end-stage renal disease. Electron microscopy is essential to diagnosis of TBMN and Alport syndrome on renal biopsy, although electron microscopy alone is of limited value in distinguishing between TBMN, the heterozygous carrier state of X-linked Alport syndrome, autosomal recessive Alport syndrome, and even early stages of X-linked Alport syndrome. To review diagnostic pathologic features of each of the above conditions, emphasizing the need for immunohistology for alpha(3)(IV) and alpha(5)(IV) in addition to electron microscopy to resolve this differential diagnosis on a renal biopsy. The diagnostic value of immunofluorescence studies for alpha(5)(IV) on a skin biopsy in family members of patients with Alport syndrome also is reviewed. Original and comprehensive review articles on the diagnosis of Alport syndrome and TBMN from the past 35 years, primarily the past 2 decades, and experience in our own renal pathology laboratory. Although Alport syndrome variants and TBMN do not show characteristic light microscopic findings and can be difficult to differentiate from each other even by electron microscopy, using a combination of electron microscopy and immunohistology for alpha(3)(IV) and alpha(5)(IV) enables pathologists to definitively diagnose these disorders on renal biopsy in most cases.

The Val{sup 192}Leu mutation in the {alpha}-subunit of {beta}-hexosaminidase A is not associated with the B1-variant form of Tay-Sachs disease

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

Hou, Y.; Vavougios, G.; Hinek, A.

1996-07-01

Substitution mutations adversely affecting the {alpha}-subunit of {beta}-hexosaminidase A ({alpha}{beta}) (EC 3.2.1.52) result in Tay-Sachs disease. The majority affect the initial folding of the pro-{alpha} chain in the endoplasmic reticulum, resulting in its retention and degradation. A much less common occurrence is a mutation that specifically affects an {open_quotes}active-site{close_quotes} residue necessary for substrate binding and/or catalysis. In this case, hexosaminidase A is present in the lysosome, but it lacks all {alpha}-specific activity. This biochemical phenotype is referred to as the {open_quotes}B1-variant form{close_quotes} of Tay-Sachs disease. Kinetic analysis of suspected B1-variant mutations is complex because hexosaminidase A is heterodimeric and bothmore » subunits possess similar active sites. In this report, we examine a previously identified B1-variant mutation, {alpha}-Val{sup 192}Leu. Chinese hamster ovary cells were permanently cotransfected with an {alpha}-cDNA-construct encoding the substitution and a mutant {beta}-cDNA ({beta}-Arg{sup 211}Lys), encoding a {beta}-subunit that is inactive but normal in all other respects. We were surprised to find that the Val{sup 192}Leu substitution produced a pro-{alpha} chain that did not form {alpha}-{beta} dimers and was not transported to the lysosome. Finally, we reexamined the hexosaminidase activity and protein levels in the fibroblasts from the original patient. These data were also not consistent with the biochemical phenotype of the B1 variant of Tay-Sachs disease previously reported to be present. Thus, we conclude that the Val{sup 192}Leu substitution does not specifically affect the {alpha}-active site. 23 refs., 4 figs., 2 tabs.« less

Protein-expression profiles in mouse blood-plasma following acute whole-body exposure to (137)Cs gamma rays.

PubMed

Rithidech, Kanokporn Noy; Honikel, Louise; Rieger, Robert; Xie, Weiping; Fischer, Thomas; Simon, Sanford R

2009-05-01

To compare the pattern of protein-expression profiles in blood-plasma after exposure of CBA/CaJ mice to 0 or 3 Gy of (137)Cs gamma rays. Two-dimensional electrophoresis gel coupled with mass spectrometry was used to analyze blood-samples collected at days 2 and 7 post-irradiation. At each sacrifice-time, alterations in expression-level of protein spots between control- and exposed-groups were analyzed statistically by the PDQuest software using Student's t-test (at the significance level of p < 0.05). Mass spectrometry was used to identify the identity of protein-spots with significantly altered expression-level. At day 2, 18 proteins were significantly up-regulated in exposed-mice. These included: alpha-2-Heremans-Schmid (HS)-glycoprotein, apolipoprotein (Apo)-AII-precursor, Apo-E, beta-2-glycoprotein-I, clusterin, fibrinogen-alpha-chain, fibrinogen-gamma-polypeptide, fetuin-B, haptoglobin, high-molecular-weight (HMW)-kininogen (Kng), low-MW-Kng, Kng1-precursor, liver-carboxylesterase-I-precursor, major-urinary-protein-6-precursor, mannose-binding-protein-C-precursor, mannose-binding-lectin-C, and prothrombin-precursor. Gelsolin was detected in control-mice only. At day 7, high expression-levels of 14 proteins were detected in control-mice (i.e., alpha-1-antitrypsin-precursor, carboxylesterase-N, cholesterol-7-alpha-hydroxylase, contraspin, coagulation-factor-II, coagulation-factor-XIII, gelsolin, immunoglobulin-G-heavy-chain, neurexin, prothrombin-precursor, protein-phosphatase, putative-calcium-influx-channel, vitamin-D-binding-protein, and 1110018G07Rik); while 15 proteins were highly expressed in exposed-mice. These included: alpha-1-acid-glycoprotein, alpha-2-HS-glycoprotein, alpha-1-protease-inhibitor-2, ApoA-IV, ApoC-I, ApoH, beta-1-globin, clusterin, complement-component-3, fibrinogen-beta-chain, HMW-Kng, major-histocompatibility-complex-class-Ia-H2-K, serine-(cysteine)-proteinase-inhibitor, retinoblastoma-associated-protein-140, and vascular-cell-adhesion-molecule-1. Although different proteins (mostly involved in inflammatory responses) were detected in exposed-mice, alterations in expression-levels of clusterin, gelsolin, kininogen, and alpha-2-HS-glycoproteins were found at both times. Despite the need for validation, the results suggested that alterations in expression-levels of specific proteins may be indicative of radiation-exposure. The results also provided the important step in an eventual establishment of blood-based biomarkers of radiation-exposure in vivo.

Acetylation and glycation of fibrinogen in vitro occur at specific lysine residues in a concentration dependent manner: A mass spectrometric and isotope labeling study

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

Svensson, Jan, E-mail: jan.svensson@ki.se; Karolinska Institutet, Department of Clinical Sciences, Danderyd Hospital, SE-182 88 Stockholm; Bergman, Ann-Charlotte

Highlights: Black-Right-Pointing-Pointer Fibrinogen was incubated in vitro with glucose or aspirin. Black-Right-Pointing-Pointer Acetylations and glycations were found at twelve lysine sites by mass spectrometry. Black-Right-Pointing-Pointer The labeling by aspirin and glucose occurred dose-dependently. Black-Right-Pointing-Pointer No competition between glucose and aspirin for binding to fibrinogen was found. -- Abstract: Aspirin may exert part of its antithrombotic effects through platelet-independent mechanisms. Diabetes is a condition in which the beneficial effects of aspirin are less prominent or absent - a phenomenon called 'aspirin resistance'. We investigated whether acetylation and glycation occur at specific sites in fibrinogen and if competition between glucose and aspirinmore » in binding to fibrinogen occurs. Our hypothesis was that such competition might be one explanation to 'aspirin resistance' in diabetes. After incubation of fibrinogen in vitro with aspirin (0.8 mM, 24 h) or glucose (100 mM, 5-10 days), we found 12 modified sites with mass spectrometric techniques. Acetylations in the {alpha}-chain: {alpha}K191, {alpha}K208, {alpha}K224, {alpha}K429, {alpha}K457, {alpha}K539, {alpha}K562, in the {beta}-chain: {beta}K233, and in the {gamma}-chain: {gamma}K170 and {gamma}K273. Glycations were found at {beta}K133 and {gamma}K75, alternatively {gamma}K85. Notably, the lysine 539 is a site involved in FXIII-mediated cross-linking of fibrin. With isotope labeling in vitro, using [{sup 14}C-acetyl]salicylic acid and [{sup 14}C]glucose, a labeling of 0.013-0.084 and 0.12-0.5 mol of acetylated and glycated adduct/mol fibrinogen, respectively, was found for clinically (12.9-100 {mu}M aspirin) and physiologically (2-8 mM glucose) relevant plasma concentrations. No competition between acetylation and glycation could be demonstrated. Thus, fibrinogen is acetylated at several lysine residues, some of which are involved in the cross-linking of fibrinogen. This may mechanistically explain why aspirin facilitates fibrin degradation. We find no support for the idea that glycation of fibrin(ogen) interferes with acetylation of fibrinogen.« less

Effects of Leucine Supplementation and Serum Withdrawal on Branched-Chain Amino Acid Pathway Gene and Protein Expression in Mouse Adipocytes

PubMed Central

Vivar, Juan C.; Knight, Megan S.; Pointer, Mildred A.; Gwathmey, Judith K.; Ghosh, Sujoy

2014-01-01

The essential branched-chain amino acids (BCAA), leucine, valine and isoleucine, are traditionally associated with skeletal muscle growth and maintenance, energy production, and generation of neurotransmitter and gluconeogenic precursors. Recent evidence from human and animal model studies has established an additional link between BCAA levels and obesity. However, details of the mechanism of regulation of BCAA metabolism during adipogenesis are largely unknown. We interrogated whether the expression of genes and proteins involved in BCAA metabolism are sensitive to the adipocyte differentiation process, and responsive to nutrient stress from starvation or BCAA excess. Murine 3T3-L1 preadipocytes were differentiated to adipocytes under control conditions and under conditions of L-leucine supplementation or serum withdrawal. RNA and proteins were isolated at days 0, 4 and 10 of differentiation to represent pre-differentiation, early differentiation and late differentiation stages. Expression of 16 BCAA metabolism genes was quantified by quantitative real-time PCR. Expression of the protein levels of branched-chain amino acid transaminase 2 (Bcat2) and branched-chain alpha keto acid dehydrogenase (Bckdha) was quantified by immunoblotting. Under control conditions, all genes displayed induction of gene expression during early adipogenesis (Day 4) compared to Day 0. Leucine supplementation resulted in an induction of Bcat2 and Bckdha genes during early and late differentiation. Western blot analysis demonstrated condition-specific concordance between gene and protein expression. Serum withdrawal resulted in undetectable Bcat2 and Bckdha protein levels at all timepoints. These results demonstrate that the expression of genes related to BCAA metabolism are regulated during adipocyte differentiation and influenced by nutrient levels. These results provide additional insights on how BCAA metabolism is associated with adipose tissue function and extends our understanding of the transcriptomic response of this pathway to variations in nutrient availability. PMID:25050624

Evolutionarily conserved TCR binding sites, identification of T cells in primary lymphoid tissues, and surprising trans-rearrangements in nurse shark.

PubMed

Criscitiello, Michael F; Ohta, Yuko; Saltis, Mark; McKinney, E Churchill; Flajnik, Martin F

2010-06-15

Cartilaginous fish are the oldest animals that generate RAG-based Ag receptor diversity. We have analyzed the genes and expressed transcripts of the four TCR chains for the first time in a cartilaginous fish, the nurse shark (Ginglymostoma cirratum). Northern blotting found TCR mRNA expression predominantly in lymphoid and mucosal tissues. Southern blotting suggested translocon-type loci encoding all four chains. Based on diversity of V and J segments, the expressed combinatorial diversity for gamma is similar to that of human, alpha and beta may be slightly lower, and delta diversity is the highest of any organism studied to date. Nurse shark TCRdelta have long CDR3 loops compared with the other three chains, creating binding site topologies comparable to those of mammalian TCR in basic paratope structure; additionally, nurse shark TCRdelta CDR3 are more similar to IgH CDR3 in length and heterogeneity than to other TCR chains. Most interestingly, several cDNAs were isolated that contained IgM or IgW V segments rearranged to other gene segments of TCRdelta and alpha. Finally, in situ hybridization experiments demonstrate a conservation of both alpha/beta and gamma/delta T cell localization in the thymus across 450 million years of vertebrate evolution, with gamma/delta TCR expression especially high in the subcapsular region. Collectively, these data make the first cellular identification of TCR-expressing lymphocytes in a cartilaginous fish.

Vascular endothelial growth factor and platelet-derived growth factor are potential angiogenic and metastatic factors in human breast cancer.

PubMed

Anan, K; Morisaki, T; Katano, M; Ikubo, A; Kitsuki, H; Uchiyama, A; Kuroki, S; Tanaka, M; Torisu, M

1996-03-01

Angiogenesis is a prerequisite for tumor growth and metastasis. Tumor angiogenesis may be mediated by several angiogenic factors such as vascular endothelial growth factor (VEGF), platelet-derived growth factor (PDGF), transforming growth factor-alpha, and basic fibroblast growth factor. Differential mRNA expressions of VEGF, PDGF (A chain), transforming growth factor-alpha and basic fibroblast growth factor in 32 primary invasive breast tumors were examined by reverse transcriptase-polymerase chain reaction. We analyzed relationships between mRNA expressions of these angiogenic factors and the degree of angiogenesis, tumor size, and metastasis. Quantification of angiogenesis was achieved by the immunohistochemical staining of endothelial cells with antibody to CD31. VEGF and PDGF-A mRNAs were expressed more frequently in breast tumors than in nontumor breast tissues, whereas no difference was found in expression frequency of either transforming growth factor-alpha or basic fibroblast growth factor mRNA. Vascular counts in tumors correlated with each expression frequency of VEGF and PDGF-A mRNA. PDGF-A mRNA was expressed more frequently in tumors with lymph node metastasis than in those without metastasis. Expression of VEGF and PDGF mRNAs detected by reverse transcriptase-polymerase chain reaction in breast tumors correlates with tumor-related characteristics of angiogenesis and metastatic potential. Analysis of these mRNAs by reverse transcriptase-polymerase chain reaction may be useful for assessing the biologic behavior of a breast tumor before surgical treatment.

Polymer translocation through a nanopore: a showcase of anomalous diffusion.

PubMed

Milchev, A; Dubbeldam, Johan L A; Rostiashvili, Vakhtang G; Vilgis, Thomas A

2009-04-01

We investigate the translocation dynamics of a polymer chain threaded through a membrane nanopore by a chemical potential gradient that acts on the chain segments inside the pore. By means of diverse methods (scaling theory, fractional calculus, and Monte Carlo and molecular dynamics simulations), we demonstrate that the relevant dynamic variable, the transported number of polymer segments, s(t), displays an anomalous diffusive behavior, both with and without an external driving force being present. We show that in the absence of drag force the time tau, needed for a macromolecule of length N to thread from the cis into the trans side of a cell membrane, scales as tauN(2/alpha) with the chain length. The anomalous dynamics of the translocation process is governed by a universal exponent alpha= 2/(2nu + 2 - gamma(1)), which contains the basic universal exponents of polymer physics, nu (the Flory exponent) and gamma(1) (the surface entropic exponent). A closed analytic expression for the probability to find s translocated segments at time t in terms of chain length N and applied drag force f is derived from the fractional Fokker-Planck equation, and shown to provide analytic results for the time variation of the statistical moments and . It turns out that the average translocation time scales as tau proportional, f(-1)N(2/alpha-1). These results are tested and found to be in perfect agreement with extensive Monte Carlo and molecular dynamics computer simulations.

Preparation of intact monomeric collagen from rat tail tendon and skin and the structure of the nonhelical ends in solution.

PubMed

Chandrakasan, G; Torchia, D A; Piez, K A

1976-10-10

Procedures for the preparation of soluble collagen from rat skin and tail tendon were reviewed and revised to permit the preparation of native monomeric collagen with intact nonhelical ends. The degree of intactness was estimated from the tyrosine content, which is present only in the nonhelical ends, and by mobility of the COOH-terminal cyanogen bromide peptide of the alpha1 chain on sodium dodecyl sulfate gels. The amount of covalently cross-linked polymeric material present was estimated by molecular sieve chromatography of denatured samples. Rapid purification in the cold was sufficient to prevent or greatly reduce proteolytic alteration. Fractionation by salt precipitation at acid pH was effective in reducing the content of polymeric material. Rat tail tendon yielded completely intact native collagen, but some high molecular weight aggregates remained. Collagen from the skin of lathyritic rats was easier to obtain free of aggregates, but contained about 1 less tyrosine residue per alpha1 chain even when isolated in the presence of enzyme inhibitors. Proton NMR spectra of denatured acidic solutions of these preparations showed that 4 to 5 tyrosine residues per alpha chain were present, confirming the chemical analysis. Spectra of the native molecule showed that about the same number of tyrosine residues per chain are in rapid motion, unlike residues in the helical portion of the molecule, a result which shows that the nonhelical ends of the native molecule are unstructured in acidic solution.

Binding of cationic pentapeptides with modified side chain lengths to negatively charged lipid membranes: Complex interplay of electrostatic and hydrophobic interactions.

PubMed

Hoernke, Maria; Schwieger, Christian; Kerth, Andreas; Blume, Alfred

2012-07-01

Basic amino acids play a key role in the binding of membrane associated proteins to negatively charged membranes. However, side chains of basic amino acids like lysine do not only provide a positive charge, but also a flexible hydrocarbon spacer that enables hydrophobic interactions. We studied the influence of hydrophobic contributions to the binding by varying the side chain length of pentapeptides with ammonium groups starting with lysine to lysine analogs with shorter side chains, namely omithine (Orn), alpha, gamma-diaminobutyric acid (Dab) and alpha, beta-diaminopropionic acid (Dap). The binding to negatively charged phosphatidylglycerol (PG) membranes was investigated by calorimetry, FT-infrared spectroscopy (FT-IR) and monolayer techniques. The binding was influenced by counteracting and sometimes compensating contributions. The influence of the bound peptides on the lipid phase behavior depends on the length of the peptide side chains. Isothermal titration calorimetry (ITC) experiments showed exothermic and endothermic effects compensating to a different extent as a function of side chain length. The increase in lipid phase transition temperature was more significant for peptides with shorter side chains. FTIR-spectroscopy revealed changes in hydration of the lipid bilayer interface after peptide binding. Using monolayer techniques, the contributions of electrostatic and hydrophobic effects could clearly be observed. Peptides with short side chains induced a pronounced decrease in surface pressure of PG monolayers whereas peptides with additional hydrophobic interactions decreased the surface pressure much less or even lead to an increase, indicating insertion of the hydrophobic part of the side chain into the lipid monolayer.

Twist-writhe partitioning in a coarse-grained DNA minicircle model

NASA Astrophysics Data System (ADS)

Sayar, Mehmet; Avşaroǧlu, Barış; Kabakçıoǧlu, Alkan

2010-04-01

Here we present a systematic study of supercoil formation in DNA minicircles under varying linking number by using molecular-dynamics simulations of a two-bead coarse-grained model. Our model is designed with the purpose of simulating long chains without sacrificing the characteristic structural properties of the DNA molecule, such as its helicity, backbone directionality, and the presence of major and minor grooves. The model parameters are extracted directly from full-atomistic simulations of DNA oligomers via Boltzmann inversion; therefore, our results can be interpreted as an extrapolation of those simulations to presently inaccessible chain lengths and simulation times. Using this model, we measure the twist/writhe partitioning in DNA minicircles, in particular its dependence on the chain length and excess linking number. We observe an asymmetric supercoiling transition consistent with experiments. Our results suggest that the fraction of the linking number absorbed as twist and writhe is nontrivially dependent on chain length and excess linking number. Beyond the supercoiling transition, chains of the order of one persistence length carry equal amounts of twist and writhe. For longer chains, an increasing fraction of the linking number is absorbed by the writhe.

Monte Carlo simulation and equation of state for flexible charged hard-sphere chain fluids: Polyampholyte and polyelectrolyte solutions

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

Jiang, Hao; Adidharma, Hertanto, E-mail: adidharm@uwyo.edu

The thermodynamic modeling of flexible charged hard-sphere chains representing polyampholyte or polyelectrolyte molecules in solution is considered. The excess Helmholtz energy and osmotic coefficients of solutions containing short polyampholyte and the osmotic coefficients of solutions containing short polyelectrolytes are determined by performing canonical and isobaric-isothermal Monte Carlo simulations. A new equation of state based on the thermodynamic perturbation theory is also proposed for flexible charged hard-sphere chains. For the modeling of such chains, the use of solely the structure information of monomer fluid for calculating the chain contribution is found to be insufficient and more detailed structure information must thereforemore » be considered. Two approaches, i.e., the dimer and dimer-monomer approaches, are explored to obtain the contribution of the chain formation to the Helmholtz energy. By comparing with the simulation results, the equation of state with either the dimer or dimer-monomer approach accurately predicts the excess Helmholtz energy and osmotic coefficients of polyampholyte and polyelectrolyte solutions except at very low density. It also well captures the effect of temperature on the thermodynamic properties of these solutions.« less

Interleukin 4: signalling mechanisms and control of T cell differentiation.

PubMed

Paul, W E

1997-01-01

Interleukin 4 (IL-4) is a pleiotropic type I cytokine that controls both growth and differentiation among haemopoietic and non-haemopoietic cells. Its receptor is a heterodimer. One chain, the IL-4R alpha chain, binds IL-4 with high affinity and determines the nature of the biochemical signals that are induced. The second chain, gamma c, is required for the induction of such signals. IL-4-mediated growth depends upon activation events that involve phosphorylation of Y497 of IL-4R alpha, leading to the binding and phosphorylation of 4PS/IRS-2 in haemopoietic cells and of IRS-1 in non-haemopoietic cells. By contrast, IL-4-mediated differentiation events depend upon more distal regions of the IL-4R alpha chain that include a series of STAT-6 binding sites. The distinctive roles of these receptor domains was verified by receptor-reconstruction experiments. The 'growth' and 'differentiation' domains of the IL-4R alpha chain, independently expressed as chimeric structures with a truncated version of the IL-2R beta chain, were shown to convey their functions to the hybrid receptor. The critical role of STAT-6 in IL-4-mediated gene activation and differentiation was made clear by the finding that lymphocytes from STAT-6 knockout mice are strikingly deficient in these functions but have retained the capacity to grow, at least partially, in response to IL-4. IL-4 plays a central role in determining the phenotype of naive CD4+ T cells. In the presence of IL-4, newly primed naive T cells develop into IL-4 producers while in its absence they preferentially become gamma-interferon (IFN-gamma) producers. Recently, a specialized subpopulation of T cells, CD4+/NK1.1+ cells, has been shown to produce large amounts of IL-4 upon stimulation. Two examples of mice with deficiencies in these cells are described--beta 2-microglobulin knockout mice and SJL mice. Both show defects in the development of IL-4-producing cells and in the increase in serum IgE in response to stimulation with the polyclonal stimulant anti-IgD. Both sets of mice have major diminutions in the number of CD4+/ NK1.1+ T cells, strongly indicating an important role of these cells in some but not all IgE responses to physiologic stimuli.

Leucine and alpha-Ketoisocaproic acid, but not norleucine, stimulate skeletal muscle protein synthesis in neonatal pigs

USDA-ARS?s Scientific Manuscript database

The branched-chain amino acid, leucine, acts as a nutrient signal to stimulate protein synthesis in skeletal muscle of young pigs. However, the chemical structure responsible for this effect has not been identified. We have shown that the other branched-chain amino acids, isoleucine and valine, are ...

The analysis of predictability of recent alpha decay formulae and the alpha partial half-lives of some exotic nuclei

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

Dasgupta-Schubert, N.; Reyes, M. A.; Tamez, V. A.

2009-04-20

Alpha decay is one of the two main decay modes of the heaviest nuclei, (SHE), and constitutes one of the dominant decay modes of highly neutron deficient medium mass nuclei ('exotics'). Thus identifying and characterizing the alpha decay chains form a crucial part of the identification of SHE. We report the extension of the previously developed method for the detailed and systematic investigation of the reliability of the three main extant analytical formulae of alpha decay half-lives: the generalized liquid drop model based formula of Royer et al. (FR), the Sobiczewski modified semi-empirical Viola-Seaborg formula (VSS) and the recent phenomenologicalmore » formula of Sobiczewski and Parkhomenko (SP)« less

Aplastic crisis revealing the diagnosis of Hb Evans [alpha62(E11)Val-->Met, GTG-->ATG (alpha2)] in a Hispanic kindred: case report and review.

PubMed

Steiner, Laurie A; Van Hoff, Jack; Kutlar, Ferdane; Gallagher, Patrick G

2007-01-01

This report describes a Hispanic girl who presented in aplastic crisis due to parvovirus infection. She was subsequently found to have a chronic hemolytic anemia secondary to an unstable hemoglobinopathy. Genetic testing revealed that the girl and two symptomatic family members had Hb Evans [alpha62(E11)Val-->Met, GTG-->ATG (alpha2)], an unstable hemoglobin (Hb) variant due to a mutation in the alpha2-globin chain. Hb Evans has been described only once previously, in a Caucasian kindred. Literature review indicates aplastic crisis is a rare initial presentation of unstable hemoglobinopathies and that these disorders are infrequent but important causes of hemolytic anemia in Hispanic patients.

The A-chain of insulin contacts the insert domain of the insulin receptor. Photo-cross-linking and mutagenesis of a diabetes-related crevice.

PubMed

Huang, Kun; Chan, Shu Jin; Hua, Qing-xin; Chu, Ying-Chi; Wang, Run-ying; Klaproth, Birgit; Jia, Wenhua; Whittaker, Jonathan; De Meyts, Pierre; Nakagawa, Satoe H; Steiner, Donald F; Katsoyannis, Panayotis G; Weiss, Michael A

2007-11-30

The contribution of the insulin A-chain to receptor binding is investigated by photo-cross-linking and nonstandard mutagenesis. Studies focus on the role of Val(A3), which projects within a crevice between the A- and B-chains. Engineered receptor alpha-subunits containing specific protease sites ("midi-receptors") are employed to map the site of photo-cross-linking by an analog containing a photoactivable A3 side chain (para-azido-Phe (Pap)). The probe cross-links to a C-terminal peptide (residues 703-719 of the receptor A isoform, KTFEDYLHNVVFVPRPS) containing side chains critical for hormone binding (underlined); the corresponding segment of the holoreceptor was shown previously to cross-link to a Pap(B25)-insulin analog. Because Pap is larger than Val and so may protrude beyond the A3-associated crevice, we investigated analogs containing A3 substitutions comparable in size to Val as follows: Thr, allo-Thr, and alpha-aminobutyric acid (Aba). Substitutions were introduced within an engineered monomer. Whereas previous studies of smaller substitutions (Gly(A3) and Ser(A3)) encountered nonlocal conformational perturbations, NMR structures of the present analogs are similar to wild-type insulin; the variant side chains are accommodated within a native-like crevice with minimal distortion. Receptor binding activities of Aba(A3) and allo-Thr(A3) analogs are reduced at least 10-fold; the activity of Thr(A3)-DKP-insulin is reduced 5-fold. The hormone-receptor interface is presumably destabilized either by a packing defect (Aba(A3)) or by altered polarity (allo-Thr(A3) and Thr(A3)). Our results provide evidence that Val(A3), a site of mutation causing diabetes mellitus, contacts the insert domain-derived tail of the alpha-subunit in a hormone-receptor complex.

Identification of a second murine interleukin-11 receptor alpha-chain gene (IL11Ra2) with a restricted pattern of expression.

PubMed

Robb, L; Hilton, D J; Brook-Carter, P T; Begley, C G

1997-03-15

The interleukin-11 receptor alpha-chain, a member of the hematopoietin receptor superfamily, forms, together with gp130, a functional high-affinity receptor complex for interleukin 11. We, and others, reported the cloning of the murine interleukin 11 receptor alpha-chain cDNA (IL11Ra) and recently described the structure of the IL11Ra locus. We also described the presence of a second IL11Ra-like locus in some mouse strains. In this study we report that the second locus, designated IL11Ra2, encodes an mRNA species. The transcript was 99% identical to the IL11Ra transcript in the coding and 3'-untranslated region, but had a different 5'-untranslated region. The complete genomic organization of the IL11Ra2 locus is presented, and the two loci are shown to be located on a 200-kb NaeI genomic fragment. Comparison of the expression pattern of the IL11Ra and IL11Ra2 genes using an RT-PCR restriction fragment length polymorphism strategy revealed that while the expression of IL11Ra was widespread, expression of IL11Ra2 was restricted to testis, lymph node, and thymus.

ENU mutagenesis identifies mice with cardiac fibrosis and hepatic steatosis caused by a mutation in the mitochondrial trifunctional protein beta-subunit.

PubMed

Kao, Hsiao-Jung; Cheng, Ching-Feng; Chen, Yen-Hui; Hung, Shuen-Iu; Huang, Cheng-Chih; Millington, David; Kikuchi, Tateki; Wu, Jer-Yuarn; Chen, Yuan-Tsong

2006-12-15

Using the metabolomics-guided screening coupled to N-ethyl-N-nitrosourea-mediated mutagenesis, we identified mice that exhibited elevated levels of long-chain acylcarnitines. Whole genome homozygosity mapping with 262 SNP markers mapped the disease gene to chromosome 5 where candidate genes Hadha and Hadhb, encoding the mitochondria trifunctional protein (MTP) alpha- and beta-subunits, respectively, are located. Direct sequencing revealed a normal alpha-subunit, but detected a nucleotide T-to-A transversion in exon 14 (c.1210T>A) of beta-subunit (Hadhb) which resulted in a missense mutation of methionine to lysine (M404K). Western blot analysis showed a significant reduction of both the alpha- and beta-subunits, consistent with reduced enzyme activity in both the long-chain 3-hydroxyacyl-CoA dehydrogenase and the long-chain 3-ketoacyl-CoA thiolase activities. These mice had a decreased weight gain and cardiac arrhythmias which manifested from a prolonged PR interval to a complete atrio-ventricular dissociation, and died suddenly between 9 and 16 months of age. Histopathological studies showed multifocal cardiac fibrosis and hepatic steatosis. This mouse model will be useful to further investigate the mechanisms underlying arrhythmogenesis relating to lipotoxic cardiomyopathy and to investigate pathophysiology and treatment strategies for human MTP deficiency.

Quantification of antibody (IgY) titers in hen eggs following immunization and their use in detecting cell surface molecules on nitrocellulose membranes.

PubMed

Ruan, Guang-Ping; Ma, Li; Meng, Xiao-Jing; Meng, Min-Jie; Wang, Xiao-Ning; Lin, Ying; Wu, Zheng-Qiang; He, Xiao-Wei; Wang, Ju-Fang; Zhu, Yong

2007-01-01

HLA-A*0201 alpha chain and beta2m were expressed from a prokaryotic system, and after refolding and purification, the alpha chain and beta2m were used to immunize eight laying hens. The titer of egg yolk antibody against alpha chain increased from 10(2) to 10(5.3) The titer of egg yolk antibody against beta2m increased from 10(1) to 10(4.7). The extent of titer increase is similar between the two antigens. An average of 135 mg purified polyclonal antibody (IgY) can be easily obtained from one egg yolk. The use of egg collection rather than serum collection is compatible with modern animal protection regulations. An average of 28 eggs were obtained from a laying hen every month, with a total amount of 3780 mg immunoglobulin extracted from one immunized hen every month, which would be equivalent to 630 mL of serum or 1260 mL of blood per month. Chickens are an optimal host for the production of polyclonal antibodies with high titer and high yield. Purified IgY was labeled with horseradish peroxidase and reacted with PBMC on nitrocellulose membranes indicating that the antibody can bind to the native conformation of class I HLA molecule on PBMC.

Localization in human interleukin 2 of the binding site to the alpha chain (p55) of the interleukin 2 receptor.

PubMed Central

Sauvé, K; Nachman, M; Spence, C; Bailon, P; Campbell, E; Tsien, W H; Kondas, J A; Hakimi, J; Ju, G

1991-01-01

Human interleukin 2 (IL-2) analogs with defined amino acid substitutions were used to identify specific residues that interact with the 55-kDa subunit (p55) or alpha chain of the human IL-2 receptor. Analog proteins containing specific substitutions for Lys-35, Arg-38, Phe-42, or Lys-43 were inactive in competitive binding assays for p55. All of these analogs retained substantial competitive binding to the intermediate-affinity p70 subunit (beta chain) of the receptor complex. The analogs varied in ability to interact with the high-affinity p55/p70 receptor. Despite the lack of binding to p55, all analogs exhibited significant biological activity, as assayed on the murine CTLL cell line. The dissociation constants of Arg-38 and Phe-42 analogs for p70 were consistent with intermediate-affinity binding; the Kd values were not significantly affected by the presence of p55 in binding to the high-affinity IL-2 receptor complex. These results confirm the importance of the B alpha-helix in IL-2 as the locus for p55-receptor binding and support a revised model of IL-2-IL-2 receptor interaction. PMID:2052547

The three-dimensional structure of a T-cell antigen receptor V alpha V beta heterodimer reveals a novel arrangement of the V beta domain.

PubMed Central

Housset, D; Mazza, G; Grégoire, C; Piras, C; Malissen, B; Fontecilla-Camps, J C

1997-01-01

The crystal structure of a mouse T-cell antigen receptor (TCR) Fv fragment complexed to the Fab fragment of a specific anti-clonotypic antibody has been determined to 2.6 A resolution. The polypeptide backbone of the TCR V alpha domain is very similar to those of other crystallographically determined V alphas, whereas the V beta structure is so far unique among TCR V beta domains in that it displays a switch of the c" strand from the inner to the outer beta-sheet. The beta chain variable region of this TCR antigen-binding site is characterized by a rather elongated third complementarity-determining region (CDR3beta) that packs tightly against the CDR3 loop of the alpha chain, without leaving any intervening hydrophobic pocket. Thus, the conformation of the CDR loops with the highest potential diversity distinguishes the structure of this TCR antigen-binding site from those for which crystallographic data are available. On the basis of all these results, we infer that a significant conformational change of the CDR3beta loop found in our TCR is required for binding to its cognate peptide-MHC ligand. PMID:9250664

Renal receptors for atrial and C-type natriuretic peptides in the rat.

PubMed

Brown, J; Zuo, Z

1992-07-01

Receptors for alpha-atrial natriuretic peptide (alpha-ANP) and C-type natriuretic peptide [CNP-(1-22)] were quantified in kidneys from adult Wistar rats by in vitro autoradiography. 125I-labeled alpha-ANP (100 pM) bound reversibly to glomeruli, outer medullary vasa recta, and inner medulla with an apparent dissociation constant (Kd) of 3-6 nM. The presence of 10 microM des-[Gln18,Ser19,Gly20,Leu21,Gly22]ANP-(4- 23) (C-ANP), a specific ligand of the ANPR-C subtype of alpha-ANP receptor, inhibited approximately 50% of the glomerular binding of 125I-alpha-ANP, and this moiety of glomerular binding was also inhibited by CNP-(1-22) with an apparent inhibitory constant (Ki) of 10.47 +/- 7.59 nM. C-ANP and CNP-(1-22) showed little affinity for the medullary binding sites of alpha-ANP. 125I-[Tyr0]CNP-(1-22) (110 pM) bound solely to glomeruli and was competitively displaced by increasing concentrations of [Tyr0]CNP-(1-22) with an apparent Kd of 1.42 +/- 0.48 nM. Binding of increasing concentrations (25 pM to 1 nM) of 125I-[Tyr0]CNP-(1-22) in the presence or absence of 1 microM [Tyr0]CNP-(1-22) also demonstrated a high affinity (Kd of 0.41 +/- 0.07 nM) for the glomerular binding of 125I-[Tyr0]CNP-(1-22). Bound 125I-[Tyr0]CNP-(1-22) could be displaced by excess alpha-ANP and excess CNP-(1-22), both with high affinities. The glomerular binding of 125I-[Tyr0]CNP-(1-22) was also prevented by 10 microM C-ANP. Guanosine 3',5'-cyclic monophosphate produced by isolated glomeruli was measured by radioimmunoassay.(ABSTRACT TRUNCATED AT 250 WORDS)

Phase equilibria and formation of vesicles of dioleoylphosphatidylcholine in glycerol/water mixtures.

PubMed

Johansson, L B; Kalman, B; Wikander, G; Fransson, A; Fontell, K; Bergenståhl, B; Lindblom, G

1993-07-04

The lipid 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) forms a lamellar liquid crystalline phase (L alpha) in arbitrary mixtures of glycerol and water. The phase has been characterized by means of X-ray diffraction, 31P-NMR spectroscopy and differential scanning calorimetry (DSC). In the L alpha state, and for DOPC concentrations greater than 50% (w/w), the thickness of the lipid bilayer decreases, while the area of the polar head group increases with increasing glycerol concentration. The phase transition from gel to L alpha state occurs in the range of 240 to 260 K. Contrary to a previous (McDaniel, R.V., McIntosh, T.J. and Simon, S.A. (1983) Biochim. Biophys. Acta 731, 97) study of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) we find that in the gel state, the thickness of the DOPC lipid bilayer is greater than that in the L alpha state. This suggests that in the gel state, the lipid acyl chains of DOPC are in extended configuration. The lamellar phase reaches its maximum swelling at about 50% (w/w) of DOPC. At lower DOPC concentrations a two-phase system is formed where the lamellar phase exists in equilibrium with excess of solvent. Unilamellar vesicles can be prepared from a diluted suspension of the lamellar phase either by using the sonicator or extruder technique. We show this by means of 31P-NMR, EPR and fluorescence spectroscopy. The mean radius of the vesicles, prepared by a sonicator, has been determined at different glycerol/water mixtures. It is found to decrease continuously from 100 A at 100% water to a minimum of 75 A at about 50% water in the solvent mixture. By further decreasing the water content in the solution, the radius rapidly increases, and a mean radius of 450 A is estimated at a water content of 10%. The rotational relaxation times of a fluorescent probe and two EPR spin probes, solubilized in DOPC vesicles, have been measured at different glycerol/water mixtures. It is found that the rotational rates are always much slower in the systems containing glycerol.

Comparative genomic and phylogenetic analysis of short-chain dehydrogenases/reductases with dual retinol/sterol substrate specificity.

PubMed

Belyaeva, Olga V; Kedishvili, Natalia Y

2006-12-01

Human short-chain dehydrogenases/reductases with dual retinol/sterol substrate specificity (RODH-like enzymes) are thought to contribute to the oxidation of retinol for retinoic acid biosynthesis and to the metabolism of androgenic and neuroactive 3alpha-hydroxysteroids. Here, we investigated the phylogeny and orthology of these proteins to understand better their origins and physiological roles. Phylogenetic and genomic analysis showed that two proteins (11-cis-RDH and RDHL) are highly conserved, and their orthologs can be identified in the lower taxa, such as amphibians and fish. Two other proteins (RODH-4 and 3alpha-HSD) are significantly less conserved. Orthologs for 3alpha-HSD are present in all mammals analyzed, whereas orthologs for RODH-4 can be identified in some mammalian species but not in others due to species-specific gene duplications. Understanding the evolution and divergence of RODH-like enzymes in various vertebrate species should facilitate further investigation of their in vivo functions using animal models.

CAWSES Related Projects in Japan : Grant-in-Aid for Creative Scientific Research ügBasic Study of Space Weather Predictionüh and CHAIN (Continuous H Alpha Imaging Network)

NASA Astrophysics Data System (ADS)

Shibata, K.; Kurokawa, H.

The Grant-in-Aid for Creative Scientific Research of the Ministry of Education Science Sports Technology and Culture of Japan The Basic Study of Space Weather Prediction PI K Shibata Kyoto Univ has started in 2005 as 5 years projects with total budget 446Myen The purpose of this project is to develop a physical model of solar-terrestrial phenomena and space storms as a basis of space weather prediction by resolving fundamental physics of key phenomena from solar flares and coronal mass ejections to magnetospheric storms under international cooperation program CAWSES Climate and Weather of the Sun-Earth System Continuous H Alpha Imaging Network CHAIN Project led by H Kurokawa is a key project in this space weather study enabling continuous H alpha full Sun observations by connecting many solar telescopes in many countries through internet which provides the basis of the study of space weather prediction

Interleukin-9 enhances interleukin-5 receptor expression, differentiation, and survival of human eosinophils.

PubMed

Gounni, A S; Gregory, B; Nutku, E; Aris, F; Latifa, K; Minshall, E; North, J; Tavernier, J; Levit, R; Nicolaides, N; Robinson, D; Hamid, Q

2000-09-15

Interleukin-9 (IL-9) has been implicated in the pathogenesis of allergic disorders. To examine the interaction between IL-9 and eosinophils, we evaluated mature peripheral blood eosinophils for their expression of the specific alpha-subunit of the IL-9 receptor (IL-9R-alpha). The expression of IL-9R-alpha by human eosinophils was detected at the messenger RNA (mRNA) and protein levels by reverse transcriptase-polymerase chain reaction (RT-PCR), flow cytometry, and immunocytochemical analysis, respectively. Functional analyses demonstrated that recombinant human (rh)IL-9 inhibited in vitro peripheral blood human eosinophil apoptosis in a concentration-dependent manner. We then examined the role of IL-9 in eosinophil differentiation using the human cord blood CD34(+) cells and human promyelocytic leukemia cells (HL-60). The addition of IL-9 to CD34(+) cells cultured in IL-3 and IL-5 enhanced eosinophil development, and IL-9 alone induced the expression of IL-5R-alpha. IL-9 also up-regulated the IL-5R-alpha chain cell surface expression during terminal eosinophil differentiation of the HL-60 cell line. Our findings suggest that IL-9 may potentiate in vivo eosinophil function by increasing their survival and IL-5-mediated differentiation and maturation. Taken together, these results suggest a mechanism by which IL-9 potentiates airway and tissue eosinophilia.

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

Zhang, Ying; Li, Jianguo, E-mail: 2010lijianguo@sina.cn

Highlights: Black-Right-Pointing-Pointer Carbachol reduced the lipopolysaccharide-induced intestinal barrier breakdown. Black-Right-Pointing-Pointer Carbachol ameliorated the lipopolysaccharide-induced ileal tight junction damage. Black-Right-Pointing-Pointer Carbachol prevented the LPS-induced NF-{kappa}{beta} and myosin light-chain kinase activation. Black-Right-Pointing-Pointer Carbachol exerted its beneficial effects in an {alpha}7 nicotinic receptor-dependent manner. -- Abstract: Carbachol is a cholinergic agonist that protects the intestines after trauma or burn injury. The present study determines the beneficial effects of carbachol and the mechanisms by which it ameliorates the lipopolysaccharide (LPS)-induced intestinal barrier breakdown. Rats were injected intraperitoneally with 10 mg/kg LPS. Results showed that the gut barrier permeability was reduced, the ultrastructural disruption ofmore » tight junctions (TJs) was prevented, the redistribution of zonula occludens-1 and claudin-2 proteins was partially reversed, and the nuclear factor-kappa beta (NF-{kappa}{beta}) and myosin light-chain kinase (MLCK) activation in the intestinal epithelium were suppressed after carbachol administration in LPS-exposed rats. Pretreatment with the {alpha}7 nicotinic acetylcholine receptor ({alpha}7nAchR) antagonist {alpha}-bungarotoxin blocked the protective action of carbachol. These results suggested that carbachol treatment can protect LPS-induced intestinal barrier dysfunction. Carbachol exerts its beneficial effect on the amelioration of the TJ damage by inhibiting the NF-{kappa}{beta} and MLCK pathways in an {alpha}7nAchR-dependent manner.« less

21 CFR 178.3780 - Polyhydric alcohol esters of long chain monobasic acids.

Code of Federal Regulations, 2013 CFR

2013-04-01

... chloride copolymer articles complying with § 177.1980 of this chapter that contact food of Types I, II, IV... 1,050 to 1,700. The esters are produced by the reaction of either ethylene glycol or glycerol with... chain alpha-olefins, the unreacted carboxylic acids in the formation of the glycerol esters being...

21 CFR 178.3780 - Polyhydric alcohol esters of long chain monobasic acids.

Code of Federal Regulations, 2010 CFR

2010-04-01

... chloride copolymer articles complying with § 177.1980 of this chapter that contact food of Types I, II, IV... 1,050 to 1,700. The esters are produced by the reaction of either ethylene glycol or glycerol with... chain alpha-olefins, the unreacted carboxylic acids in the formation of the glycerol esters being...

21 CFR 178.3780 - Polyhydric alcohol esters of long chain monobasic acids.

Code of Federal Regulations, 2012 CFR

2012-04-01

... chloride copolymer articles complying with § 177.1980 of this chapter that contact food of Types I, II, IV... 1,050 to 1,700. The esters are produced by the reaction of either ethylene glycol or glycerol with... chain alpha-olefins, the unreacted carboxylic acids in the formation of the glycerol esters being...

21 CFR 178.3780 - Polyhydric alcohol esters of long chain monobasic acids.

Code of Federal Regulations, 2011 CFR

2011-04-01

... chloride copolymer articles complying with § 177.1980 of this chapter that contact food of Types I, II, IV... 1,050 to 1,700. The esters are produced by the reaction of either ethylene glycol or glycerol with... chain alpha-olefins, the unreacted carboxylic acids in the formation of the glycerol esters being...

The energy content of restaurant foods without stated calorie information.

PubMed

Urban, Lorien E; Lichtenstein, Alice H; Gary, Christine E; Fierstein, Jamie L; Equi, Ashley; Kussmaul, Carolyn; Dallal, Gerard E; Roberts, Susan B

2013-07-22

National recommendations for the prevention and treatment of obesity emphasize reducing energy intake through self-monitoring food consumption. However, little information is available on the energy content of foods offered by nonchain restaurants, which account for approximately 50% of restaurant locations in the United States. To measure the energy content of foods from independent and small-chain restaurants that do not provide stated information on energy content. We used bomb calorimetry to determine the dietary energy content of the 42 most frequently purchased meals from the 9 most common restaurant categories. Independent and small-chain restaurants were randomly selected, and 157 individual meals were analyzed. Area within 15 miles of downtown Boston. A random sample of independent and small-chain restaurants. Dietary energy. All meal categories provided excessive dietary energy. The mean energy content of individual meals was 1327 (95% CI, 1248-1406) kcal, equivalent to 66% of typical daily energy requirements. We found a significant effect of food category on meal energy (P ≤ .05), and 7.6% of meals provided more than 100% of typical daily energy requirements. Within-meal variability was large (average SD, 271 kcal), and we found no significant effect of restaurant establishment or size. In addition, meal energy content averaged 49% greater than those of popular meals from the largest national chain restaurants (P < .001) and in subset analyses contained 19% more energy than national food database information for directly equivalent items (P < .001). National chain restaurants have been criticized for offering meals with excess dietary energy. This study finds that independent and small-chain restaurants, which provide no nutrition information, also provide excessive dietary energy in amounts apparently greater than popular meals from chain restaurants or information in national food databases. A national requirement for accurate calorie labeling in all restaurants may discourage menus offering unhealthy portions and would allow consumers to make informed choices about ordering meals that promote weight gain and obesity.

Long-chain n-3 PUFA: plant v. marine sources.

PubMed

Williams, Christine M; Burdge, Graham

2006-02-01

Increasing recognition of the importance of the long-chain n-3 PUFA, EPA and DHA, to cardiovascular health, and in the case of DHA to normal neurological development in the fetus and the newborn, has focused greater attention on the dietary supply of these fatty acids. The reason for low intakes of EPA and DHA in most developed countries (0.1-0.5 g/d) is the low consumption of oily fish, the richest dietary source of these fatty acids. An important question is whether dietary intake of the precursor n-3 fatty acid, alpha-linolenic acid (alphaLNA), can provide sufficient amounts of tissue EPA and DHA by conversion through the n-3 PUFA elongation-desaturation pathway. alphaLNA is present in marked amounts in plant sources, including green leafy vegetables and commonly-consumed oils such as rape-seed and soyabean oils, so that increased intake of this fatty acid would be easier to achieve than via increased fish consumption. However, alphaLNA-feeding studies and stable-isotope studies using alphaLNA, which have addressed the question of bioconversion of alphaLNA to EPA and DHA, have concluded that in adult men conversion to EPA is limited (approximately 8%) and conversion to DHA is extremely low (<0.1%). In women fractional conversion to DHA appears to be greater (9%), which may partly be a result of a lower rate of utilisation of alphaLNA for beta-oxidation in women. However, up-regulation of the conversion of EPA to DHA has also been suggested, as a result of the actions of oestrogen on Delta6-desaturase, and may be of particular importance in maintaining adequate provision of DHA in pregnancy. The effect of oestrogen on DHA concentration in pregnant and lactating women awaits confirmation.

Increased levels of advanced glycation end products positively correlate with iron overload and oxidative stress markers in patients with β-thalassemia major.

PubMed

Mirlohi, Maryam Sadat; Yaghooti, Hamid; Shirali, Saeed; Aminasnafi, Ali; Olapour, Samaneh

2018-04-01

The impaired biosynthesis of the β-globin chain in β-thalassemia leads to the accumulation of unpaired alpha globin chains, failure in hemoglobin formation, and iron overload due to frequent blood transfusion. Iron excess causes oxidative stress and massive tissue injuries. Advanced glycation end products (AGEs) are harmful agents, and their production accelerates in oxidative conditions. This study was conducted on 45 patients with major β-thalassemia who received frequent blood transfusions and chelation therapy and were compared to 40 healthy subjects. Metabolic parameters including glycemic and iron indices, hepatic and renal functions tests, oxidative stress markers, and AGEs (carboxymethyl-lysine and pentosidine) levels were measured. All parameters were significantly increased in β-thalassemia compared to the control except for glutathione levels. Blood glucose, iron, serum ferritin, non-transferrin-bound iron (NTBI), MDA, soluble form of low-density lipoprotein receptor, glutathione peroxidase, total reactive oxygen species (ROS), and AGE levels were significantly higher in the β-thalassemia patients. Iron and ferritin showed a significant positive correlation with pentosidine (P < 0.01) but not with carboxymethyl-lysine. The NTBI was markedly increased in the β-thalassemia patients, and its levels correlated significantly with both carboxymethyl-lysine and pentosidine (P < 0.05). Our findings confirm the oxidative status generated by the iron overload in β-thalassemia major patients and highlight the enhanced formation of AGEs, which may play an important role in the pathogenesis of β-thalassemia major.

Alpha-helix to beta-sheet transition in long-chain poly-l-lysine: Formation of alpha-helical fibrils by poly-l-lysine.

PubMed

Cieślik-Boczula, Katarzyna

2017-06-01

The temperature-induced α-helix to β-sheet transition in long-chain poly-l-lysine (PLL), accompanied by the gauche-to-trans isomerization of CH 2 groups in the hydrocarbon side chains of Lys amino acid residues, and formation of β-sheet as well as α-helix fibrillar aggregates of PLL have been studied using Fourier-transform infrared (FT-IR) and vibrational circular dichroism (VCD) spectroscopy, and transmission electron microscopy (TEM). In a low-temperature alkaline water solution or in a methanol-rich water mixture, the secondary structure of PLL is represented by α-helical conformations with unordered and gauche-rich hydrocarbon side chains. Under these conditions, PLL molecules aggregate into α-helical fibrils. PLLs dominated by extended antiparallel β-sheet structures with highly ordered trans-rich hydrocarbon side chains are formed in a high-temperature range at alkaline pD and aggregate into fibrillar, protofibrillar, and spherical forms. Presented data support the idea that fibrillar aggregation is a varied phenomenon possible in repetitive structural elements with not only a β-sheet-rich conformation, but also an α-helical-rich conformation. Copyright © 2017 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.

Specific reaction of alpha,beta-unsaturated carbonyl compounds such as 6-shogaol with sulfhydryl groups in tubulin leading to microtubule damage.

PubMed

Ishiguro, Kazuhiro; Ando, Takafumi; Watanabe, Osamu; Goto, Hidemi

2008-10-15

6-Shogaol and 6-gingerol are ginger components with similar chemical structures. However, while 6-shogaol damages microtubules, 6-gingerol does not. We have investigated the molecular mechanism of 6-shogaol-induced microtubule damage and found that the action of 6-shogaol results from the structure of alpha,beta-unsaturated carbonyl compounds. alpha,beta-Unsaturated carbonyl compounds such as 6-shogaol react with sulfhydryl groups of cysteine residues in tubulin, and impair tubulin polymerization. The reaction with sulfhydryl groups depends on the chain length of alpha,beta-unsaturated carbonyl compounds. In addition, alpha,beta-unsaturated carbonyl compounds are more reactive with sulfhydryl groups in tubulin than in 2-mercaptoethanol, dithiothreitol, glutathione and papain, a cysteine protease.

[A compound heterozygosity mutation in the interleukin-7 receptor-alpha gene resulted in severe combined immunodeficiency in a Chinese patient].

PubMed

Zhang, Zhi-yong; Zhao, Xiao-dong; Wang, Mo; Yu, Jie; An, Yun-fei; Yang, Xi-qiang

2009-09-01

Mutation in the interleukin-7 receptor-alpha (IL-7R alpha) chain causes a rare type of severe combined immunodeficiency (SCID) with presence of NK cells in the peripheral blood. Here we report the molecular and clinical characterization of a compound heterozygosity mutation in the interleukin-7 receptor-alpha gene that resulted in SCID in a patient firstly from China. A 5 month-old male patient and his parents were enrolled in this study. Since 15 days of age, the patient had had recurrent fever, persistent cough and diarrhea. He was in poor general condition with pyorrhea and ulceration of the BCG scar. His brother died of severe infection at 4 months of age. He was initially diagnosed as SCID according to clinical manifestation and immunological analysis. A panel of SCID candidate genes including IL-2RG, RAG1/RAG2 and IL-7R alpha of patient and his parents were amplified by polymerase chain reaction (PCR) from genomic DNA. Reverse transcription polymerase chain reaction (RT-PCR) was used to amplify the IL-7R alpha transcripts. Sequencing was performed directly on the PCR products forward and reversely. The serum immunoglobulin (Ig) profile was IgG 6867 mg/L (normal range, 3050 - 8870 mg/L); IgM 206 mg/L and IgA 249 mg/L, IgE 2.3 IU/ml (normal range < 150 IU/ml). The patient was treated with IVIG previously. There were no T-cells but increased percentage of B-cells (58%) and NK cells (42%) in the peripheral blood was found. Needle biopsies from enlarged axillary lymph node was identified positive for Mycobacterium bovis under microscope and by culture. The patient had a compound heterozygosity mutation in the IL-7R alpha gene:on one allele, there was a splice-junction mutation in intron 4 (intron 4(+1)G > A), for which his father was a carrier; whereas on the other allele, a nonsense mutation at position 638 in exon 5 with a premature stop codon (638 C > T, R206X) was identified, for which his mother was a carrier. The splice-junction mutation in intron 4 of IL-7R alpha was firstly reported. The IL-7R alpha mRNA expression of the patient was remarkably reduced whereas the parents had relatively normal IL-7R alpha mRNA expression. IL-7R alpha cDNA of the patient was amplified by nested PCR. The PCR products were purified, cloned with a TA Cloning Kit and sequenced directly. A 64 bp deletion was found in exon 4 of IL-7R alpha. No mutation was found in IL-2RG and RAG1/RAG2 of the patient and his parents. This is the first case with a compound heterozygosity mutation in the IL-7R receptor alpha gene and T-B+NK+ phenotype from China. Intron 4(+1)G > A was a novel mutation.

Biological agents for controlling excessive scarring.

PubMed

Berman, Brian

2010-01-01

The potential of various biological agents to reduce or prevent excessive scar formation has now been evaluated in numerous in-vitro studies, experimental animal models and preliminary clinical trials, in some cases with particularly promising results. Perhaps prominent among this group of biological agents, and, to some degree, possibly representing marketed compounds already being used 'off label' to manage excessive scarring, are the tumor necrosis factor alpha antagonist etanercept, and immune-response modifiers such as IFNalpha2b and imiquimod. Additional assessment of these novel agents is now justified with a view to reducing or preventing hypertrophic scars, keloid scars and the recurrence of post-excision keloid lesions.

Clinical and laboratory features of patients with an inherited deficiency of neutrophil membrane complement receptor type 3 (CR3) and the related membrane antigens LFA-1 and p150,95.

PubMed

Ross, G D

1986-03-01

Over the last 3 years a group of more than 20 patients has been described worldwide who have a similar history of recurrent bacterial infections and an inherited deficiency of three related leukocyte membrane surface antigens known as CR3, LFA-1 (lymphocyte function-associated antigen type 1), and p150,95 (function unknown). These antigens share a common beta-chain structure linked noncovalently to one of three distinct alpha-chain types. It is believed that the patients with this disease have a reduced or absent ability to synthesize the common beta subunit of the antigen family, resulting in absent or reduced expression of all three antigen family members on different leukocyte types. Neutrophils have a reduced phagocytic and respiratory burst response to bacteria and yeast as well as a reduced ability to adhere to various substrates and migrate into sites of infection. In vitro functional studies of normal neutrophils, monocytes, and lymphocytes treated with monoclonal antibodies to the individual alpha and beta chains of these antigens suggest that most of the clinical features of the patients may be due to the neutrophil and monocyte deficiency of CR3. Although natural killer-cell activity is diminished or absent, no immune deficiency of the patients' lymphocytes attributable to the absence of LFA-1 has been detected. Diagnosis of this disease has been facilitated by the commercial availability of monoclonal antibodies specific for the alpha chains of CR3 and p150,95.

Complementary DNA sequences encoding the multimammate rat MHC class II DQ alpha and beta chains and cross-species sequence comparison in rodents.

PubMed

de Bellocq, J Goüy; Leirs, H

2009-09-01

Sequences of the complete open reading frame (ORF) for rodents major histocompatibility complex (MHC) class II genes are rare. Multimammate rat (Mastomys natalensis) complementary DNA (cDNA) encoding the alpha and beta chains of MHC class II DQ gene was cloned from a rapid amplifications of cDNA Emds (RACE) cDNA library. The ORFs consist of 801 and 771 bp encoding 266 and 256 amino acid residues for DQB and DQA, respectively. The genomic structure of Mana-DQ genes is globally analogous to that described for other rodents except for the insertion of a serine residue in the signal peptide of Mana-DQB, which is unique among known rodents.

Amide-based inhibitors of p38alpha MAP kinase. Part 2: design, synthesis and SAR of potent N-pyrimidyl amides.

PubMed

Tester, Richland; Tan, Xuefei; Luedtke, Gregory R; Nashashibi, Imad; Schinzel, Kurt; Liang, Weiling; Jung, Joon; Dugar, Sundeep; Liclican, Albert; Tabora, Jocelyn; Levy, Daniel E; Do, Steven

2010-04-15

Optimization of a tri-substituted N-pyridyl amide led to the discovery of a new class of potent N-pyrimidyl amide based p38alpha MAP kinase inhibitors. Initial SAR studies led to the identification of 5-dihydrofuran as an optimal hydrophobic group. Additional side chain modifications resulted in the introduction of hydrogen bond interactions. Through extensive SAR studies, analogs bearing free amino groups and alternatives to the parent (S)-alpha-methyl benzyl moiety were identified. These compounds exhibited improved cellular activities and maintained balance between p38alpha and CYP3A4 inhibition. Copyright 2010 Elsevier Ltd. All rights reserved.

Observation of the anisotropy of 10 TeV primary cosmic ray nuclei flux with the Super-Kamiokande-I detector

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

Guillian, G.; Kibayashi, A.; Learned, J. G.

2007-03-15

The relative sidereal variation in the arrival direction of primary cosmic ray nuclei of median energy 10 TeV was measured using downward, through-going muons detected with the Super-Kamiokande-I detector. The projection of the anisotropy map onto the right ascension axis has a first harmonic amplitude of (6.64{+-}0.98 stat{+-}0.55 syst)x10{sup -4} and a phase at maximum at (33.2 deg. {+-}8.2 deg. stat{+-}5.1 deg. syst) right ascension. A sky map in equatorial coordinates indicates an excess region in the constellation of Taurus and a deficit region toward Virgo. The excess region is centered at ({alpha}{sub T},{delta}{sub T})=(75 deg. {+-}7 deg., -5 deg.more » {+-}9 deg.) with a half-opening angle {chi}{sub T}=(39{+-}7) deg.; the excess flux is (0.104{+-}0.020)% above the isotropic expectation. The corresponding parameters for the deficit region are ({alpha}{sub V},{delta}{sub V})=(205 deg. {+-}7 deg., 5 deg. {+-}10 deg.), {chi}{sub V}=(54{+-}7) deg., and (-0.094{+-}0.014)%. The data do not allow us to rule out a pure dipole form for the anisotropy (allowed at 13% confidence level); they are better described by the excess and deficit cones described above. We explored the implications under the assumption that the true anisotropy is not distorted too much by the analysis filter so that it is well-described by the observed excess and deficit cones.« less

Unusual Nonterrestrial L-proteinogenic Amino Acid excesses in the Tagish Lake Meteorite

NASA Technical Reports Server (NTRS)

Glavin, Daniel P.; Elsila, Jamie E.; Burton, Aaron S.; Callahan, Michael P.; Dworkin, Jason P.; Hilts, Robert W.; Herd, D. K.

2012-01-01

The distribution and isotopic and enantiomeric compositions of amino acids found in three distinct fragments of the Tagish Lake C2-type carbonaceous chondrite were investigated via liquid chromatography with fluorescence detection and time-of-flight mass spectrometry and gas chromatography isotope ratio mass spectrometry. Large L-enantiomeric excesses (L(sub ee) approximately 43-59%) of the alpha-hydrogen aspartic and glutamic amino acids were measured in Tagish Lake, whereas alanine, another alpha hydrogen protein amino acid, was found to be nearly racemic (D much approximately L) using both techniques. Carbon isotope measurements of D- and L-aspartic acid and 1)- and L-alanine in Tagish Lake fall well outside of the terrestrial range and indicate that the measured aspartic acid enantioenrichment is indigenous to the meteorite. Alternate explanations for the L-excesses of aspartic acid such as interference from other compounds present in the sample, analytical biases, or terrestrial amino acid contamination were investigated and rejected. These results can be explained by differences in the solid-solution phase behavior of aspartic acid, which can form conglomerate enantiopure solids during crystallization, and alanine, which can only form racemic crystals. Amplification of a small initial L-enantiomer excess during aqueous alteration on the meteorite parent body could have led to the large L-enrichments observed for aspartic acid and other conglomerate amino acids in Tagish Lake. The detection of non terrestrial L-proteinogenic amino acid excesses in the Tagish Lake meteorite provides support for the hypothesis that significant enantiomeric enrichments for some amino acids could form by abiotic processes prior to the emergence of life.

Zinc Binding and Dimerization of Streptococcus pyogenes Pyrogenic Exotoxin C Are Not Essential for T-Cell Stimulation

DTIC Science & Technology

2003-03-14

streptococcal superantigen binding to MHCII on the surface of cells (7–9), suggesting an essential role in both MHCII molecular recognition and TCR-mediated...extent, mutations of side chains found in a second conserved MHCII alpha-chain-binding site consisting of a hydrophobic surface loop decreased T-cell...fraction of dimer is present at T-cell stimulatory concentrations of Spe-C following mutation of the unpaired side chain of cys- teine at residue 27 to

Novel Humanized mice to test Therapeutics for Human Type 1 Diabetes

DTIC Science & Technology

2014-01-06

performed in non- obese diabetic (NOD) mice, the closest animal model for human T1D, have identified different immune cells involved in pancreatic β-cell...periphery. Each MHC class II chain contributes to the formation of a groove where the peptide is embedded (78). 17 The polygenetic factor underlying...diabetes mellitus in non- obese diabetic mice by transgenes encoding modified I-A beta-chain or normal I-E alpha-chain. Nature 345:727-9 63. Marek

Genotyping of alpha-thalassemia deletions using multiplex polymerase chain reactions and gold nanoparticle-filled capillary electrophoresis.

PubMed

Chen, Yen-Ling; Shih, Chi-Jen; Ferrance, Jerome; Chang, Ya-Sian; Chang, Jan-Gowth; Wu, Shou-Mei

2009-02-13

A gold nanoparticle-filled capillary electrophoresis method combined with three multiplex polymerase chain reactions (PCRs) was established for simultaneous diagnosis of five common alpha-thalassemia deletions, including the -alpha(3.7) deletion, -alpha(4.2) deletion, Southeast Asian (--(SEA)), Filipino (--(FIL)) and Thai (--(THAI)) deletions. Gold nanoparticles (GNPs) were used as a pseudostationary phase to improve the resolution between DNA fragments in a low-viscosity polymer. To achieve the best CE separation, several parameters were evaluated for optimizing the separation conditions, including the capillary coating, the concentrations of polymer sieving matrix, the sizes and concentrations of GNPs, the buffer concentrations, and the pH. The final CE method for separating a 200-base pair (bp) DNA ladder and alpha-thalassemia deletions used a DB-17 capillary, 0.6% poly(ethylene oxide) (PEO) prepared in a mixture of GNP(32nm) solution and glycine buffer (25mM, pH 9.0) (80:20, v/v) as the sieving matrix with 1microM YO-PRO-1 for fluorescence detection; the applied voltage was -10kV (detector at anode side) and the separation temperature was 25 degrees C. Under these optimal conditions, 15 DNA fragments with sizes ranging from 0.2kb to 3.0kb were resolved within 11.5min. The RSDs of migration times were less than 2.81%. A total of 21 patients with alpha-thalassemia deletions were analyzed using this method, and all results showed good agreement with those obtained by gel electrophoresis.

An algorithm for converting a virtual-bond chain into a complete polypeptide backbone chain

NASA Technical Reports Server (NTRS)

Luo, N.; Shibata, M.; Rein, R.

1991-01-01

A systematic analysis is presented of the algorithm for converting a virtual-bond chain, defined by the coordinates of the alpha-carbons of a given protein, into a complete polypeptide backbone. An alternative algorithm, based upon the same set of geometric parameters used in the Purisima-Scheraga algorithm but with a different "linkage map" of the algorithmic procedures, is proposed. The global virtual-bond chain geometric constraints are more easily separable from the loal peptide geometric and energetic constraints derived from, for example, the Ramachandran criterion, within the framework of this approach.

Proteome analysis of the cardiac and pericardial tissue of Biomphalaria tenagophila populations susceptible and resistant to Schistosoma mansoni infection.

PubMed

Jannotti-Passos, Liana K; Andrade, Hélida M; Caldeira, Roberta L; Romanha, Alvaro J; Murta, Silvane M F; Chapeaurouge, Donat A; Perales, Jonas; Coelho, Paulo Marcos Z; Carvalho, Omar S

2008-03-01

For a better comprehension of the parasite-host interaction, proteins expressed by the cardiac and pericardial tissues were compared between susceptible (Cabo Frio) and resistant (Taim) Biomphalaria tenagophila populations, challenged (c) and non-challenged (nc) with Schistosoma mansoni. Proteins were separated by two-dimensional gel electrophoresis (2DE) and stained with Coomassie blue. A total of 146 and 135 spots were observed in Cabo Frio (CFnc) and in Taim (Tnc) non-challenged populations, respectively, whereas 153 spots were detected in both Cabo Frio (CFc) and Taim (Tc) challenged populations. Regarding comparisons between CFnc and CFc, the numbers of exclusive spots obtained were one and nine, respectively, whereas Tnc yielded 17 and Tc eight exclusive spots. By comparing the total of spots in CF (nc+c) with T (nc+c) populations, we obtained: four exclusive spots for CFc; zero for CFnc; four for Tc and; one for Tnc. A quantitative comparison (reason>2.5) of the total spots of CF (nc+c) with T (nc+c) populations allowed us to distinguish five more intense spots for Tc, 14 for Tnc, 15 for CFnc and 11 for CFc. In the CFnc population, two proteins were identified: actin and ATP synthase alpha chain; in the CFc population, four proteins: actin, calmodulin, HSP70, and dehydrogenase; in the Tnc population, five proteins: matrilin, HSP70, actin, ATP synthase alpha chain and intermediate filament of the protein; and in the Tc population, three proteins: actin, alpha-S1 casein and ATP synthase alpha chain. Out of a total of 79 spots, only nine proteins were identified due to the low number of available nucleotide sequences in the GenBank. Nevertheless, knowing proteins regarded as differentially expressed is indispensable for hitherto unidentified genes implicated in B. tenagophila resistance and or susceptibility to S. mansoni infection.

The C-terminal region of alpha-crystallin: involvement in protection against heat-induced denaturation

NASA Technical Reports Server (NTRS)

Takemoto, L.; Emmons, T.; Horwitz, J.; Spooner, B. S. (Principal Investigator)

1993-01-01

Recent studies have demonstrated that the alpha-crystallins can protect other proteins against heat-induced denaturation and aggregation. To determine the possible involvement of the C-terminal region in this activity, the alpha-crystallins were subjected to limited tryptic digestion, and the amount of cleavage from the N-terminal and C-terminal regions of the alpha-A and alpha-B crystallin chains was assessed using antisera specific for these regions. Limited tryptic digestion resulted in cleavage only from the C-terminal region of alpha-A crystallin. This trypsin-treated alpha-A crystallin preparation showed a decreased ability to protect proteins from heat-induced aggregation using an in vitro assay. Together, these results demonstrate that the C-terminal region of alpha-A crystallin is important for its ability to protect against heat-induced aggregation, which is consistent with the hypothesis that post-translational changes that are known to occur at the C-terminal region may have significant effects on the ability of alpha-A crystallin to protect against protein denaturation in vivo.

Determination of the parameters controlling swelling of chemically cross-linked pH-sensitive poly(N-vinylimidazole) hydrogels.

PubMed

Molina, M Jesús; Gómez-Antón, M Rosa; Piérola, Inés F

2007-10-25

The number of variables controlling the behavior of ionic gels is large and very often some of them are unknown. The aim of this work is to interpret quantitatively the swelling behavior of pH sensitive gels, with the minimum number of simplifying assumptions. With this purpose, the equilibrium degree of swelling (S) and protonation (alpha) of chemically cross-linked poly(N-vinylimidazole) (PVI) immersed in aqueous salt solutions were measured as a function of the ionic strength (mu), in the whole range of pH. In acid solutions with pH in the range 0 to 4, imidazole moieties become protonated, and PVI behaves as a polyelectrolyte gel: S decreases upon increasing mu both for NaCl and for CaCl(2), with HCl as protonating acid. In aqueous solutions with larger pH, between 4 and 12, the hydrogel is practically neutral, and S increases as mu rises, showing a salting-in effect. From the quantitative analysis of these results, the following facts emerged. Protonation induces chain stiffness (as measured by the non-Gaussian factor) and worsening of the solvent quality of the aqueous media (as measured by the polymer-solvent interaction parameter). For alpha below 33%, swelling seems to be governed by the excess of mobile counterions inside the gel with respect to the bath, with a minor but still significantly negative contribution of the osmotic swelling pressure due to polymer-solvent mixing. Above 33% protonation, it is necessary to consider Manning counterion condensation to get parameters with physical meaning. The crossover between polyelectrolyte and salting-in effects corresponds to alpha and mu values with the same ionic and mixing contributions to the osmotic swelling pressure. The formation of ionic nonpermanent cross-links, with H(2)SO(4) as the protonating acid, was discarded.

Mapping High-Velocity H-alpha and Lyman-alpha Emission from Supernova 1987A

NASA Technical Reports Server (NTRS)

France, Kevin; McCray, Richard; Fransson, Claes; Larsson, Josefin; Frank, Kari A.; Burrows, David N.; Challis, Peter; Kirshner, Robert P.; Chevalier, Roger A.; Garnavich, Peter;

Polymorphism in 'L' shaped lipids: structure of N-, O-diacylethanolamines with mixed acyl chains.

PubMed

Tarafdar, Pradip K; Swamy, Musti J

2009-11-01

Although solid state polymorphism in lipids has been established by spectroscopic and calorimetric studies long ago, only in a few cases crystal structures of different polymorphs of the same compound have been reported, possibly due to difficulties in obtaining high quality single crystals of individual polymorphs. Recent studies show that N-, O-diacylethanolamines (DAEs) can be derived by the O-acylation of the stress-related lipids, the N-acylethanolamines under physiological conditions. In this study, two DAEs with mixed acyl chains, namely N-palmitoyl, O-octanoylethanolamine and N-palmitoyl, O-decanoylethanolamine have been synthesized and their three-dimensional structures were determined. Both the compounds were found to adopt 'L' shaped structures and exist in two polymorphic forms, alpha and beta. In the alpha form a mixed-type chain packing has been observed whereas in the beta form the chain packing is symmetric. Similar polymorphic forms are likely to exist in other 'L' shaped lipids such as 1,3-diacylglycerols and ceramides, where polymorphism has been detected earlier, but three-dimensional structures - which can give precise information about the packing at atomic resolution - have not been reported.

The electrophoretically 'slow' and 'fast' forms of the alpha 2-macroglobulin molecule.

PubMed Central

Barrett, A J; Brown, M A; Sayers, C A

1979-01-01

alpha 2-Macroglobulin (alpha 2M) was isolated from human plasma by a four-step procedure: poly(ethylene glyco) fractionation, gel chromatography, euglobulin precipitation and immunoadsorption. No contaminants were detected in the final preparations by electrophoresis or immunoprecipitation. The protein ran as a single slow band in gel electrophoresis, and was designated 'S-alpha 2M'. S-alpha 2M bound about 2 mol of trypsin/mol. Treatment of S-alpha 2M with a proteinase or ammonium salts produced a form of the molecule more mobile in electrophoresis, and lacking proteinase-binding activity (F-alpha 2M). The electrophoretic mobility of the F-alpha 2M resulting from reaction with NH4+ salts was identical with that of proteinase complexes. We attribute the change in electrophoretic mobility of the alpha 2M to a conformation change, but there was no evidence of a change in pI or Strokes radius. Electrophoresis of S-alpha 2M in the presence of sodium dodecylsulphate gave results consistent with the view that the alpha 2M molecule is a tetramer of identical subunits, assembled as a non-covalent pair of disulphide-linked dimers. Some of the subunits seemed to be 'nicked' into two-thires-length and one-third-length chains, however. This was not apparent with F-alpha 2M produced by ammonium salts. F-alpha 2M produced by trypsin showed two new bands attributable to cleavage of the subunit polypeptide chain near the middle. Immunoassays of F-alpha 2M gave 'rockets' 12-29% lower than those with S-alpha 2M. The nature of the interactions between subunits in S-alpha 2M and F-alpha 2M was investigated by treating each form with glutaraldehyde before electrophoresis in the presence of sodium dodecyl sulphate. A much greater degree of cross-linking was observed with the F-alpha 2M, indicating that the subunits interact most closely in this form of the molecule. Exposure of S-alpha 2M to 3 M-urea or pH3 resulted in dissociation to the disulphide-bonded half-molecules; these did not show the proteinase-binding activity characteristic of the intact alpha 2M. F-alpha 2M was less easily dissociated than was S-alpha 2M. S-alpha 2M was readily dissociated to the quarter-subunits by mild reduction, with the formation of 3-4 new thiol groups per subunit. Inact reactive alpha 2M could then be regenerated in high yield by reoxidation of the subunits. F-alpha 2M formed by reaction with a proteinase or ammonium salts was not dissociated under the same conditions, although the interchain disulphide bonds were reduced. If the thiol groups of the quarter-subunits of S-alpha 2M were blocked by carboxymethylation, oxidative reassociation did not occur. Nevertheless treatment of these subunits with methylammonium salts or a proteinase caused the reassembly of half-molecules and intact (F-) tetramers. It is emphasized that F-alpha 2M does not have the properties of a denatured form of the protein... Images Fig. 3. Fig. 4. Fig. 5. Fig. 6. PMID:91367

Action of transglucosidase from Aspergillus niger on maltoheptaose and [U-(13)C]maltose.

PubMed

Ota, Masafumi; Okamoto, Takeshi; Wakabayashi, Hidehiko

2009-03-10

Oligosaccharides synthesized from a mixture of maltoheptaose and [U-(13)C]maltose with transglucosidase [EC 2.4.1.24] from Aspergillus niger were investigated. When the reaction mixture was incubated at 15 degrees C for 1h, several types of oligosaccharides with DP (degree of polymerization) 2 to DP8 containing alpha-D-Glcp-(1-->6)-maltoheptaose were detected by liquid chromatography-mass spectrometry (LC-MS) and methylation analysis. Most of these compounds consisted of alpha-(1-->4) linkages in the main chain and alpha-(1-->6) linkages at the non-reducing ends. However, when the reaction mixture was incubated for 96h, most of these products were converted into oligosaccharides with DP2 to DP5 consisting of only alpha-(1-->6) linkages. These results suggested that A. niger transglucosidase rapidly transferred glucosyl residues to maltooligosaccharides, and gradually hydrolyzed both alpha-(1-->4) linkages and alpha-(1-->6) linkages at the non-reducing end, and transformed these into smaller molecules of mainly alpha-(1-->6) linkages.

[Trofosides A and B and other cytostatic steroid-derived compounds from the Far East starfish Trofodiscus über].

PubMed

Levina, E V; Kalinovskiĭ, A I; Andriiashchenko, P V; Menzorova, N I; Dmitrenok, P S

2007-01-01

Three new polar steroids identified as trofoside A, (20R,24S)-24-O-(3-O-methyl-beta-D-xylopyranosyl)-3beta,6alpha,8,15beta,24-pentahydroxy-5alpha-cholestane, its 22(23)-dehydro derivative (trofoside B), and 15-sulfoxy-(20R,24S)-5alpha-cholestane-3beta,6beta,8,15alpha,24-pentaol sodium salt, were isolated from Trofodiscus uber starfish extracts collected in the Sea of Okhotsk. Two known compounds, trofoside A aglycone, (20R,24S)-3beta,6alpha,8,15beta,24-pentahydroxy-5alpha-cholestane, and triseramide, (20R,24R,25S,22E)-24-methyl-3beta,6alpha,8,15beta-tetrahydroxy-5alpha-cholest-22-en-27-oic acid (2-sulfoethyl)amide sodium salt, were also found. The structures of the isolated polyoxysteroids were established from their spectra. Minimal concentrations causing degradation of unfertilized egg-cells of the sea-urchin Strongylocentrotus intermedius (C(min)) and terminating the cell division at the stage of the first division (C(min) embr.), as well as the concentrations causing 50% immobilization of sperm cells (ImC50) and inhibiting their ability to fertilize egg-cells by 50% (IC50) were determined for the isolated compounds. Of three compounds highly toxic in embryos and sea-urchin sperm cells, the polyol with a sulfo group in the steroid core was the most active; two glycosides with monosaccharide chains located at C3 and C24 atoms were less toxic. Note that all the compounds with the spermiotoxic activities differently affected the embryo development. The positions of monosaccharide residues in the core considerably influence the compound activity. For example, both mono- and double chained glycosides with the monosaccharide fragment at C3 and C24 atoms are active against sea-urchin sperm cells and embryos, whereas the C24 glycosylated trofoside A does not affect embryos and displays a poor spermiotoxicity.

[Immunohistochemistry contribution in Alport syndrome diagnosis].

PubMed

Krichen Makni, S; Kharrat, M; Ben Hmida, M; Chaker, H; Gubler, M-C; Antignac, C; Jlidi, R; Hachicha, J; Sellami Boudwara, T

2005-07-01

Alport syndrome (AS) is an hereditary disease characterised by the association of progressive hematuria nephritis. The diagnosis is based on clinical genetic and ultrastructural findings. Nowadays, immunohistochemical technique is of great interest. It enables us to analyze the distribution of the different chains of the type IV collagen in renal basement membrane (RBM) and epidermal basement membrane (EBM) which appeared to be abnormal in 70% of cases. We report a prospective study of five families affected with AS. Six patients were investigated by immunohistochemical studies of kidney (3 cases) and skin (6 cases) frozen specimens. Monoclonal antibodies recognizing the collagenous domain of alpha1 (MAB1), alpha3 (MAB3) and alpha5 (MAB5) chains of type IV collagen were used. Two methods were performed: direct immunofluorescence and immunohistochemical (ultravision) analysis. The different chains distribution of type IV collagen in the EBM and RBM was normal in four cases (4 men), abnormal in two patients (1 man and woman). Based on the clinical, genetical and immunohistochemical findings we established three transmission modes: autosomal recessive in two families, dominant X linked in two other familiales, and autosomal dominant in one family. Immunohistochemical studies is a simple technique of an easy interpretation accomplished on kidney frozen specimen, or even on a simple cutaneous biopsy. It could be very useful for the diagnosis and enables us in addition to determine the mode of transmission of AS.

Guanine nucleotide-binding regulatory proteins in retinal pigment epithelial cells

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

Jiang, Meisheng; Tran, V.T.; Fong, H.K.W.

1991-05-01

The expression of GTP-binding regulatory proteins (G proteins) in retinal pigment epithelial (RPE) cells was analyzed by RNA blot hybridization and cDNA amplification. Both adult and fetal human RPE cells contain mRNA for multiple G protein {alpha} subunits (G{alpha}) including G{sub s}{alpha}, G{sub i-1}{alpha}, G{sub i-2}{alpha}, G{sub i-3}{alpha}, and G{sub z}{alpha} (or G{sub x}{alpha}), where G{sub s} and G{sub i} are proteins that stimulate or inhibit adenylyl cyclase, respectively, and G{sub z} is a protein that may mediate pertussis toxin-insensitive events. Other G{alpha}-related mRNA transcripts were detected in fetal RPE cells by low-stringency hybridization to G{sub i-2}{alpha} and G{sub s}{alpha}more » protein-coding cDNA probes. The diversity of G proteins in RPE cells was further studied by cDNA amplification with reverse transcriptase and the polymerase chain reaction. This approach revealed that, besides the above mentioned members of the G{alpha} gene family, at least two other G{alpha} subunits are expressed in RPE cells. Human retinal cDNA clones that encode one of the additional G{alpha} subunits were isolated and characterized. The results indicate that this G{alpha} subunit belongs to a separate subfamily of G proteins that may be insensitive to inhibition by pertussis toxin.« less

An RNA-splicing mutation (G{sup +51VS20}) in the Type II collagen gene (COL2A1) in a family with spondyloepiphyseal dysplasia congenita

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

Tiller, G.E.; Polumbo, P.A.; Weis, M.A.

1995-02-01

Defects in type II collagen have been demonstrated in a phenotypic continuum of chondrodysplasias that includes achondrogenesis II, hypochondrogenesis, spondyloepiphyseal dysplasia congenita (SEDC), Kniest dysplasia, and Stickler syndrome. We have determined that cartilage from a terminated fetus with an inherited form of SEDC contained both normal {alpha}1(II) collagen chains and chains that lacked amino acids 256-273 of the triple-helical domain. PCR amplification of this region of COL2A1, from genomic DNA, yielded products of normal size, while amplification of cDNA yielded a normal sized species and a shorter fragment missing exon 20. Sequence analysis of genomic DNA from the fetus revealedmore » a G{yields}T transversion at position +5 of intron 20; the affected father was also heterozygous for the mutation. Allele-specific PCR and heteroduplex analysis of a VNTR in COL2A1 independently confirmed the unaffected status of a fetus in a subsequent pregnancy. Thermodynamic calculations suggest that the mutation prevents normal splicing of exon 20 by interfering with binding of U{sub 1} small-nuclear RNA to pre-mRNA, thus leading to skipping of exon 20 in transcripts from the mutant allele. Electron micrographs of diseased cartilage showed intracellular inclusion bodies, which were stained by an antibody to {alpha}1(II) procollagen. Our findings support the hypothesis that {alpha}-chain length alterations that preserve the Gly-X-Y repeat motif of the triple helix result in partial intracellular retention of {alpha}1(II) procollagen and produce mild to moderate chondrodysplasia phenotypes. 50 refs., 6 figs., 1 tab.« less

Supramolecular structure of the casein micelle.

PubMed

McMahon, D J; Oommen, B S

2008-05-01

The supramolecular structure of colloidal casein micelles in milk was investigated by using a sample preparation protocol based on adsorption of proteins onto a poly-l-lysine and parlodion-coated copper grid, staining of proteins and calcium phosphate by uranyl oxalate, instantaneous freezing, and drying under a high vacuum. High-resolution transmission electron microscopy stereo-images were obtained showing the interior structure of casein micelles. On the basis of our interpretation of these images, an interlocked lattice model was developed in which both casein-calcium phosphate aggregates and casein polymer chains act together to maintain casein micelle integrity. The caseins form linear and branched chains (2 to 5 proteins long) interlocked by the casein-stabilized calcium phosphate nanoclusters. This model suggests that stabilization of calcium phosphate nanoclusters by phosphoserine domains of alpha(s1)-, alpha(s2)-, or beta-casein, or their combination, would orient their hydrophobic domains outward, allowing interaction and binding to other casein molecules. Other interactions between the caseins, such as calcium bridging, could also occur and further stabilize the supramolecule. The combination of having an interlocked lattice structure and multiple interactions results in an open, sponge-like colloidal supramolecule that is resistant to spatial changes and disintegration. Hydrophobic interactions between caseins surrounding a calcium phosphate nanocluster would prevent complete dissociation of casein micelles when the calcium phosphate nanoclusters are solubilized. Likewise, calcium bridging and other electrostatic interactions between caseins would prevent dissociation of the casein micelles into casein-calcium phosphate nanocluster aggregates when milk is cooled or urea is added to milk, and hydrophobic interactions are reduced. The appearance of both polymer chains and small aggregate particles during milk synthesis would also be expected based on this interlocked lattice model of casein micelles, and its supramolecule structure thus exhibits the principles of self-aggregation, interdependence, and diversity observed in nature.

Structural characterization of alpha-terminal group of natural rubber. 1. Decomposition of branch-points by lipase and phosphatase treatments.

PubMed

Tarachiwin, Lucksanaporn; Sakdapipanich, Jitladda; Ute, Koichi; Kitayama, Tatsuki; Bamba, Takashi; Fukusaki, Ei-Ichiro; Kobayashi, Akio; Tanaka, Yasuyuki

2005-01-01

Deproteinized natural rubber latex (DPNR-latex) was treated with lipase and phosphatase in order to analyze the structure of the chain-end group (alpha-terminal). The enzymatic treatment decreased the content of long-chain fatty acid ester groups in DPNR from about 6 to 2 mol per rubber molecule. The molecular weight and intrinsic viscosity were reduced to about one-third after treatment with lipase and phosphatase. The Huggins' k' constant of the enzyme-treated DPNR showed the formation of linear rubber molecules. The molecular weight distribution of DPNR changed apparently after treatment with lipase and phosphatase. (1)H NMR spectrum of rubber obtained from DPNR-latex showed small signals due to monophosphate, di-phosphate and phospholipids at the alpha-terminus. Treatment of DPNR-latex with lipase and phosphatase decreased the relative intensity of the (1)H NMR signals corresponding to phospholipids, whereas no change was observed for the signals due to mono- and diphosphates. The residual mono- and diphosphate signals as well as some phospholipid signals after lipase and phosphatase treatments indicate that mono- and diphosphate groups are directly linked at the alpha-terminus with the modified structure, expected by aggregation or linking with phospholipid molecules.

Hydrophobic interactions in complexes of antimicrobial peptides with bacterial polysaccharides.

PubMed

Kuo, Hsin H; Chan, Celine; Burrows, Lori L; Deber, Charles M

2007-06-01

Biofilms of Pseudomonas aeruginosa are responsible for chronic lung infections in cystic fibrosis patients, where they are characterized by overproduction of the exopolysaccharide alginate and are recalcitrant to treatment with conventional antibiotics. Cationic antimicrobial peptides (CAPs) are potential alternatives for the treatment of multi-drug-resistant P. aeruginosa. However, alginate in P. aeruginosa biofilms has been proposed to bind these peptides through hydrophobic interactions, consequently reducing their activity [Chan et al., J Biol Chem 2004; 279: 38749-38754]. Here we perform biophysical analyses of the interactions of alginate with a series of novel peptide antibiotics (alpha-CAPs) of prototypic sequence KK-AAAXAAAAAXAAWAAXAAA-KKKK (where X = Phe, Trp or Leu). The hydrophobic interaction interface in alginate was investigated by examining (i) the effects of polysaccharide composition with respect to D-mannuronate and L-guluronate content; (ii) glycan chain length; (iii) alpha-CAP Trp fluorescence; and (iv) 1-anilinonaphthalene-8-sulfonate fluorescence. The results show that, while M and G residues produce equivalent effects, hydrophobic interactions between alginate and alpha-CAPs require a minimal glycan chain length. Peptide interactions with alginate are deduced to be mediated by hydrophobic microdomains comprised of pyranosyl C-H groups that are inducible upon formation of alpha-CAP-alginate complexes due to charge neutralization between the two species.

Excess Volumes and Excess Isentropic Compressibilities of Binary Liquid Mixtures of Trichloroethylene with Esters at 303.15 K

NASA Astrophysics Data System (ADS)

Ramanaiah, S.; Rao, C. Narasimha; Nagaraja, P.; Venkateswarlu, P.

2015-11-01

Exces volumes, VE, and excess isentropic compressibilities, κSE, have been reported as a function of composition for binary liquid mixtures of trichloroethylene with ethyl acetate, n-propyl acetate, and n-butyl acetate at 303.15 K. Isentropic compressibilities are calculated using measured sound speeds and density data for pure components and for binary mixtures. Excess volumes and excess isentropic compressibilities are found to be negative for the three systems studied over the entire composition range at 303.15 K, whereas these values become more negative with an increase of carbon chain length. The results are discussed in terms of intermolecular interactions between unlike molecules.

Naturally occurring disulfide-bound dimers of three-fingered toxins: a paradigm for biological activity diversification.

PubMed

Osipov, Alexey V; Kasheverov, Igor E; Makarova, Yana V; Starkov, Vladislav G; Vorontsova, Olga V; Ziganshin, Rustam Kh; Andreeva, Tatyana V; Serebryakova, Marina V; Benoit, Audrey; Hogg, Ronald C; Bertrand, Daniel; Tsetlin, Victor I; Utkin, Yuri N

2008-05-23

Disulfide-bound dimers of three-fingered toxins have been discovered in the Naja kaouthia cobra venom; that is, the homodimer of alpha-cobratoxin (a long-chain alpha-neurotoxin) and heterodimers formed by alpha-cobratoxin with different cytotoxins. According to circular dichroism measurements, toxins in dimers retain in general their three-fingered folding. The functionally important disulfide 26-30 in polypeptide loop II of alpha-cobratoxin moiety remains intact in both types of dimers. Biological activity studies showed that cytotoxins within dimers completely lose their cytotoxicity. However, the dimers retain most of the alpha-cobratoxin capacity to compete with alpha-bungarotoxin for binding to Torpedo and alpha7 nicotinic acetylcholine receptors (nAChRs) as well as to Lymnea stagnalis acetylcholine-binding protein. Electrophysiological experiments on neuronal nAChRs expressed in Xenopus oocytes have shown that alpha-cobratoxin dimer not only interacts with alpha7 nAChR but, in contrast to alpha-cobratoxin monomer, also blocks alpha3beta2 nAChR. In the latter activity it resembles kappa-bungarotoxin, a dimer with no disulfides between monomers. These results demonstrate that dimerization is essential for the interaction of three-fingered neurotoxins with heteromeric alpha3beta2 nAChRs.

29 CFR 1915.112 - Ropes, chains and slings.

Code of Federal Regulations, 2012 CFR

2012-07-01

..., defective welds, deformation and increase in length or stretch. (3) Employers must note interlink wear, not accompanied by stretch in excess of 5 percent, and remove the chain from service when maximum allowable wear... shall be removed from service when, due to stretch, the increase in length of a measured section exceeds...

29 CFR 1915.112 - Ropes, chains and slings.

Code of Federal Regulations, 2014 CFR

2014-07-01

..., defective welds, deformation and increase in length or stretch. (3) Employers must note interlink wear, not accompanied by stretch in excess of 5 percent, and remove the chain from service when maximum allowable wear... shall be removed from service when, due to stretch, the increase in length of a measured section exceeds...

29 CFR 1915.112 - Ropes, chains and slings.

Code of Federal Regulations, 2013 CFR

2013-07-01

..., defective welds, deformation and increase in length or stretch. (3) Employers must note interlink wear, not accompanied by stretch in excess of 5 percent, and remove the chain from service when maximum allowable wear... shall be removed from service when, due to stretch, the increase in length of a measured section exceeds...

Characterization of synthesis and storage of TGF-alpha in rat parotid acinar and intercalated duct cells.

PubMed

Login, G R; Yang, J; Bryan, K P; Digenis, E C; McBride, J; Elovic, A; Quissell, D O; Dvorak, A M; Wong, D T

1997-03-01

Although the expression and biological role of transforming growth factor-alpha (TGF-alpha) have been explored in a variety of normal cells in mammalian species, little is known about the storage of TGF-alpha in secretory cells of exocrine organs. Parotid glands from four rats were homogenized for RNA isolation followed by reverse transcription-polymerase chain reaction to determine the presence of TGF-alpha message. In situ hybridization using a hamster-specific TGF-alpha riboprobe was done on paraffin sections. Parotid gland and isolated acinar cells were processed for transmission electron microscopy (TEM) and postembedding immunogold labeled for TGF-alpha. Gold particles were counted on approximately 200 granules in 10 acinar cells and in 10 intercalated duct cells. Labeling density was calculated as the number of gold particles per square micrometer +/- SD. Statistical significance was calculated using one-way analysis of variance. Using multiple technologies, we have established that rat parotid acinar and intercalated duct cells synthesize TGF-alpha and store the precursor form of this cytokine in their secretory granules.

Characterization and charge distribution of the asparagine-linked oligosaccharides on secreted mouse thyrotropin and free alpha-subunits

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

Gesundheit, N.; Gyves, P.W.; DeCherney, G.S.

1989-06-01

Mouse hemipituitaries in vitro secrete TSH, composed of an alpha-beta heterodimer, as well as excess (free) alpha-subunits. By dual metabolic labeling with (35S)sulfate and (3H)mannose, we have characterized oligosaccharides from secreted TSH alpha, TSH beta, and free alpha-subunits released from the apoprotein by enzymatic deglycosylation. Oligosaccharides from each subunit displayed a distinct anion exchange HPLC profile due to a specific pattern of sialylation and sulfation. Six species were obtained from TSH alpha (with two glycosylation sites), including neutral oligosaccharides as well as those with one or two negative charges. For TSH beta (with one glycosylation site) at least eight oligosaccharidemore » species were noted, representing nearly every permutation of sialylation and sulfation; approximately 30% contained three or more negative charges. Analysis of (3H)mannose-labeled oligosaccharides on Concanavalin-A-agarose showed 85% binding for those from TSH alpha, 70% for free alpha, and 50% for those from TSH beta. These data demonstrate that oligosaccharides from secreted TSH beta were more sialylated and sulfated, consistent with a more complex branching pattern, than those from TSH alpha. Oligosaccharides from free alpha-subunit were more sialylated than those from TSH alpha, and the net negative charge was intermediate between those of TSH alpha and TSH beta. Although great microheterogeneity is present even at the single glycosylation site on the beta-subunit of secreted TSH, a pattern of sialylation and sulfation could be discerned.« less

Assessing the Origins of Aliphatic Amines in the Murchison Meteorite from their Compound-Specific Carbon Isotopic Ratios and Enantiomeric Composition

NASA Technical Reports Server (NTRS)

Aponte, Jose; Dworkin, Jason; Elsila, Jamie E.

2014-01-01

The study of meteoritic organic compounds provides a unique window into the chemical inventory of the early Solar System and prebiotic chemistry that may have been important for the origin of life on Earth. Multiple families of organic compounds have been extracted from the Murchison meteorite, which is one of the most thoroughly studied carbonaceous chondrites. The amino acids extracted from Murchison have been extensively analyzed, including measurements of non-terrestrial stable isotopic ratios and discoveries of L-enantiomeric excesses for alpha-dialkyl amino acids, notably isovaline. However, although the isotopic signatures of bulk amine-containing fractions have been measured, the isotopic ratios and enantiomeric composition of individual aliphatic amines, compounds that are chemically related to amino acids, remain unknown. Here, we report a novel method for the extraction, separation, identification and quantitation of aliphatic monoamines extracted from the Murchison meteorite. Our results show a complete suite of structural isomers, with a larger concentration of methylamine and ethylamine and decreasing amine concentrations with increasing carbon number. The carbon isotopic compositions of fourteen meteoritic aliphatic monoamines were measured, with delta C-13 values ranging from +21% to +129%, showing a decrease in C-13 with increasing carbon number, a relationship that may be consistent with the chain elongation mechanism under kinetic control previously proposed for meteoritic amino acids. We also found the enantiomeric composition of sec-butylamine, a structural analog to isovaline, was racemic within error, while the isovaline extracted from the same Murchison piece showed an L-enantiomeric excess of 9.7; this result suggested that processes leading to enantiomeric excess in the amino acid did not affect the amine. We used these collective data to assess the primordial synthetic origins of these meteoritic aliphatic amines and their potential linkage to meteoritic amino acids.

Hb Adana (HBA2 or HBA1: c.179G > A) and alpha thalassemia: Genotype-phenotype correlation.

PubMed

Singh, Sharon A; Sarangi, Susmita; Appiah-Kubi, Abena; Hsu, Peihong; Smith, W Byron; Gallagher, Patrick G; Glader, Bertil; Chui, David H K

2018-05-11

Alpha thalassemia due to nondeletional mutations usually leads to more severe disease than that caused by deletional mutations. Devastating outcomes such as hydrops fetalis can occur with two nondeletional mutations, therefore warranting DNA-based workup for suspected carriers with subtle hematological abnormalities for family counseling purposes. We describe three cases with hemoglobin (Hb) Adana, a nondeletional alpha chain mutation, compounded with an alpha globin gene deletion resulting in thalassemia intermedia. We review the literature, draw genotype-phenotype correlations from published cases of Hb Adana, and propose that this correlation can be used by clinicians to help direct diagnostic studies and urge hematologists to thoroughly workup high-risk patients. © 2018 Wiley Periodicals, Inc.

Effect of alpha-tocopherol on lipid peroxidation and antioxidant system in fibrosarcoma bearing rats.

PubMed

Vasavi, H; Thangaraju, M; Sachdanandam, P

1994-02-23

The anticarcinogenic activity of alpha-tocopherol (Vitamin E) was tried in fibrosarcoma induced rats through its antioxidative potential. The rate of formation of malondialdehyde (MDA), the end product of lipid peroxidation was analysed in alpha-tocopherol (400 mg/kg body weight) treated and untreated fibrosarcoma bearing rats with respective controls. The levels of non-enzymic antioxidants like, glutathione and vitamin E, and enzymic antioxidants viz., catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GSH-PX), and glutathione-S-transferase (GST) were assayed as well. Significantly increased (p < 0.001) level of lipid peroxide was observed with concomitant decreases in the level of enzymic and non-enzymic antioxidants in fibrosarcoma bearing rats when compared with control animals. In alpha-tocopherol supplemented animals, the corrected level of these parameters were observed likely to near normal values. Thus, alpha-tocopherol can be accepted to pose first line of defense mechanism against excessively formed reactive species due to impaired antioxidant systems in fibrosarcoma conditions, that cause membrane damage leading to deleterious effects.

Neurofeedback in Learning Disabled Children: Visual versus Auditory Reinforcement.

PubMed

Fernández, Thalía; Bosch-Bayard, Jorge; Harmony, Thalía; Caballero, María I; Díaz-Comas, Lourdes; Galán, Lídice; Ricardo-Garcell, Josefina; Aubert, Eduardo; Otero-Ojeda, Gloria

2016-03-01

Children with learning disabilities (LD) frequently have an EEG characterized by an excess of theta and a deficit of alpha activities. NFB using an auditory stimulus as reinforcer has proven to be a useful tool to treat LD children by positively reinforcing decreases of the theta/alpha ratio. The aim of the present study was to optimize the NFB procedure by comparing the efficacy of visual (with eyes open) versus auditory (with eyes closed) reinforcers. Twenty LD children with an abnormally high theta/alpha ratio were randomly assigned to the Auditory or the Visual group, where a 500 Hz tone or a visual stimulus (a white square), respectively, was used as a positive reinforcer when the value of the theta/alpha ratio was reduced. Both groups had signs consistent with EEG maturation, but only the Auditory Group showed behavioral/cognitive improvements. In conclusion, the auditory reinforcer was more efficacious in reducing the theta/alpha ratio, and it improved the cognitive abilities more than the visual reinforcer.

Displacement chromatography on cyclodextrin silicas. IV. Separation of the enantiomers of ibuprofen.

PubMed

Farkas, G; Irgens, L H; Quintero, G; Beeson, M D; al-Saeed, A; Vigh, G

1993-08-13

A displacement chromatographic method has been developed for the preparative separation of the enantiomers of ibuprofen using a beta-cyclodextrin silica stationary phase. The retention behavior of ibuprofen was studied in detail: the log k' vs. polar organic modifier concentration, the log k' vs. pH, the log k' vs. buffer concentration and the log k' vs. 1/T relationships; also, the alpha vs. polar organic modifier concentration, the alpha vs. pH, the alpha vs. buffer concentration and the log alpha vs. 1/T relationships have been determined in order to find the carrier solution composition which results in maximum chiral selectivity and sufficient, but not excessive solute retention (1 < k' < 30). 4-tert.-Butylcyclohexanol, a structurally similar but more retained compound than ibuprofen, was selected as displacer for the separation. Even with an alpha value as small as 1.08, good preparative chiral separations were observed both in the displacement mode and in the overloaded elution mode, up to a sample load of 0.5 mg.

Properties of polyproline II, a secondary structure element implicated in protein-protein interactions.

PubMed

Cubellis, M V; Caillez, F; Blundell, T L; Lovell, S C

2005-03-01

The polyproline II (PPII) conformation of protein backbone is an important secondary structure type. It is unusual in that, due to steric constraints, its main-chain hydrogen-bond donors and acceptors cannot easily be satisfied. It is unable to make local hydrogen bonds, in a manner similar to that of alpha-helices, and it cannot easily satisfy the hydrogen-bonding potential of neighboring residues in polyproline conformation in a manner analogous to beta-strands. Here we describe an analysis of polyproline conformations using the HOMSTRAD database of structurally aligned proteins. This allows us not only to determine amino acid propensities from a much larger database than previously but also to investigate conservation of amino acids in polyproline conformations, and the conservation of the conformation itself. Although proline is common in polyproline helices, helices without proline represent 46% of the total. No other amino acid appears to be greatly preferred; glycine and aromatic amino acids have low propensities for PPII. Accordingly, the hydrogen-bonding potential of PPII main-chain is mainly satisfied by water molecules and by other parts of the main-chain. Side-chain to main-chain interactions are mostly nonlocal. Interestingly, the increased number of nonsatisfied H-bond donors and acceptors (as compared with alpha-helices and beta-strands) makes PPII conformers well suited to take part in protein-protein interactions. Copyright 2005 Wiley-Liss, Inc.

Ornithorhynchus anatinus (platypus) links the evolution of immunoglobulin genes in eutherian mammals and nonmammalian tetrapods.

PubMed

Zhao, Yaofeng; Cui, Huiting; Whittington, Camilla M; Wei, Zhiguo; Zhang, Xiaofeng; Zhang, Ziding; Yu, Li; Ren, Liming; Hu, Xiaoxiang; Zhang, Yaping; Hellman, Lars; Belov, Katherine; Li, Ning; Hammarström, Lennart

2009-09-01

The evolutionary origins of mammalian immunoglobulin H chain isotypes (IgM, IgD, IgG, IgE, and IgA) are still incompletely understood as these isotypes differ considerably in structure and number from their counterparts in nonmammalian tetrapods. We report in this study that the platypus (Ornithorhynchus anatinus) Ig H chain constant region gene locus contains eight Ig encoding genes, which are arranged in an mu-delta-omicron-gamma2-gamma1-alpha1-epsilon-alpha2 order, spanning a total of approximately 200 kb DNA, encoding six distinct isotypes. The omicron (omicron for Ornithorhynchus) gene encodes a novel Ig H chain isotype that consists of four constant region domains and a hinge, and is structurally different from any of the five known mammalian Ig classes. This gene is phylogenetically related to upsilon (epsilon) and gamma, and thus appears to be a structural intermediate between these two genes. The platypus delta gene encodes ten heavy chain constant region domains, lacks a hinge region and is similar to IgD in amphibians and fish, but strikingly different from that in eutherian mammals. The platypus Ig H chain isotype repertoire thus shows a unique combination of genes that share similarity both to those of nonmammalian tetrapods and eutherian animals and demonstrates how phylogenetically informative species can be used to reconstruct the evolutionary history of functionally important genes.

X-ray Crystallographic Analysis of [alpha]-Ketoheterocycle Inhibitors Bound to a Humanized Variant of Fatty Acid Amide Hydrolase

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

Mileni, Mauro; Garfunkle, Joie; Ezzili, Cyrine

2010-11-03

Three cocrystal X-ray structures of the {alpha}-ketoheterocycle inhibitors 3-5 bound to a humanized variant of fatty acid amide hydrolase (FAAH) are disclosed and comparatively discussed alongside those of 1 (OL-135) and its isomer 2. These five X-ray structures systematically probe each of the three active site regions key to substrate or inhibitor binding: (1) the conformationally mobile acyl chain-binding pocket and membrane access channel responsible for fatty acid amide substrate and inhibitor acyl chain binding, (2) the atypical active site catalytic residues and surrounding oxyanion hole that covalently binds the core of the {alpha}-ketoheterocycle inhibitors captured as deprotonated hemiketals mimickingmore » the tetrahedral intermediate of the enzyme-catalyzed reaction, and (3) the cytosolic port and its uniquely important imbedded ordered water molecules and a newly identified anion binding site. The detailed analysis of their key active site interactions and their implications on the interpretation of the available structure-activity relationships are discussed providing important insights for future design.« less

H-alpha Fabry-Perot interferometric observations of blue compact dwarf galaxies

NASA Technical Reports Server (NTRS)

Thuan, Trinh Xuan; Williams, T. B.; Malumuth, E.

1987-01-01

H-alpha Fabry-Perot interferometric observations of the two blue compact dwarf galaxies (BCDs) 7 Zw 403 and 1 Zw 49 are presented. The velocity field of 7 Zw 403 shows no clear large-scale organized motion but the velocity field is not completely chaotic either. The gas associated with the 8 H II regions in 7 Zw 403 has neither the highest nor lowest velocities. The BCD 1 Zw 49 is dominated by a single H II region which is about 50 times brighter than any other feature in the galaxy. There is a chain of fainter H II regions extending across the galaxy. The velocity field is well ordered along the H II region chain, but it is very complex around the dominant H II region, suggesting H-alpha loops and filaments around the latter. Both BCDs show velocity gradients of about 25 km/s on scales of about 10 pc in 7 Zw 403 and of about 50 pc in 1 Zw 49. These velocity discontinuities compress the gas and are probably responsible for the star formation.

Fcγ receptor I alpha chain (CD64) expression in macrophages is critical for the onset of meningitis by Escherichia coli K1.

PubMed

Mittal, Rahul; Sukumaran, Sunil K; Selvaraj, Suresh K; Wooster, David G; Babu, M Madan; Schreiber, Alan D; Verbeek, J Sjef; Prasadarao, Nemani V

2010-11-18

Neonatal meningitis due to Escherichia coli K1 is a serious illness with unchanged morbidity and mortality rates for the last few decades. The lack of a comprehensive understanding of the mechanisms involved in the development of meningitis contributes to this poor outcome. Here, we demonstrate that depletion of macrophages in newborn mice renders the animals resistant to E. coli K1 induced meningitis. The entry of E. coli K1 into macrophages requires the interaction of outer membrane protein A (OmpA) of E. coli K1 with the alpha chain of Fcγ receptor I (FcγRIa, CD64) for which IgG opsonization is not necessary. Overexpression of full-length but not C-terminal truncated FcγRIa in COS-1 cells permits E. coli K1 to enter the cells. Moreover, OmpA binding to FcγRIa prevents the recruitment of the γ-chain and induces a different pattern of tyrosine phosphorylation of macrophage proteins compared to IgG2a induced phosphorylation. Of note, FcγRIa(-/-) mice are resistant to E. coli infection due to accelerated clearance of bacteria from circulation, which in turn was the result of increased expression of CR3 on macrophages. Reintroduction of human FcγRIa in mouse FcγRIa(-/-) macrophages in vitro increased bacterial survival by suppressing the expression of CR3. Adoptive transfer of wild type macrophages into FcγRIa(-/-) mice restored susceptibility to E. coli infection. Together, these results show that the interaction of FcγRI alpha chain with OmpA plays a key role in the development of neonatal meningitis by E. coli K1.

An automated method for modeling proteins on known templates using distance geometry.

PubMed

Srinivasan, S; March, C J; Sudarsanam, S

1993-02-01

We present an automated method incorporated into a software package, FOLDER, to fold a protein sequence on a given three-dimensional (3D) template. Starting with the sequence alignment of a family of homologous proteins, tertiary structures are modeled using the known 3D structure of one member of the family as a template. Homologous interatomic distances from the template are used as constraints. For nonhomologous regions in the model protein, the lower and the upper bounds for the interatomic distances are imposed by steric constraints and the globular dimensions of the template, respectively. Distance geometry is used to embed an ensemble of structures consistent with these distance bounds. Structures are selected from this ensemble based on minimal distance error criteria, after a penalty function optimization step. These structures are then refined using energy optimization methods. The method is tested by simulating the alpha-chain of horse hemoglobin using the alpha-chain of human hemoglobin as the template and by comparing the generated models with the crystal structure of the alpha-chain of horse hemoglobin. We also test the packing efficiency of this method by reconstructing the atomic positions of the interior side chains beyond C beta atoms of a protein domain from a known 3D structure. In both test cases, models retain the template constraints and any additionally imposed constraints while the packing of the interior residues is optimized with no short contacts or bond deformations. To demonstrate the use of this method in simulating structures of proteins with nonhomologous disulfides, we construct a model of murine interleukin (IL)-4 using the NMR structure of human IL-4 as the template. The resulting geometry of the nonhomologous disulfide in the model structure for murine IL-4 is consistent with standard disulfide geometry.

Disruption of the processing alpha-mannosidase gene does not prevent outer chain synthesis in Saccharomyces cerevisiae.

PubMed

Puccia, R; Grondin, B; Herscovics, A

1993-02-15

Processing of N-linked oligosaccharides in Saccharomyces cerevisiae begins with the removal of glucose and mannose residues from Glc3Man9GlcNAc2 to form a single isomer of Man8GlcNAc2. The importance of mannose removal for subsequent outer chain synthesis was examined in strains of S. cerevisiae disrupted in the MNS1 gene encoding a specific alpha 1,2-mannosidase responsible for Man8GlcNAc2 synthesis [Camirand, Heysen, Grondin and Herscovics (1991) J. Biol. Chem. 266, 15120-15127]. Both MNS1 transcripts of 1.85 kb and 1.7 kb were not observed in Northern blots of mns1 cells (i.e. cells containing the disrupted gene). Analysis on Bio-Gel P-6 of endo-beta-N-acetylglucosaminidase-H-sensitive oligosaccharides following a 10 min pulse with [2-3H]mannose revealed similar amounts of labelled outer chains excluded from the gel in both control and mns1 cells. H.p.l.c. of the included oligosaccharides showed that a Man9GlcNAc, rather than a Man8GlcNAc, intermediate was formed in mns1 cells. Analysis of [3H]mannose-labelled core oligosaccharides from immunoprecipitated CPY and invertase by h.p.l.c. showed a similar size distribution in mns1 and control cells. Invertase immunoprecipitated from [35S]methionine-labelled mns1 cells was highly glycosylated, but migrated slightly faster than that from control cells on denaturing PAGE, indicating a small difference in glycosylation. A similar difference in mobility was observed for invertase activity stain following non-denaturing gel electrophoresis. It is concluded that the alpha-mannosidase encoded by MNS1 is the only enzyme responsible for mannose removal in vivo, and that this processing step is not essential for outer chain synthesis.

GammaM23K, gammaM232K, and gammaL77K single substitutions in the TF1-ATPase lower ATPase activity by disrupting a cluster of hydrophobic side chains.

PubMed

Bandyopadhyay, Sanjay; Allison, William S

2004-07-27

In crystal structures of the bovine F(1)-ATPase (MF(1)), the side chains of gammaMet(23), gammaMet(232), and gammaLeu(77) interact in a cluster. Substitution of the corresponding residues in the alpha(3)beta(3)gamma subcomplex of TF(1) with lysine lowers the ATPase activity to 2.3, 11, and 15%, respectively, of that displayed by wild-type. In contrast, TF(1) subcomplexes containing the gammaM(23)C, gammaM(232)C, and gammaL(77)C substitutions display 36, 36, and 130%, respectively, of the wild-type ATPase activity. The ATPase activity of the gammaM(23)C/gammaM(232)C double mutant subcomplex is 36% that of the wild-type subcomplex before and after cross-linking the introduced cysteines, whereas the ATPase activity of the gammaM(23)C/L(77)C double mutant increased from 50 to 85% that of wild-type after cross-linking the introduced cysteines. Only beta-beta cross-links formed when the alpha(3)(betaE(395)C)(3)gammaM(23)C double mutant was inactivated with CuCl(2). The overall results suggest that the attenuated ATPase of the mutant subcomplexes containing the gammaM(23)K, gammaL(77)K, and gammaM(232)K substitutions is caused by disruption of the cluster of hydrophobic amino acid side chains and that the midregion of the coiled-coil comprised of the amino- and carboxyl-terminal alpha helices of the gamma subunit does not undergo unwinding or major displacement from the side chain of gammaLeu(77) during ATP-driven rotation of the gamma subunit.

Synthesis and characterization of poly-L-leucine initialized and immobilized by rehydrated hydrotalcite: understanding stability and the nature of interaction.

PubMed

Miranda, Ronald-Alexander; Finocchio, Elisabetta; Llorca, Jordi; Medina, Francisco; Ramis, Gianguido; Sueiras, Jesús E; Segarra, Anna M

2013-10-07

PLLs were synthesized by the ring-opening polycondensation (ROP) method using α-L-leucine N-carboxyanhydride (NCA) and initialized by triethylamine (Et3N), water or rehydrated hydrotalcite (HTrus). The role of temperature, different initiators and water in ROP was further investigated. In general, the initiators used in the polymerization reaction lead to PLL alpha-helical chains containing 5-40 monomers with NCA endgroups via a monomer-activated mechanism. However, the water has a twofold effect on ROP, as both a nucleophile and a base, which involves competition between two different types of initiating mechanisms (nucleophilic attack or deprotonation of the NCA monomer) in the polymerization reaction. This competition provides as a main product NCA endgroups with an alpha-helical structure and leads to the formation of the PLL cyclic-chains and beta-sheet structures which reduce the polymer Mw and the PD of the polypeptide. Furthermore, the water can hydrolyze the NCA endgroups resulting in PLL alpha-helical chains that contain living groups as the main product. On the other hand, the HTrus presents a double role: as both an initiator and a support. The polymers synthesized in the presence of HTrus presented a HT-carboxylate endgroup. The PLLs immobilized in HTrus through an anion-exchange method performed for just 30 minutes presented the PLL immobilized in the interlayer space of the HTrus. The PLL chains of the immobilized counterpart are stabilized by H-bonding with the M-OH of the HT structure. All the polypeptides and biohybrid materials synthesized have been characterized using different techniques (EA, ICP, XRD, Raman, MALDI-TOF, ESI-TOF, FT-IR at increasing temperatures, TG/DT analyses and TEM).

Analysis of transcriptional isoforms of collagen types IX, II, and I in the developing avian cornea by competitive polymerase chain reaction.

PubMed

Fitch, J M; Gordon, M K; Gibney, E P; Linsenmayer, T F

1995-01-01

The genes for the alpha 1(IX), alpha 1(II), and alpha 2(I) collagen chains can give rise to different isoforms of mRNA, generated by alternative promotor usage [for alpha 1(IX) and alpha 2(I)] or alternative splicing [for alpha 1(II)]. In this study, we employed competitive reverse transcriptase PCR to quantitate the amounts of transcriptional isoforms for these genes in the embryonic avian cornea from its inception (about 3 1/2 days of development) to 11 days. In order to compare values at different time points, the results were normalized to those obtained for the "housekeeping" enzyme, glycerol-3-phosphate dehydrogenase (G3PDH). These values were compared to those obtained from other tissues (anterior optic cup and cartilage) that synthesize different combinations of the collagen isoforms. We found that, in the cornea, transcripts from the upstream promotor of alpha 1(IX) collagen (termed "long IX") were predominant at stage 18-20 (about 3 1/2 days), but then fell rapidly, and remained at a low level. By 5 days (just before stromal swelling) the major mRNA isoform of alpha 1(IX) was from the downstream promoter (termed "short IX"). The relative amount of transcript for the short form of type IX collagen rose to a peak at about 6 days of development, and then declined. Throughout this period, the predominant transcriptional isoform of the collagen type II gene was IIA (i.e., containing the alternatively spliced exon 2). This indicates that the molecules of type II collagen that are assembled into heterotypic fibrils with type I collagen possess, at least transiently, an amino-terminal globular domain similar to that found in collagen types I, III, and V. For type I, the "bone/tendon" mRNA isoform of the alpha 2(I) collagen gene was predominant; transcripts from the downstream promotor were at basal levels. In other tissues expressing collagen types IX and II, long IX was expressed predominantly with the IIA form in the anterior optic cup at stage 22/23; in 14 1/2 day cartilage, long IX was expressed predominantly along with the IIB form of alpha 1(II). The downstream transcript of the alpha 2(I) gene (Icart) was found at high levels only in cartilage.

Mitogen-activated protein kinase inhibitors suppress prostaglandin F(2alpha)-induced myosin-light chain phosphorylation and contraction in iris sphincter smooth muscle.

PubMed

Yousufzai, S Y; Gao, G; Abdel-Latif, A A

2000-10-27

The purpose of this study was to investigate the potential role of mitogen-activated protein (MAP) kinase in contraction by monitoring MAP kinase phosphorylation (activation) and contraction during agonist stimulation of cat iris sphincter smooth muscle. Changes in tension in response to prostaglandin F(2alpha), latanoprost, a prostaglandin F(2alpha) analog used as an anti-glaucoma drug, and carbachol were recorded isometrically, and MAP kinase activation was monitored by Western blot using a phosphospecific p42/p44 MAP kinase antibody. We found that treatment of the muscle with 2'-Amino-3'-methoxyflavone (PD98059) (10 microM), a specific inhibitor of MAP kinase kinase (MEK), inhibited significantly prostaglandin F(2alpha)- and latanoprost-induced phosphorylation and contraction, but had little effect on those evoked by carbachol. Prostaglandin F(2alpha) increased MAP kinase phosphorylation in a concentration-dependent manner with EC(50) value of 1.1 x 10(-8) M and increased contraction with EC(50) of 0.92 x 10(-9) M. The MAP kinase inhibitors PD98059, Apigenin and 1,4-Diamino-2,3-dicyano-1, 4bis(2-aminophenylthio)butadiene (UO126) inhibited prostaglandin F(2alpha)-induced contraction in a concentration-dependent manner with IC(50) values of 2.4, 3.0 and 4.8 microM, respectively. PD98059 had no effect on prostaglandin F(2alpha)- or on carbachol-stimulated inositol-1,4,5-trisphosphate (IP(3)) production. In contrast, the MAP kinase inhibitor inhibited prostaglandin F(2alpha)-induced myosin-light chain (MLC) phosphorylation, but had no effect on that of carbachol. N-[2-(N-(4-Chloro-cinnamyl)-N-methylaminomethyl)phenyl]-N-[2- hydroxyethyl]-4-methoxybenzenesulfonamide (KN-93) (10 microM), a Ca(2+)-calmodulin-dependent protein kinase inhibitor, and Wortmannin (10 microM), an MLC kinase inhibitor, inhibited significantly (by 80%) prostaglandin F(2alpha)- and carbachol-induced contraction. It can be concluded that in this smooth muscle p42/p44 MAP kinases are involved in the mechanism of prostaglandin F(2alpha)-, but not in that of carbachol, induced contraction. In addition, these data clearly indicate that the stimulation of the iris sphincter with prostaglandin F(2alpha) and carbachol activate two distinct pathways, the MAP kinase pathway and the Ca(2+) mobilization pathway.

Conditional solvation thermodynamics of isoleucine in model peptides and the limitations of the group-transfer model.

PubMed

Tomar, Dheeraj S; Weber, Valéry; Pettitt, B Montgomery; Asthagiri, D

2014-04-17

The hydration thermodynamics of the amino acid X relative to the reference G (glycine) or the hydration thermodynamics of a small-molecule analog of the side chain of X is often used to model the contribution of X to protein stability and solution thermodynamics. We consider the reasons for successes and limitations of this approach by calculating and comparing the conditional excess free energy, enthalpy, and entropy of hydration of the isoleucine side chain in zwitterionic isoleucine, in extended penta-peptides, and in helical deca-peptides. Butane in gauche conformation serves as a small-molecule analog for the isoleucine side chain. Parsing the hydrophobic and hydrophilic contributions to hydration for the side chain shows that both of these aspects of hydration are context-sensitive. Furthermore, analyzing the solute-solvent interaction contribution to the conditional excess enthalpy of the side chain shows that what is nominally considered a property of the side chain includes entirely nonobvious contributions of the background. The context-sensitivity of hydrophobic and hydrophilic hydration and the conflation of background contributions with energetics attributed to the side chain limit the ability of a single scaling factor, such as the fractional solvent exposure of the group in the protein, to map the component energetic contributions of the model-compound data to their value in the protein. But ignoring the origin of cancellations in the underlying components the group-transfer model may appear to provide a reasonable estimate of the free energy for a given error tolerance.

A theoretical case study of type I and type II beta-turns.

PubMed

Czinki, Eszter; Császár, Attila G; Perczel, András

2003-03-03

NMR chemical shielding anisotropy tensors have been computed by employing a medium size basis set and the GIAO-DFT(B3LYP) formalism of electronic structure theory for all of the atoms of type I and type II beta-turn models. The models contain all possible combinations of the amino acid residues Gly, Ala, Val, and Ser, with all possible side-chain orientations where applicable in a dipeptide. The several hundred structures investigated contain either constrained or optimized phi, psi, and chi dihedral angles. A statistical analysis of the resulting large database was performed and multidimensional (2D and 3D) chemical-shift/chemical-shift plots were generated. The (1)H(alpha-13)C(alpha), (13)C(alpha-1)H(alpha-13)C(beta), and (13)C(alpha-1)H(alpha-13)C' 2D and 3D plots have the notable feature that the conformers clearly cluster in distinct regions. This allows straightforward identification of the backbone and side-chain conformations of the residues forming beta-turns. Chemical shift calculations on larger For-(L-Ala)(n)-NH(2) (n=4, 6, 8) models, containing a single type I or type II beta-turn, prove that the simple models employed are adequate. A limited number of chemical shift calculations performed at the highly correlated CCSD(T) level prove the adequacy of the computational method chosen. For all nuclei, statistically averaged theoretical and experimental shifts taken from the BioMagnetic Resonance Bank (BMRB) exhibit good correlation. These results confirm and extend our previous findings that chemical shift information from selected multiple-pulse NMR experiments could be employed directly to extract folding information for polypeptides and proteins.

Regulation of hypoxia-inducible factor-1alpha by nitric oxide through mitochondria-dependent and -independent pathways.

PubMed Central

Mateo, Jesús; García-Lecea, Marta; Cadenas, Susana; Hernández, Carlos; Moncada, Salvador

2003-01-01

Nitric oxide (NO) has been reported both to promote and to inhibit the activity of the transcription factor hypoxia-inducible factor-1 (HIF-1). In order to avoid the pitfalls associated with the use of NO donors, we have developed a human cell line (Tet-iNOS 293) that expresses the inducible NO synthase (iNOS) under the control of a tetracycline-inducible promoter. Using this system to generate finely controlled amounts of NO, we have demonstrated that the stability of the alpha-subunit of HIF-1 is regulated by NO through two separate mechanisms, only one of which is dependent on a functional respiratory chain. HIF-1alpha is unstable in cells maintained at 21% O(2), but is progressively stabilized as the O(2) concentration decreases, resulting in augmented HIF-1 DNA-binding activity. High concentrations of NO (>1 microM) stabilize HIF-1alpha at all O(2) concentrations tested. This effect does not involve the respiratory chain, since it is preserved in cells lacking functional mitochondria (rho(0)-cells) and is not reproduced by other inhibitors of the cytochrome c oxidase. By contrast, lower concentrations of NO (<400 nM) cause a rapid decrease in HIF-1alpha stabilized by exposure of the cells to 3% O(2). This effect of NO is dependent on the inhibition of mitochondrial respiration, since it is mimicked by other inhibitors of mitochondrial respiration, including those not acting at cytochrome c oxidase. We suggest that, although stabilization of HIF-1alpha by high concentrations of NO might have implications in pathophysiological processes, the inhibitory effect of lower NO concentrations is likely to be of physiological relevance. PMID:14531732

Access to enantioenriched alpha-amino esters via rhodium-catalyzed 1,4-addition/enantioselective protonation.

PubMed

Navarre, Laure; Martinez, Rémi; Genet, Jean-Pierre; Darses, Sylvain

2008-05-14

Conjugate addition of potassium trifluoro(organo)borates 2 to dehydroalanine derivatives 1, mediated by a chiral rhodium catalyst and in situ enantioselective protonation, afforded straightforward access to a variety of protected alpha-amino esters 3 with high yields and enantiomeric excesses up to 95%. Among the tested chiral ligands and proton sources, Binap, in combination with guaiacol (2-methoxyphenol), an inexpensive and nontoxic phenol, afforded the highest asymmetric inductions. Organostannanes have also shown to participate in this reaction. By a fine-tuning of the ester moiety, and using Difluorophos as chiral ligand, increased levels of enantioselectivity, generally close to 95%, were achieved. Deuterium labeling experiments revealed, and DFT calculation supported, an unusual mechanism involving a hydride transfer from the amido substituent to the alpha carbon explaining the high levels of enantioselectivity attained in controlling this alpha chiral center.

Dementia with Lewy bodies in an elderly Greek male due to alpha-synuclein gene mutation.

PubMed

Morfis, Litsa; Cordato, Dennis John

2006-11-01

We report the case of an elderly man of Greek background who presented with progressive cognitive decline and motor parkinsonism on a background of a strong family history of Parkinson's disease. Associated symptoms included visual hallucinations, excessive daytime drowsiness, recurrent falls, orthostatic hypotension and urinary incontinence. His major clinical symptoms and signs fulfilled consensus criteria for a clinical diagnosis of dementia with Lewy bodies. An alpha-synuclein gene mutation analysis for the G209A substitution was positive. We conclude that the alpha-synuclein (G209A) gene mutation genotype should be considered in the differential diagnosis of dementia with Lewy bodies, particularly in patients with European ancestry and a family history of Parkinson's disease.

β-Thalassemia.

PubMed

Origa, Raffaella

2017-06-01

β-Thalassemia is caused by reduced (β + ) or absent (β 0 ) synthesis of the β-globin chains of hemoglobin. Three clinical and hematological conditions of increasing severity are recognized: the β-thalassemia carrier state, thalassemia intermedia, and thalassemia major, a severe transfusion-dependent anemia. The severity of disease expression is related mainly to the degree of α-globin chain excess, which precipitates in the red blood cell precursors, causing both mechanic and oxidative damage (ineffective erythropoiesis). Any mechanism that reduces the number of unbound α-globin chains in the red cells may ameliorate the detrimental effects of excess α-globin chains. Factors include the inheritance of mild/silent β-thalassemia mutations, the coinheritance of α-thalassemia alleles, and increased γ-globin chain production. The clinical severity of β-thalassemia syndromes is also influenced by genetic factors unlinked to globin genes as well as environmental conditions and management. Transfusions and oral iron chelation therapy have dramatically improved the quality of life for patients with thalassemia major. Previously a rapidly fatal disease in early childhood, β-thalassemia is now a chronic disease with a greater life expectancy. At present, the only definitive cure is bone marrow transplantation. Therapies undergoing investigation are modulators of erythropoiesis and stem cell gene therapy.Genet Med advance online publication 03 November 2016.

Cr(II) reactivity of taurine/alpha-ketoglutarate dioxygenase.

PubMed

Grzyska, Piotr K; Hausinger, Robert P

2007-11-26

The interaction of CrII with taurine/alpha-ketoglutarate (alphaKG) dioxygenase (TauD) was examined. CrII replaces FeII and binds stoichiometrically with alphaKG to the FeII/alphaKG binding site of the protein, with additional CrII used to generate a chromophore attributed to a CrIII-semiquinone in a small percentage of the sample. Formation of the latter oxygen-sensitive species requires the dihydroxyphenylalanine (DOPA) quinone form of Tyr-73. This preformed side chain is generated by intracellular self-hydroxylation of Tyr-73 to form DOPA, which is subsequently oxidized to the quinone. No chromophore is generated when using NaBH4-treated sample, protein isolated from anaerobically grown cells, inactive TauD variants that are incapable of self-hydroxylation, or the Y73F active mutant of TauD. A CrIII-DOPA semiquinone also was observed in the herbicide hydroxylase SdpA.

Theoretical and experimental {alpha} decay half-lives of the heaviest odd-Z elements and general predictions

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

Zhang, H. F.; Royer, G.

Theoretical {alpha} decay half-lives of the heaviest odd-Z nuclei are calculated using the experimental Q{sub {alpha}} value. The barriers in the quasimolecular shape path are determined within a Generalized Liquid Drop Model (GLDM) and the WKB approximation is used. The results are compared with calculations using the Density-Dependent M3Y (DDM3Y) effective interaction and the Viola-Seaborg-Sobiczewski (VSS) formulas. The calculations provide consistent estimates for the half-lives of the {alpha} decay chains of these superheavy elements. The experimental data stand between the GLDM calculations and VSS ones in the most time. Predictions are provided for the {alpha} decay half-lives of other superheavymore » nuclei within the GLDM and VSS approaches using the recent extrapolated Q{sub {alpha}} of Audi, Wapstra, and Thibault [Nucl. Phys. A729, 337 (2003)], which may be used for future experimental assignment and identification.« less

viking: identification and characterization of a second type IV collagen in Drosophila.

PubMed

Yasothornsrikul, S; Davis, W J; Cramer, G; Kimbrell, D A; Dearolf, C R

1997-10-01

We have taken an enhancer trap approach to identify genes that are expressed in hematopoietic cells and tissues of Drosophila. We conducted a molecular analysis of two P-element insertion strains that have reporter gene expression in embryonic hemocytes, strain 197 and vikingICO. This analysis has determined that viking encodes a collagen type IV gene, alpha2(IV). The viking locus is located adjacent to the previously described DCg1, which encodes collagen alpha1(IV), and in the opposite orientation. The alpha2(IV) and alpha1(IV) collagens are structurally very similar to one another, and to vertebrate type IV collagens. In early development, viking and DCg1 are transcribed in the same tissue-specific pattern, primarily in the hemocytes and fat body cells. Our results suggest that both the alpha1 and alpha2 collagen IV chains may contribute to basement membranes in Drosophila. This work also provides the foundation for a more complete genetic dissection of collagen type IV molecules and their developmental function in Drosophila.

Molecular chaperone properties of the high molecular weight aggregate from aged lens

NASA Technical Reports Server (NTRS)

Takemoto, L.; Boyle, D.; Spooner, B. S. (Principal Investigator)

1994-01-01

The high molecular weight aggregate (HMWA) fraction was isolated from the water soluble proteins of aged bovine lenses. Its composition and ability to inhibit heat-induced denaturation and aggregation were compared with the lower molecular weight, oligomeric fraction of alpha isolated from the same lens. Although the major components of both fractions were the alpha-A and alpha-B chains, the HMWA fraction possessed a decreased ability to protect other proteins against heat-induced denaturation and aggregation. Immunoelectron microscopy of both fractions demonstrated that alpha particles from the HMWA fraction contained increased amounts of beta and gamma crystallins, bound to a central region of the supramolecular complex. Together, these results demonstrate that alpha crystallins found in the HMWA fraction possess a decreased ability to protect against heat-induced denaturation and aggregation, and suggest that at least part of this decrease could be due to the increased presence of beta and gamma crystallins complexed to the putative chaperone receptor site of the alpha particles.

Herschel Discovery of a New class of Cold, Faint Debris Discs

NASA Technical Reports Server (NTRS)

Eiroa, C.; Marshall, J. P.; Mora, A.; Krivov, A. V.; Montesinos, B.; Absil, O.; Ardila, D.; Arevalo, M.; Augereau, J. -Ch.; Bayo, A.;

Significance of two distinct types of tryptophan synthase beta chain in Bacteria, Archaea and higher plants.

PubMed

Xie, Gary; Forst, Christian; Bonner, Carol; Jensen, Roy A

2002-01-01

Tryptophan synthase consists of two subunits, alpha and beta. Two distinct subgroups of beta chain exist. The major group (TrpEb_1) includes the well-studied beta chain of Salmonella typhimurium. The minor group of beta chain (TrpEb_2) is most frequently found in the Archaea. Most of the amino-acid residues important for catalysis are highly conserved between both TrpE subfamilies. Conserved amino-acid residues of TrpEb_1 that make allosteric contact with the TrpEa subunit (the alpha chain) are absent in TrpEb_2. Representatives of Archaea, Bacteria and higher plants all exist that possess both TrpEb_1 and TrpEb_2. In those prokaryotes where two trpEb genes coexist, one is usually trpEb_1 and is adjacent to trpEa, whereas the second is trpEb_2 and is usually unlinked with other tryptophan-pathway genes. TrpEb_1 is nearly always partnered with TrpEa in the tryptophan synthase reaction. However, by default at least six lineages of the Archaea are likely to use TrpEb_2 as the functional beta chain, as TrpEb_1 is absent. The six lineages show a distinctive divergence within the overall TrpEa phylogenetic tree, consistent with the lack of selection for amino-acid residues in TrpEa that are otherwise conserved for interfacing with TrpEb_1. We suggest that the standalone function of TrpEb_2 might be to catalyze the serine deaminase reaction, an established catalytic capability of tryptophan synthase beta chains. A coincident finding of interest is that the Archaea seem to use the citramalate pathway, rather than threonine deaminase (IlvA), to initiate the pathway of isoleucine biosynthesis.

Relationship between peroxisome proliferator-activated receptor alpha activity and cellular concentration of 14 perfluoroalkyl substances in HepG2 cells.

PubMed

Rosenmai, Anna Kjerstine; Ahrens, Lutz; le Godec, Théo; Lundqvist, Johan; Oskarsson, Agneta

2018-02-01

Peroxisome proliferator-activated receptor alpha (PPARα) is a molecular target for perfluoroalkyl substances (PFASs). Little is known about the cellular uptake of PFASs and how it affects the PPARα activity. We investigated the relationship between PPARα activity and cellular concentration in HepG2 cells of 14 PFASs, including perfluoroalkyl carboxylates (PFCAs), perfluoroalkyl sulfonates and perfluorooctane sulfonamide (FOSA). Cellular concentrations were determined by high-performance liquid chromatography-tandem mass spectrometry and PPARα activity was determined in transiently transfected cells by reporter gene assay. Cellular uptake of the PFASs was low (0.04-4.1%) with absolute cellular concentrations in the range 4-2500 ng mg -1 protein. Cellular concentration of PFCAs increased with perfluorocarbon chain length up to perfluorododecanoate. PPARα activity of PFCAs increased with chain length up to perfluorooctanoate. The maximum induction of PPARα activity was similar for short-chain (perfluorobutanoate and perfluoropentanoate) and long-chain PFCAs (perfluorododecanoate and perfluorotetradecanoate) (approximately twofold). However, PPARα activities were induced at lower cellular concentrations for the short-chain homologs compared to the long-chain homologs. Perfluorohexanoate, perfluoroheptanoate, perfluorooctanoate, perfluorononanoate (PFNA) and perfluorodecanoate induced PPARα activities >2.5-fold compared to controls. The concentration-response relationships were positive for all the tested compounds, except perfluorooctane sulfonate PFOS and FOSA, and were compound-specific, as demonstrated by differences in the estimated slopes. The relationships were steeper for PFCAs with chain lengths up to and including PFNA than for the other studied PFASs. To our knowledge, this is the first report establishing relationships between PPARα activity and cellular concentration of a broad range of PFASs. Copyright © 2017 John Wiley & Sons, Ltd.

Regulation of hepatic branched-chain alpha-keto acid dehydrogenase kinase in a rat model for type 2 diabetes mellitus at different stages of the disease.

PubMed

Doisaki, Masao; Katano, Yoshiaki; Nakano, Isao; Hirooka, Yoshiki; Itoh, Akihiro; Ishigami, Masatoshi; Hayashi, Kazuhiko; Goto, Hidemi; Fujita, Yuko; Kadota, Yoshihiro; Kitaura, Yasuyuki; Bajotto, Gustavo; Kazama, Shunsuke; Tamura, Tomohiro; Tamura, Noriko; Feng, Guo-Gang; Ishikawa, Naohisa; Shimomura, Yoshiharu

2010-03-05

Branched-chain alpha-keto acid dehydrogenase (BCKDH) kinase (BDK) is responsible for the regulation of BCKDH complex, which is the rate-limiting enzyme in the catabolism of branched-chain amino acids (BCAAs). In the present study, we investigated the expression and activity of hepatic BDK in spontaneous type 2 diabetes using hyperinsulinemic Zucker diabetic fatty rats aged 9weeks and hyperglycemic, but not hyperinsulinemic rats aged 18weeks. The abundance of hepatic BDK mRNA and total BDK protein did not correlate with changes in serum insulin concentrations. On the other hand, the amount of BDK bound to the complex and its kinase activity were correlated with alterations in serum insulin levels, suggesting that hyperinsulinemia upregulates hepatic BDK. The activity of BDK inversely corresponded with the BCKDH complex activity, which was suppressed in hyperinsulinemic rats. These results suggest that insulin regulates BCAA catabolism in type 2 diabetic rats by modulating the hepatic BDK activity. 2010 Elsevier Inc. All rights reserved.

The conformational requirements for the mechanical precipitation of hemoglobin S and other mutants.

PubMed

Roth, E F; Elbaum, D; Bookchin, R M; Nagel, R L

1976-08-01

The mechanical stability of human hemoglobin mutants was studied for the specific effects of single and double amino acid substitutions, the ligand state of each chain, and the effect of hybrids between oxy and cyanmet partners on precipitability. It was found that the beta6Glu leads to Val and the beta73 Asp leads to Asn mutations increased the degree of mechanical precipitation in the liganded but not in the deoxy form. When these mutations occurred on the same chain, the effects were approximately additive. Heat labile mutants such as Hb Gun Hill and Hb Leiden exhibited mechanical instability, but probably through a different mechanism, as very little dependence on ligand state was apparent. Studies with valency hybrids of HbS(alpha2 betas2-and-alpha2 betas2 where = cyanmet) revealed that instability was primarily determined by the state of the betas chain, which must be liganded to confer instability on the tetramer. A good agreement between surface activity and mechanical precipitability of these mutants has been found.

alpha-decay half-lives and Q{sub a}lpha values of superheavy nuclei

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

Dong Jianmin; Graduate University of Chinese Academy of Sciences, Beijing 100049; School of Nuclear Science and Technology, Lanzhou University, Lanzhou 730000

2010-06-15

The alpha-decay half-lives of recently synthesized superheavy nuclei (SHN) are investigated by employing a unified fission model (UFM) where a new method to calculate the assault frequency of alpha emission is used. The excellent agreement with the experimental data indicates the UFM is a useful tool to investigate these alpha decays. It is found that the alpha-decay half-lives become more and more insensitive to the Q{sub a}lpha values as the atomic number increases on the whole, which is favorable for us to predict the half-lives of SHN. In addition, a formula is proposed to compute the Q{sub a}lpha values formore » the nuclei with Z>=92 and N>=140 with a good accuracy, according to which the long-lived SHN should be neutron rich. Several weeks ago, two isotopes of a new element with atomic number Z=117 were synthesized and their alpha-decay chains have been observed. The Q{sub a}lpha formula is found to work well for these nuclei, confirming its predictive power. The experimental half-lives are well reproduced by employing the UFM with the experimental Q{sub a}lpha values. This fact that the experimental half-lives are compatible with experimental Q{sub a}lpha values supports the synthesis of a new element 117 and the experimental measurements to a certain extent.« less

Adipocytokine concentrations in children with different types of beta-thalassemia.

PubMed

Enli, Yaşar; Balci, Yasemin I; Gönen, Cafer; Uzun, Ebru; Polat, Aziz

2014-06-01

Beta-thalassemia is an inherited blood disorder. It results from the impaired production of β-globin chains, leading to a relative excess of alpha-globin chains. Clinical severity separates this disease into three main subtypes: β- thalassemia major, β-thalassemia intermedia and β-thalassemia minor, the former two being clinically more significant. Inflammatory processes may play an important role in some of the complications of thalassemia. Adipose tissue is one of the most important endocrine and secretory organs that release adipocytokines like adiponectin, resistin and visfatin. The aim of our study was to analyze adipocytokine concentrations (adiponectin, resistin and visfatin) in different types of β-thalassemia patients and determine any possible correlations with disease severity. We recruited 29 patients who were transfusion-dependent β-thalassemia-major patients, 17 patients with β-thalassemia intermedia, 30 β-thalassemia minor patients. The control group consisted of 30 healthy children. Anthropometric measurements, complete blood count, biochemical parameters, serum concentrations of adiponectin, resistin, visfatin were performed for all subjects. Resistin and visfatin concentrations were significantly higher in β-thalassemia minor patients than in controls. Adiponetin, resistin and visfatin concentrations were significantly higher in both β-thalassemia intermedia and major patients than in controls. The concentrations of adiponectin, resistin and visfatin were significantly higher in both β-thalassemia intermedia and major patients than in β-thalassemia minor patients. There was no significant difference between β-thalassemia intermedia and β-thalassemia major patients for adipocytokines concentrations. We speculate that these adipocytokines may play a role in the development of complications in β-thalassaemia.

Neurocognitive effects of an omega-3 fatty acid and vitamins E+C in schizophrenia: A randomised controlled trial.

PubMed

Bentsen, H; Landrø, N I

2017-10-16

There is need for more efficient treatment of neurocognitive deficits in schizophrenia. In this 16 weeks randomised, placebo-controlled trial, we examined neurocognitive effects of adding ethyl-eicosapentaenoate 2g/day and/or vitamins E 364mg/day + C 1000mg/day to antipsychotics in 53 patients aged 18-39 years with acute schizophrenia. For the sake of validating neurocognitive tests, healthy subjects, not taking trial drugs, were also included in the study. Ethyl-EPA given alone to patients with low baseline RBC polyunsaturated fatty acids (PUFA), and Vitamins E+C given alone to high PUFA patients, impaired sustained attention (Continuous Performance Test, CPT-IP d prime score), standardised effect sizes d = 0.78 and d = 0.69, respectively. These adverse effects were paralleled by excessive increases in long-chain PUFA and serum alpha-tocopherol, respectively. They were counteracted by combining ethyl-EPA and vitamins, d = 0.80 and d = 0.74 in low and high PUFA patients, respectively. No other neurocognitive tests yielded significant results. Plausible mechanisms of harmful effects are oxidative stress and lipid raft disruption. Copyright © 2017. Published by Elsevier Ltd.

Alpha-Glucan, Water Dikinase 1 Affects Starch Metabolism and Storage Root Growth in Cassava (Manihot esculenta Crantz).

PubMed

Zhou, Wenzhi; He, Shutao; Naconsie, Maliwan; Ma, Qiuxiang; Zeeman, Samuel C; Gruissem, Wilhelm; Zhang, Peng

2017-08-29

Regulation of storage root development by source strength remains largely unknown. The cassava storage root delay (srd) T-DNA mutant postpones storage root development but manifests normal foliage growth as wild-type plants. The SRD gene was identified as an orthologue of α-glucan, water dikinase 1 (GWD1), whose expression is regulated under conditions of light/dark cycles in leaves and is associated with storage root development. The GWD1-RNAi cassava plants showed both retarded plant and storage root growth, as a result of starch excess phenotypes with reduced photosynthetic capacity and decreased levels of soluble saccharides in their leaves. These leaves contained starch granules having greatly increased amylose content and type C semi-crystalline structures with increased short chains that suggested storage starch. In storage roots of GWD1-RNAi lines, maltose content was dramatically decreased and starches with much lower phosphorylation levels showed a drastically reduced β-amylolytic rate. These results suggested that GWD1 regulates transient starch morphogenesis and storage root growth by decreasing photo-assimilation partitioning from the source to the sink and by starch mobilization in root crops.

The role of autophagy in Parkinson's disease: rotenone-based modeling

PubMed Central

2013-01-01

Background Autophagy-mediated self-digestion of cytoplasmic inclusions may be protective against neurodegenerative diseases such as Parkinson’s disease (PD). However, excessive autophagic activation evokes autophagic programmed cell death. Methods In this study, we aimed at exploring the role of autophagy in the pathogenesis of rotenone-induced cellular and animal models for PD. Results Reactive oxygen species over-generation, mitochondrial membrane potential reduction or apoptosis rate elevation occurred in a dose-dependent fashion in rotenone-treated human neuroblastoma cell line SH-SY5Y. The time- and dose-dependent increases in autophagic marker microtubule-associated protein1 light chain 3 (LC3) expression and decreases in autophagic adaptor protein P62 were observed in this cellular model. LC3-positive autophagic vacuoles were colocalized with alpha-synuclein-overexpressed aggregations. Moreover, the number of autophagic vacuoles was increased in rotenone-based PD models in vitro and in vivo. Conclusions These data, along with our previous finding showing rotenone-induced toxicity was prevented by the autophagy enhancers and was aggravated by the autophagy inhibitors in SH-SY5Y, suggest that autophagy contributes to the pathogenesis of PD, attenuates the rotenone toxicity and possibly represents a new subcellular target for treating PD. PMID:23497442

Behavior of a cloned murine interferon alpha/beta receptor expressed in homospecific or heterospecific background.

PubMed

Uzé, G; Lutfalla, G; Bandu, M T; Proudhon, D; Mogensen, K E

1992-05-15

A murine interferon (IFN) alpha/beta receptor was cloned from the IFN-sensitive L1210 cell line on the basis of its homology with the human receptor. A combination of methods that includes the screening of random-primed and oligo(dT)-primed cDNA libraries and polymerase chain reactions with a single-side specificity was used. At the amino acid level, the murine IFN-alpha/beta shows 46% identity with its human counterpart. Both human WISH cells presenting a low sensitivity to mouse IFN and a murine L1210 mutant subline that does not express the receptor have been stably transfected with the murine IFN-alpha/beta receptor. Whereas transfected human cells became sensitive to a limited number of mouse IFN-alpha/beta subtypes, the transfected murine L1210 mutant was found to be fully complemented and became sensitive to all mouse IFN-alpha/beta subtypes tested, including those that were not active on transfected human cells. These results strongly suggest that the receptor described here is implicated in the mediation of the activities of all murine IFN-alpha/beta subtypes.

Study on molecular epidemiology of the alpha-thalassemias in Liuzhou City, Guangxi Autonomous Region, China.

PubMed

Cai, Ren; Liu, Jingzhong; Wang, Lirong; Liang, Xin; Xiao, Bai; Su, Liu; Zhou, Yan; Pan, Lizhen

2004-01-01

Guangxi is one of the provinces of Southern China with the highest incidence of alpha-thalassemia (thal). Liuzhou is the second biggest city in Guangxi. To find out the incidence of the various alpha-thal genotypes, and their distribution in the Liuzhou area, an F820 Blood Cell Analysis System was used to measure the parameters of red blood cells. A SPIFE Rapid Auto-Electrophoresis System was used to analyze the normal and abnormal hemoglobins (Hbs). Multiplex polymerase chain reaction (mPCR) was used to detect the alpha-globin genotypes. Thirty-two (2.05%) out of 7805 young couples undergoing pre-marriage counseling, were diagnosed as having an Hb H (beta4) disease. The study of 1228 cord blood samples revealed 138 newborn children carrying an alpha-thal determinant with nine different genotypes, thus making the total incidence of alpha-thal 11.24%. Among 185 cases of Hb H, 119 (64.1%) were confirmed as being deletional, and 66 cases (35.7%) nondeletional types. The severity of the Hb H diseases could be classified in the following order: alphaCSalpha/--SEA (alphaConstant (Spring)alpha/--Southeast Asia); alpha(-4.2)/--SEA; alpha(-3.7)/--SEA. Ten cases of alpha-thal determinants were found in combination with beta-thal. The mPCR technique can detect all kinds of combinations of the three common large deletions (--SEA, alpha(-4.2) and alpha(-3.7)) accurately and conveniently.

Management of Excess Material in the Navys Real Time Reutilization Asset Management Facilities Needs Improvement

DTIC Science & Technology

2017-01-23

operations for financial and supply chain management . ii │ DODIG-2017-043 ( Project No. D2016-D000RD-0052.000) Results in Brief Management of Excess...following page. Recommendations (cont’d) DODIG-2017-043 ( Project No. D2016-D000RD-0052.000) │ iii Recommendations Table Management Recommendations...No. DODIG-2017-043 J A N U A R Y 2 3 , 2 0 1 7 Management of Excess Material in the Navy’s Real-Time Reutilization Asset Management Facilities

Aggregation of alpha-synuclein by a coarse-grained Monte Carlo simulation

NASA Astrophysics Data System (ADS)

Farmer, Barry; Pandey, Ras

Alpha-synuclein, an intrinsic protein abundant in neurons, is believed to be a major cause of neurodegenerative diseases (e.g. Alzheimer, Parkinson's disease). Abnormal aggregation of ASN leads to Lewy bodies with specific morphologies. We investigate the self-organizing structures in a crowded environment of ASN proteins by a coarse-grained Monte Carlo simulation. ASN is a chain of 140 residues. Structure detail of residues is neglected but its specificity is captured via unique knowledge-based residue-residue interactions. Large-scale simulations are performed to analyze a number local and global physical quantities (e.g. mobility profile, contact map, radius of gyration, structure factor) as a function of temperature and protein concentration. Trend in multi-scale structural variations of the protein in a crowded environment is compared with that of a free protein chain.

Open conformers: the hidden face of MHC-I molecules.

PubMed

Arosa, Fernando A; Santos, Susana G; Powis, Simon J

2007-03-01

A pool of MHC-I molecules present at the plasma membrane can dissociate from the peptide and/or the light chain, becoming open MHC-I conformers. Whereas peptide-bound MHC-I molecules have an important role in regulating adaptive and innate immune responses, through trans-interactions with T cell and NK cell receptors, the function of the open MHC-I conformers is less clear but seems to be related to their inherent ability to cis-associate, both with themselves and with other receptors. Here, we review data indicating the open MHC-I conformers as regulators of ligand-receptor interactions and discuss the biological implications for immune and non-immune cells. The likelihood that the MHC-I heavy chains have hidden functions that are determined by the amino acid sequence of the alpha1 and alpha2 domains are discussed.

Differences among the cell wall galactomannans from Aspergillus wentii and Chaetosartorya chrysella and that of Aspergillus fumigatus.

PubMed

Gómez-Miranda, Begoña; Prieto, Alicia; Leal, Juan Antonio; Ahrazem, Oussama; Jiménez-Barbero, Jesús; Bernabé, Manuel

2004-01-01

The alkali extractable and water-soluble cell wall polysaccharides F1SS from Aspergillus wentii and Chaetosartorya chrysella have been studied by methylation analysis, 1D- and 2D-NMR, and MALDI-TOF analysis. Their structures are almost identical, corresponding to the following repeating unit: [--> 3)-beta-D-Gal f -(1 --> 5)-beta-D-Gal f-(1 -->]n --> mannan core. The structure of this galactofuranose side chain differs from that found in the pathogenic fungus Aspergillus fumigatus, in other Aspergillii and members of Trichocomaceae: [--> 5)-beta-D-Gal f-(1 -->]n --> mannan core. The mannan cores have also been investigated, and are constituted by a (1 --> 6)-alpha-mannan backbone, substituted at positions 2 by chains from 1 to 7 residues of (1 --> 2) linked alpha-mannopyranoses. Copyright 2004 Kluwer Academic Publishers

The Chlamydomonas Dhc1 gene encodes a dynein heavy chain subunit required for assembly of the I1 inner arm complex.

PubMed Central

Myster, S H; Knott, J A; O'Toole, E; Porter, M E

1997-01-01

Multiple members of the dynein heavy chain (Dhc) gene family have been recovered in several organisms, but the relationships between these sequences and the Dhc isoforms that they encode are largely unknown. To identify Dhc loci and determine the specific functions of the individual Dhc isoforms, we have screened a collection of motility mutants generated by insertional mutagenesis in Chlamydomonas. In this report, we characterize one strain, pf9-3, in which the insertion event was accompanied by a deletion of approximately 13 kb of genomic DNA within the transcription unit of the Dhc1 gene. Northern blot analysis confirms that pf9-3 is a null mutation. Biochemical and structural studies of isolated axonemes demonstrate that the pf9-3 mutant fails to assemble the I1 inner arm complex, a two-headed dynein isoform composed of two Dhcs (1 alpha and 1 beta) and three intermediate chains. To determine if the Dhc1 gene product corresponds to one of the Dhcs of the I1 complex, antibodies were generated against a Dhc1-specific peptide sequence. Immunoblot analysis reveals that the Dhc1 gene encodes the 1 alpha Dhc subunit. These studies thus, identify the first inner arm Dhc locus to be described in any organism and further demonstrate that the 1 alpha Dhc subunit plays an essential role in the assembly of the I1 inner arm complex. Images PMID:9247642

Altered cortisol metabolism in polycystic ovary syndrome: insulin enhances 5alpha-reduction but not the elevated adrenal steroid production rates.

PubMed

Tsilchorozidou, Tasoula; Honour, John W; Conway, Gerard S

2003-12-01

Androgen excess in women with polycystic ovary syndrome (PCOS) may be ovarian and/or adrenal in origin, and one proposed contributing mechanism is altered cortisol metabolism. Increased peripheral metabolism of cortisol may occur by enhanced inactivation of cortisol by 5alpha-reductase (5alpha-R) or impaired reactivation of cortisol from cortisone by 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD1) resulting in decreased negative feedback suppression of ACTH secretion maintaining normal plasma cortisol concentrations at the expense of androgen excess. We have tested whether any enzyme dysregulation was related to circulating insulin or androgen concentrations in women with PCOS and have sought to clarify their relationship with obesity. First, to avoid obesity-related effects on cortisol metabolism, 18 lean women with PCOS were compared with 19 lean controls who were closely matched for body mass index (BMI). Second, the impact of obesity was studied in a cross-section of 42 PCOS women of a broad range of BMI. We measured 24-h urinary excretion of steroid metabolites by gas chromatography/mass spectrometry and fasting metabolic and hormone profiles. Urinary excretion of androgens [androsterone (P = 0.003), etiocholanolone (P = 0.02), and C19 steroid sulfates (P = 0.009)], cortisone metabolites [tetrahydrocortisone (THE) (P = 0.02), alpha-cortolone (P < 0.001), beta-cortol + beta-cortolone (P < 0.001), cortolones (P < 0.001), and E metabolites (P < 0.001)], and TCM (P = 0.002) were raised in lean PCOS subjects when compared with controls. A significantly higher 5alpha-tetrahydrocortisol (5alpha-THF)/5beta-THF ratio (P = 0.04) and a significantly lower alpha-THF + THF + alpha-cortol/THE + cortolones ratio (P = 0.01) were found in lean PCOS women compared with lean controls, indicating both enhanced 5alpha-R and reduced 11beta-HSD1 activities. A decreased THE/cortolones ratio (P = 0.03) was also found in lean PCOS women compared with lean controls, indicating increased 20 alpha/beta-HSD activity. In the group of 42 PCOS subjects, measures of 5alpha/5beta reduction were positively correlated with the homeostasis model insulin resistance index (HOMA-R): alpha-THF/THF and HOMA-R (r = 0.34; P = 0.03), androsterone/etiocholanolone and HOMA-R (r = 0.32; P = 0.04), and total 5alpha /total 5beta and HOMA-R (r = 0.37; P = 0.02). A positive correlation was also found between measures of 5alpha-R and BMI (r = 0.37; P = 0.02). No correlation was found between measures of 11beta-HSD1 activity and indices of insulin sensitivity or BMI. We have demonstrated that there is an increased production rate of cortisol and androgens as measured in vivo in lean PCOS women. Insulin seems to enhance 5alpha reduction of steroids in PCOS but was not associated with the elevated cortisol production rate. The changes in 5alpha-R, 11beta-HSD1, and 20alpha/beta-HSD enzyme activities observed in PCOS may contribute to the increased production rates of cortisol and androgens, supporting the concept of a widespread dysregulation of steroid metabolism. This dysregulation does not seem to be the primary cause of PCOS because no correlation was found between serum androgen levels or urinary excretion of androgens with measurements of either 5alpha-R or 11beta-HSD1 activities.

Dronedarone administration prevents body weight gain and increases tolerance of the heart to ischemic stress: a possible involvement of thyroid hormone receptor alpha1.

PubMed

Pantos, Constantinos; Mourouzis, Iordanis; Malliopoulou, Vassiliki; Paizis, Ioannis; Tzeis, Stylianos; Moraitis, Panagiotis; Sfakianoudis, Konstantinos; Varonos, Dennis D; Cokkinos, Dennis V

2005-01-01

Hypothyroid heart displays a phenotype of cardioprotection against ischemia and this study investigated whether administration of dronedarone, an amiodarone-like compound that has been shown to preferentially antagonize thyroid hormone binding to thyroid hormone receptor alpha1 (TRalpha1), results in a similar effect. Dronedarone was given in Wistar rats (90 mg/kg, once daily (od) for 2 weeks) (DRON), while untreated animals served as controls (CONT). Hypothyroidism (HYPO) was induced by propylthiouracil administration. Isolated rat hearts were perfused in Langendorff mode and subjected to 20 minutes of zero-flow global ischemia (I) followed by 45 minutes of reperfusion (R). 3,5,3' Triiodothyronine remained unchanged while body weight and food intake were reduced. alpha-Myosin heavy chain (alpha-MHC) decreased in DRON while beta-myosin heavy chain (beta-MHC) and sarcoplasmic reticulum Ca2+ adenosine triphosphatase (ATPase) expression (SERCA) was similar to CONT. In HYPO, alpha-MHC and SERCA were decreased while beta-MHC was increased. Myocardial glycogen content was increased in both DRON and HYPO. In DRON, resting heart rate and contractility were reduced and ischemic contracture was significantly suppressed while postischemic left ventricular end-diastolic pressure and lactate dehydrogenase release (IU/L min) after I/R were significantly decreased. In conclusion, dronedarone treatment results in cardioprotection by selectively mimicking hypothyroidism. This is accompanied by a reduction in body weight because of the suppression of food intake. TRs might prove novel pharmacologic targets for the treatment of cardiovascular illnesses.

Variability and repertoire size of T-cell receptor V alpha gene segments.

PubMed

Becker, D M; Pattern, P; Chien, Y; Yokota, T; Eshhar, Z; Giedlin, M; Gascoigne, N R; Goodnow, C; Wolf, R; Arai, K

The immune system of higher organisms is composed largely of two distinct cell types, B lymphocytes and T lymphocytes, each of which is independently capable of recognizing an enormous number of distinct entities through their antigen receptors; surface immunoglobulin in the case of the former, and the T-cell receptor (TCR) in the case of the latter. In both cell types, the genes encoding the antigen receptors consist of multiple gene segments which recombine during maturation to produce many possible peptides. One striking difference between B- and T-cell recognition that has not yet been resolved by the structural data is the fact that T cells generally require a major histocompatibility determinant together with an antigen whereas, in most cases, antibodies recognize antigen alone. Recently, we and others have found that a series of TCR V beta gene sequences show conservation of many of the same residues that are conserved between heavy- and light-chain immunoglobulin V regions, and these V beta sequences are predicted to have an immunoglobulin-like secondary structure. To extend these studies, we have isolated and sequenced eight additional alpha-chain complementary cDNA clones and compared them with published sequences. Analyses of these sequences, reported here, indicate that V alpha regions have many of the characteristics of V beta gene segments but differ in that they almost always occur as cross-hybridizing gene families. We conclude that there may be very different selective pressures operating on V alpha and V beta sequences and that the V alpha repertoire may be considerably larger than that of V beta.

A sulfated alpha-L-fucan from sea cucumber.

PubMed

Ribeiro, A C; Vieira, R P; Mourão, P A; Mulloy, B

1994-03-04

A purified sulfated alpha-L-fucan from the sea cucumber body wall was studied, before and after almost complete desulfation, using methylation analysis and NMR spectroscopy. NMR analysis indicates that 2,4-di-O-sulfo-L-fucopyranose and unsubstituted fucopyranose are present in equal proportions, and that 2-O-sulfo-L-fucopyranose is present in twice that proportion. There is some NMR evidence that a regular repeating sequence of four residues comprises most or all of the polysaccharide chain.

IL-4 function can be transferred to the IL-2 receptor by tyrosine containing sequences found in the IL-4 receptor alpha chain.

PubMed

Wang, H Y; Paul, W E; Keegan, A D

1996-02-01

IL-4 binds to a cell surface receptor complex that consists of the IL-4 binding protein (IL-4R alpha) and the gamma chain of the IL-2 receptor complex (gamma c). The receptors for IL-4 and IL-2 have several features in common; both use the gamma c as a receptor component, and both activate the Janus kinases JAK-1 and JAK-3. In spite of these similarities, IL-4 evokes specific responses, including the tyrosine phosphorylation of 4PS/IRS-2 and the induction of CD23. To determine whether sequences within the cytoplasmic domain of the IL-4R alpha specify these IL-4-specific responses, we transplanted the insulin IL-4 receptor motif (I4R motif) of the huIL-4R alpha to the cytoplasmic domain of a truncated IL-2R beta. In addition, we transplanted a region that contains peptide sequences shown to block Stat6 binding to DNA. We analyzed the ability of cells expressing these IL-2R-IL-4R chimeric constructs to respond to IL-2. We found that IL-4 function could be transplanted to the IL-2 receptor by these regions and that proliferative and differentiative functions can be induced by different receptor sequences.

Grape seed proanthocyanidins ameliorates isoproterenol-induced myocardial injury in rats by stabilizing mitochondrial and lysosomal enzymes: an in vivo study.

PubMed

Karthikeyan, K; Sarala Bai, B R; Niranjali Devaraj, S

2007-11-30

This study was designed to examine the effects of grape seed proanthocyanidins (GSP) against myocardial injury (MI) induced by isoproterenol (ISO), in a rat model. Induction of rats with ISO (85 mg/kg body weight, subcutaneously) for 2 days resulted in a significant decrease in the activities of heart mitochondrial enzymes (isocitrate dehydrogenase, succinate dehydrogenase, malate dehydrogenase and alpha-ketoglutarate dehydrogenase) and respiratory chain enzymes (NADH dehydrogenase and cytochrome c oxidase). The activities of lysosomal enzymes (alpha-d-glucuronidase, alpha-d-N-acetylglucosaminidase, cathepsin-D, acid phosphatases and alpha-d-galactosidase) were increased significantly in the heart and serum of ISO-induced rats. The prior administration of GSP for 6 days a week for 5 weeks significantly increased the activities of mitochondrial and respiratory chain enzymes and significantly decreased the activities of lysosomal enzymes in the heart tissues of ISO-induced rats, which proves the stress stabilizing action of GSP. Oral administration of grape seed proanthocyanidins alone (50, 100 and 150 mg/kg) to normal rats did not show any significant effect in all the parameters studied. These biochemical functional alterations were supported by the macroscopic enzyme mapping assay of ischemic myocardium. Thus, this study shows that 100 and 150 mg/kg of GSP gives protection against ISO-induced MI and demonstrates that GSP has a significant effect in the protection of heart.

Localization of functional receptor epitopes on the structure of ciliary neurotrophic factor indicates a conserved, function-related epitope topography among helical cytokines.

PubMed

Panayotatos, N; Radziejewska, E; Acheson, A; Somogyi, R; Thadani, A; Hendrickson, W A; McDonald, N Q

1995-06-09

By rational mutagenesis, receptor-specific functional analysis, and visualization of complex formation in solution, we identified individual amino acid side chains involved specifically in the interaction of ciliary neurotrophic factor (CNTF) with CNTFR alpha and not with the beta-components, gp130 and LIFR. In the crystal structure, the side chains of these residues, which are located in helix A, the AB loop, helix B, and helix D, are surface accessible and are clustered in space, thus constituting an epitope for CNTFR alpha. By the same analysis, a partial epitope for gp130 was also identified on the surface of helix A that faces away from the alpha-epitope. Superposition of the CNTF and growth hormone structures showed that the location of these epitopes on CNTF is analogous to the location of the first and second receptor epitopes on the surface of growth hormone. Further comparison with proposed binding sites for alpha- and beta-receptors on interleukin-6 and leukemia inhibitory factor indicated that this epitope topology is conserved among helical cytokines. In each case, epitope I is utilized by the specificity-conferring component, whereas epitopes II and III are used by accessory components. Thus, in addition to a common fold, helical cytokines share a conserved order of receptor epitopes that is function related.

Radical-cationic gaseous amino acids: a theoretical study.

PubMed

Sutherland, Kailee N; Mineau, Philippe C; Orlova, Galina

2007-08-16

Three major forms of gaseous radical-cationic amino acids (RCAAs), keto (COOH), enolic (C(OH)OH), and zwitterionic (COO(-)), as well as their tautomers, are examined for aliphatic Ala(.+), Pro(.+), and Ser(.+), sulfur-containing Cys(.+), aromatic Trp(.+), Tyr(.+), and Phe(.+), and basic His(.+). The hybrid B3LYP exchange-correlation functional with various basis sets along with the highly correlated CCSD(T) method is used. For all RCAAs considered, the main stabilizing factor is spin delocalization; for His(.+), protonation of the basic side chain is equally important. Minor stabilizing factors are hydrogen bonding and 3e-2c interactions. An efficient spin delocalization along the N-C(alpha)-C(O-)O moiety occurs upon H-transfer from C(alpha) to the carboxylic group to yield the captodative enolic form, which is the lowest-energy isomer for Ala(.+), Pro(.+), Ser(.+), Cys(.+), Tyr(.+), and Phe(.+). This H-transfer occurs in a single step as a 1,3-shift through the sigma-system. For His(.+), the lowest-energy isomer is formed upon H-transfer from C(alpha) to the basic side chain, which results in a keto form, with spin delocalized along the N-C(alpha)-C=O fragment. Trp(.+) is the only RCAA that favors spin delocalization over an aromatic system given the low ionization energy of indole. The lowest-energy isomer of Trp(.+) is a keto form, with no H-transfer.

Engineering acyclic stereocontrol in the alkylation of vinylglycine-derived dianions: asymmetric synthesis of higher alpha-vinyl amino acids.

PubMed

Berkowitz, D B; McFadden, J M; Sloss, M K

2000-05-19

A generalizable synthesis of higher L-alpha-vinyl amino acids is presented. The strategy pursued here involves the introduction of the amino acid side chain via the alkylation of a chiral, vinylglycine-derived dianionic dienolate, bearing the (-)-8-(beta-naphthyl)menthyl (d'Angelo) auxiliary. A model is presented that postulates a favored "exo-entended" conformation for this dienolate, leading to C(alpha)-alkylation at the si face. The model invokes internal amidate chelation to control ester enolate geometry and soft-soft interactions between the polarizable beta-naphthyl ring of the auxiliary and the extended pi-system of the dienolate to shield the re face. Heats of formation for four conformers of this dianion were calculated for their semiempirical optimized geometries (PM3). The results support the notion that in these vinylglycine-derived dianionic dienolates, "exo" conformations are considerable lower in energy than their "endo" counterparts, with the "exo-entended" conformation being most favorable. In fact, the d'Angelo auxiliary gives a greater degree of acyclic stereocontrol in this system when compared with the (-)-8-phenylmenthyl (Corey) and trans-2-(beta-naphthyl)cyclohexyl auxiliaries, using isobutyl iodide and benzyl bromide as model electrophiles. These dianions are generated from the corresponding dehydrobutyrine esters via sequential deprotonation with LDA and n-BuLi (2 equiv). When alkylations are carried out at -78 degrees C in THF-HMPA, they proceed in 65-81% yields, with both regiocontrol (deconjugative alpha-alkylation is preferred over gamma-alkylation) and a great degree of acyclic stereocontrol [91:9 to >/=98:2 diastereomeric ratios (10 examples)]. The auxiliary may be recovered in high yield (generally 90%) using a modification of Gassman's "anhydrous hydroxide" conditions, in which considerably higher temperatures are employed. Among the side chains introduced directly are those of butyrine, leucine, ornithine, phenylalanine, aspartate, valine, and norvaline. The lysine side chain is elaborated via a 4-step sequence from the alkylation product obtained with 1-chloro-4-iodobutane as electrophile. Importantly, to our knowledge, this work represents the first asymmetric synthesis of L-alpha-vinyl analogues of m-tyrosine, ornithine, and lysine, known time-dependent inhibitors for amino acid decarboxylases.

AtFXG1, an Arabidopsis gene encoding alpha-L-fucosidase active against fucosylated xyloglucan oligosaccharides.

PubMed

de La Torre, Francisco; Sampedro, Javier; Zarra, Ignacio; Revilla, Gloria

2002-01-01

An alpha-L-fucosidase (EC 3.2.1.51) able to release the t-fucosyl residue from the side chain of xyloglucan oligosaccharides has been detected in the leaves of Arabidopsis plants. Moreover, an alpha-L-fucosidase with similar substrate specificity was purified from cabbage (Brassica oleracea) leaves to render a single band on SDS-PAGE. Two peptide sequences were obtained from this protein band, and they were used to identify an Arabidopsis gene coding for an alpha-fucosidase that we propose to call AtFXG1. In addition, an Arabidopsis gene with homology with known alpha-L-fucosidases has been also found, and we proposed to name it as AtFUC1. Both AtFXG1 and ATFUC1 were heterologously expressed in Pichia pastoris cells and the alpha-L-fucosidase activities secreted to the culture medium. The alpha-L-fucosidase encoded by AtFXG1 was active against the oligosaccharides from xyloglucan XXFG as well as against 2'-fucosyl-lactitol but not against p-nitrophenyl-alpha-L-fucopyranoside. However, the AtFUC1 heterologously expressed was active only against 2'-fucosyl-lactitol. Thus, the former must be related to xyloglucan metabolism.

H NMR studies of substrate hydrogen exchange reactions catalyzed by L-methionine gamma-lyase

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

Esaki, N.; Nakayama, T.; Sawada, S.

Hydrogen exchange reactions of various L-amino acids catalyzed by L-methionine gamma-lyase (EC 4.4.1.11) have been studied. The enzyme catalyzes the rapid exchange of the alpha- and beta-hydrogens of L-methionine and S-methyl-L-cysteine with deuterium from the solvent. The rate of alpha-hydrogen exchange was about 40 times faster than that of the enzymatic elimination reaction of the sulfur-containing amino acids. The enzyme also catalyzes the exchange reaction of alpha- and beta-hydrogens of the straight-chain L-amino acids which are not susceptible to elimination. The exchange rates of the alpha-hydrogen and the total beta-hydrogens of L-alanine and L-alpha-aminobutyrate with deuterium followed first-order kinetics. Formore » L-norvaline, L-norleucine, S-methyl-L-cysteine, and L-methionine, the rate of alpha-hydrogen exchange followed first-order kinetics, but the rate of total beta-hydrogen exchange decreased due to a primary isotope effect at the alpha-position. L-Phenylalanine and L-tryptophan slowly underwent alpha-hydrogen exchange. The pro-R hydrogen of glycine was deuterated stereospecifically.« less

Folding and trimerization of clathrin subunits at the triskelion hub.

PubMed

Näthke, I S; Heuser, J; Lupas, A; Stock, J; Turck, C W; Brodsky, F M

1992-03-06

The triskelion shape of the clathrin molecule enables it to form the polyhedral protein network that covers clathrin-coated pits and vesicles. Domains within the clathrin heavy chain that are responsible for maintaining triskelion shape and function were identified and localized. Sequences that mediate trimerization are distal to the carboxyl terminus and are adjacent to a domain that mediates both light chain binding and clathrin assembly. Structural modeling predicts that within this domain, the region of heavy chain-light chain interaction is a bundle of three or four alpha helices. These studies establish a low resolution model of clathrin subunit folding in the central portion (hub) of the triskelion, thus providing a basis for future mutagenesis experiments.

Energy contents of frequently ordered restaurant meals and comparison with human energy requirements and US Department of Agriculture database information: a multisite randomized study

USDA-ARS?s Scientific Manuscript database

BACKGROUND: Excess energy intake from meals consumed away from home is implicated as a major contributor to obesity, and ~50% of US restaurants are individual or small-chain (non-chain) establishments that do not provide nutrition information. OBJECTIVE: To measure the energy content of frequently o...

A self-consistent model of cosmic-ray fluxes and positron excess: roles of nearby pulsars and a sub-dominant source population

NASA Astrophysics Data System (ADS)

Joshi, Jagdish C.; Razzaque, Soebur

2017-09-01

The cosmic-ray positron flux calculated using the cosmic-ray nuclei interactions in our Galaxy cannot explain observed data above 10 GeV. An excess in the measured positron flux is therefore open to interpretation. Nearby pulsars, located within sub-kiloparsec range of the Solar system, are often invoked as plausible sources contributing to the excess. We show that an additional, sub-dominant population of sources together with the contributions from a few nearby pulsars can explain the latest positron excess data from the Alpha Magnetic Spectrometer (AMS). We simultaneously model, using the DRAGON code, propagation of cosmic-ray proton, Helium, electron and positron and fit their respective flux data. Our fit to the Boron to Carbon ratio data gives a diffusion spectral index of 0.45, which is close to the Kraichnan turbulent spectrum.

A self-consistent model of cosmic-ray fluxes and positron excess: roles of nearby pulsars and a sub-dominant source population

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

Joshi, Jagdish C.; Razzaque, Soebur, E-mail: jjagdish@uj.ac.za, E-mail: srazzaque@uj.ac.za

The cosmic-ray positron flux calculated using the cosmic-ray nuclei interactions in our Galaxy cannot explain observed data above 10 GeV. An excess in the measured positron flux is therefore open to interpretation. Nearby pulsars, located within sub-kiloparsec range of the Solar system, are often invoked as plausible sources contributing to the excess. We show that an additional, sub-dominant population of sources together with the contributions from a few nearby pulsars can explain the latest positron excess data from the Alpha Magnetic Spectrometer (AMS). We simultaneously model, using the DRAGON code, propagation of cosmic-ray proton, Helium, electron and positron and fitmore » their respective flux data. Our fit to the Boron to Carbon ratio data gives a diffusion spectral index of 0.45, which is close to the Kraichnan turbulent spectrum.« less

Primary amyloidosis

MedlinePlus

Amyloidosis - primary; Immunoglobulin light chain amyloidosis ... The cause of primary amyloidosis is not well understood. Genes may play a role. The condition is related to abnormal and excess production of proteins. ...

Activation of peroxisome proliferator-activated receptor-{alpha} enhances fatty acid oxidation in human adipocytes

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

Lee, Joo-Young; Hashizaki, Hikari; Goto, Tsuyoshi

2011-04-22

Highlights: {yields} PPAR{alpha} activation increased mRNA expression levels of adipocyte differentiation marker genes and GPDH activity in human adipocytes. {yields} PPAR{alpha} activation also increased insulin-dependent glucose uptake in human adipocytes. {yields} PPAR{alpha} activation did not affect lipid accumulation in human adipocytes. {yields} PPAR{alpha} activation increased fatty acid oxidation through induction of fatty acid oxidation-related genes in human adipocytes. -- Abstract: Peroxisome proliferator-activated receptor-{alpha} (PPAR{alpha}) is a key regulator for maintaining whole-body energy balance. However, the physiological functions of PPAR{alpha} in adipocytes have been unclarified. We examined the functions of PPAR{alpha} using human multipotent adipose tissue-derived stem cells as a humanmore » adipocyte model. Activation of PPAR{alpha} by GW7647, a potent PPAR{alpha} agonist, increased the mRNA expression levels of adipocyte differentiation marker genes such as PPAR{gamma}, adipocyte-specific fatty acid-binding protein, and lipoprotein lipase and increased both GPDH activity and insulin-dependent glucose uptake level. The findings indicate that PPAR{alpha} activation stimulates adipocyte differentiation. However, lipid accumulation was not changed, which is usually observed when PPAR{gamma} is activated. On the other hand, PPAR{alpha} activation by GW7647 treatment induced the mRNA expression of fatty acid oxidation-related genes such as CPT-1B and AOX in a PPAR{alpha}-dependent manner. Moreover, PPAR{alpha} activation increased the production of CO{sub 2} and acid soluble metabolites, which are products of fatty acid oxidation, and increased oxygen consumption rate in human adipocytes. The data indicate that activation of PPAR{alpha} stimulates both adipocyte differentiation and fatty acid oxidation in human adipocytes, suggesting that PPAR{alpha} agonists could improve insulin resistance without lipid accumulation in adipocytes. The expected effects of PPAR{alpha} activation are very valuable for managing diabetic conditions accompanied by obesity, because PPAR{gamma} agonists, usually used as antidiabetic drugs, induce excessive lipid accumulation in adipocytes in addition to improvement of insulin resistance.« less

Conditional Solvation Thermodynamics of Isoleucine in Model Peptides and the Limitations of the Group-Transfer Model

PubMed Central

2015-01-01

The hydration thermodynamics of the amino acid X relative to the reference G (glycine) or the hydration thermodynamics of a small-molecule analog of the side chain of X is often used to model the contribution of X to protein stability and solution thermodynamics. We consider the reasons for successes and limitations of this approach by calculating and comparing the conditional excess free energy, enthalpy, and entropy of hydration of the isoleucine side chain in zwitterionic isoleucine, in extended penta-peptides, and in helical deca-peptides. Butane in gauche conformation serves as a small-molecule analog for the isoleucine side chain. Parsing the hydrophobic and hydrophilic contributions to hydration for the side chain shows that both of these aspects of hydration are context-sensitive. Furthermore, analyzing the solute–solvent interaction contribution to the conditional excess enthalpy of the side chain shows that what is nominally considered a property of the side chain includes entirely nonobvious contributions of the background. The context-sensitivity of hydrophobic and hydrophilic hydration and the conflation of background contributions with energetics attributed to the side chain limit the ability of a single scaling factor, such as the fractional solvent exposure of the group in the protein, to map the component energetic contributions of the model-compound data to their value in the protein. But ignoring the origin of cancellations in the underlying components the group-transfer model may appear to provide a reasonable estimate of the free energy for a given error tolerance. PMID:24650057

Familial hypofibrinogenaemia associated with heterozygous substitution of a conserved arginine residue; Bbeta255 Arg-->His (Fibrinogen Merivale).

PubMed

Maghzal, Ghassan J; Brennan, Stephen O; Fellowes, Andrew P; Spearing, Ruth; George, Peter M

2003-02-21

Sequencing of all three fibrinogen genes from an individual with hypofibrinogenaemia led to the identification of two new point mutations in the Bbeta gene. Family studies showed the mutations Bbeta255 Arg-->His (Fibrinogen Merivale) and Bbeta148 Lys-->Asn (Fibrinogen Merivale II) were on different alleles and that only the Bbeta255 Arg-->His mutation segregated with hypofibrinogenaemia. Three simple heterozygotes for this mutation had mean fibrinogen concentrations of 1.4 mg/ml, while heterozygotes for the Bbeta148 Lys-->Asn mutation had normal fibrinogen concentrations. ESI MS analysis of endoproteinase Asp-N digests of Bbeta chains showed that the Bbeta255 Arg-->His substitution was not expressed in plasma, confirming it as the cause of the hypofibrinogenaemia. The Bbeta148 Lys-->Asn chains, on the other hand, were equally expressed with wild-type Bbeta chains in simple heterozygotes. Genotype analysis failed to detect either substitution in 182 healthy controls. Arg(255) is located in the first strand of the five-stranded sheet that forms the main feature of the betaD domain and appears to form an essential H bond with Gly(414). Both the Arg and Gly are absolutely conserved, not only in all known Bbeta chains, but also in all homologous alphaE and gamma chains and in all fibrinogen-related proteins. Protein instability from loss of this contact could easily explain the association of this mutation with hypofibrinogenaemia.

O-linked oligosaccharides on insulin receptor

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

Collier, E.; Gorden, P.

1991-02-01

The insulin receptor, an integral membrane glycoprotein, is synthesized as a single-chain precursor that is cleaved to produce two mature subunits, both of which contain N-linked oligosaccharide chains and covalently linked fatty acids. We report that the beta-subunit also contains O-linked oligosaccharides. The proreceptor, alpha-subunit, and beta-subunit were labeled with (3H)mannose and (3H)galactose in the presence or absence of an inhibitor of O-linked glycosylation. Tryptic peptides from each component were separated by reverse-phase high-performance liquid chromatography. N- and O-linked oligosaccharide chains were identified on these peptides by specific enzymatic digestions. The proreceptor and alpha-subunit contained only N-linked oligosaccharides, whereas themore » beta-subunit contained both N- and O-linked oligosaccharides. The O-linked oligosaccharide chains were attached to a single tryptic fraction of the beta-subunit, which also contained N-linked chains. This fraction was further localized to the NH2-terminal tryptic peptide of the beta-subunit by specific immunoprecipitation with an anti-peptide antibody with specificity for this region. Binding of insulin and autophosphorylation of the beta-subunit were not dependent on O-linked glycosylation, because cells grown in the presence of the inhibitor exhibited a normal dose response to insulin. Therefore, the insulin receptor contains O-linked oligosaccharides on the NH2-terminal tryptic peptide of the beta-subunit, and these O-linked oligosaccharides are not necessary to the binding or autophosphorylation function of the receptor.« less

Nicotinic Receptor Alpha7 Expression during Tooth Morphogenesis Reveals Functional Pleiotropy

PubMed Central

Rogers, Scott W.; Gahring, Lorise C.

2012-01-01

The expression of nicotinic acetylcholine receptor (nAChR) subtype, alpha7, was investigated in the developing teeth of mice that were modified through homologous recombination to express a bi-cistronic IRES-driven tau-enhanced green fluorescent protein (GFP); alpha7GFP) or IRES-Cre (alpha7Cre). The expression of alpha7GFP was detected first in cells of the condensing mesenchyme at embryonic (E) day E13.5 where it intensifies through E14.5. This expression ends abruptly at E15.5, but was again observed in ameloblasts of incisors at E16.5 or molar ameloblasts by E17.5–E18.5. This expression remains detectable until molar enamel deposition is completed or throughout life as in the constantly erupting mouse incisors. The expression of alpha7GFP also identifies all stages of innervation of the tooth organ. Ablation of the alpha7-cell lineage using a conditional alpha7Cre×ROSA26-LoxP(diphtheria toxin A) strategy substantially reduced the mesenchyme and this corresponded with excessive epithelium overgrowth consistent with an instructive role by these cells during ectoderm patterning. However, alpha7knock-out (KO) mice exhibited normal tooth size and shape indicating that under normal conditions alpha7 expression is dispensable to this process. The function of ameloblasts in alpha7KO mice is altered relative to controls. High resolution micro-computed tomography analysis of adult mandibular incisors revealed enamel volume of the alpha7KO was significantly reduced and the organization of enamel rods was altered relative to controls. These results demonstrate distinct and varied spatiotemporal expression of alpha7 during tooth development, and they suggest that dysfunction of this receptor would have diverse impacts upon the adult organ. PMID:22666322

Substrate specificity and kinetic properties of alpha-galactosidases from Vicia faba.

PubMed

Dey, P M; Pridham, J B

1969-10-01

1. The hydrolysis of a variety of galactosides and other glycosides by alpha-galactosidases I and II of Vicia faba was studied. 2. The effect of temperature on kinetic parameters was also examined. 3. Both enzymes are inhibited by excess of substrate (p-nitrophenyl alpha-d-galactoside); with enzyme I this is competitive and is caused by the galactosyl moiety. 4. Enzyme I is inhibited by oligosaccharides possessing terminal non-reducing galactose residues and to a smaller extent by l-arabinose and d-fucose. 5. The effect of pH on K(m) and V(max.) values suggests that carboxyl and imidazole groups are involved in the catalytic activity of enzyme I. 6. Photo-oxidation experiments with enzyme I also suggest that an imidazole group is present at the active site.

Tryptophan W207 in transducin T alpha is the fluorescence sensor of the G protein activation switch and is involved in the effector binding.

PubMed Central

Faurobert, E; Otto-Bruc, A; Chardin, P; Chabre, M

1993-01-01

We have produced a recombinant transducin alpha subunit (rT alpha) in sf9 cells, using a baculovirus system. Deletion of the myristoylation site near the N-terminal increased the solubility and allowed the purification of rT alpha. When reconstituted with excess T beta gamma on retinal membrane, rT alpha displayed functional characteristics of wild-type T alpha vis à vis its coupled receptor, rhodopsin and its effector, cGMP phosphodiesterase (PDE). We further mutated a tryptophan, W207, which is conserved in all G proteins and is suspected to elicit the fluorescence change correlated to their activation upon GDP/GTP exchange or aluminofluoride (AlFx) binding. [W207F]T alpha mutant displayed high affinity receptor binding and underwent a conformational switch upon receptor-catalysed GTP gamma S binding or upon AlFx binding, but this did not elicit any fluorescence change. Thus W207 is the only fluorescence sensor of the switch. Upon the switch the mutant remained unable to activate the PDE. To characterize better its effector-activating interaction we measured the affinity of [W207F]T alpha GDP-AlFx for PDE gamma, the effector subunit that binds most tightly to T alpha. [W207F]T alpha still bound in an activation-dependent way to PDE gamma, but with a 100-fold lower affinity than rT alpha. This suggests that W207 contributes to the G protein effector binding. Images PMID:8223434

Hypotensive effect of alpha-lipoic acid after a single administration in rats.

PubMed

Dudek, Magdalena; Razny, Katarzyna; Bilska-Wilkosz, Anna; Iciek, Malgorzata; Sapa, Jacek; Wlodek, Lidia; Filipek, Barbara

2016-05-01

The effect of alpha-lipoic acid on blood pressure was investigated many times in chronic studies, but there are no studies on the effect of this compound after a single administration. Alpha-lipoic acid is a drug used in diabetic neuropathy, often in obese patients, to treat hypertension. Therefore, knowledge of the potential antihypertensive effect of alpha-lipoic acid even after a single dose and possibly too much pressure reduction is interesting and useful. The mechanism of the hypotensive effect of alpha-lipoic acid was examined in normotensive rats in vivo after a single intraperitoneal administration, blood pressure in the left carotid artery of the rats was measured prior to the administration of the compounds (alpha- lipoic acid and/or glibenclamide) and 80 min thereafter. Alpha-lipoic acid at a dosage of 50 mg/kg b.w. i.p. significantly decreased the blood pressure from the 50th min after drug administration. This cardiovascular effect of this compound was reversed by glibenclamide, a selective KATP blocker. Glibenclamide alone at this dose did not significantly affect the blood pressure. Statistical significance was evaluated using two-way ANOVA. This suggests that alpha-lipoic acid affects ATP-dependent potassium channels. It is possible that this is an indirect effect of hydrogen sulfide because alpha-lipoic acid can increase its concentration. The results obtained in this study are very important because the patients taking alpha-lipoic acid may be treated for co-existing hypertension. Therefore, the possibility of blood pressure lowering by alpha-lipoic acid should be taken into account, although it does not lead to excessive orthostatic hypotension.

High-Molecular Compounds (Selected Articles).

DTIC Science & Technology

1987-10-15

us ions The method of potentiometric titration in dimethylformamide was used to study the structure of macro molecular chain of copolymers based on...macromolecular chain we used the method of potential metric titration . The objects of the study uere alpha chloroacrylic acid (KO)K) in monomer...homopolymer, and copolymer (with methylmethacrylate) form 131. I e d* r. I%0 I 0 12 # z Curves of potential metric titration of solutions of copolymers of KjAK

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

Antonacci, R.; Colombo, I.; Volta, M.

The electron-transfer flavoprotein (ETF), located in the mitochondrial matrix, is a nuclear-encoded enzyme delivering to the respiratory chain electrons by straight-chain acyl-CoA dehydrogenases and other dehydrogenases. ETF is composed of a 35-kDa [alpha]-subunit that is cleaved to a 32-kDa protein during mitochondrial import (ETFA) and a [beta]-subunit that reaches the mitochondrion unmodified (ETFB). The cDNA encoding both these subunits has been cloned and sequenced. 14 refs., 1 fig.

Microstructure of Mixed Surfactant Solutions by Electron Microscopy

NASA Astrophysics Data System (ADS)

Naranjo, Edward

1995-01-01

Surfactant mixtures add a new dimension to the design of complex fluid microstructure. By combining different surfactants it is not only possible to modify aggregate morphology and control the macrascopic properties of colloidal dispersions but also to produce a variety of novel synergistic phases. Mixed systems produce new microstructures by altering the intermolecular and interaggregate forces in ways impossible for single component systems. In this dissertation, we report on the phase behavior and microstructure of several synthetic and biological surfactant mixtures as elucidated by freeze-fracture and cryo-transmission electron microscopy. We have discovered that stable, spontaneous unilamellar vesicles can be prepared from aqueous mixtures of commercially available single-tailed cationic and anionic surfactants. Vesicle stability is determined by the length and volume of the hydrocarbon chains of the "catanionic" pairs. Mixtures containing bulky or branched surfactant pairs (C _{16}/C_{12 -14}) in water produce defect-free fairly monodisperse equilibrium vesicles at high dilution. In contrast, mixtures of catanionic surfactants with highly asymmetric tails (C_{16}/C_8 ) form phases of porous vesicles, dilute lamellar L_{alpha}, and anomalous isotropic L_3 phases. Images of the microstructure by freeze-fracture microscopy show that the L_3 phase consists of multiconnected self-avoiding bilayers with saddle shaped curvature. The forces between bilayers of vesicle-forming cationic and anionic surfactant mixtures were also measured using the Surface Force Apparatus (SFA). We find that the vesicles are stabilized by a long range electrostatic repulsion at large separations (>20 A) and an additional salt-independent repulsive force below 20 A. The measured forces correlate very well with the ternary phase diagram and the vesicle microstructures observed by electron microscopy. In addition to studying ionic surfactants, we have also done original work with biological surfactants. We have found that subtle changes by surfactant additives to phosphatidylcholines (PC) produce dramatic changes in the microstructure of the composite that are impossible to determine from simple scattering experiments. Novel microstructures were observed at mole ratios from 4/1 to 9/1 long chain (Di-C_{16}PC)/short chain lipid (Di-C_7PC), including disc-like micelles and rippled bilayers at room temperature. We have also observed for the first time the formation of single layered ripple phase bilayer fragments. The formation of such fragments eliminates a number of theories of formation of this unique structure that depend on coupling between bilayers. In a similar system, dimyristoyl phosphatidylcholine (DMPC) mixed with the branched alcohol geraniol produces a bluish and extremely viscoelastic phase of giant multilamellar wormy vesicles. This phase shows the Weissenberg effect under flow due to the distortion of the entangled vesicles and may be related to fluid lamellar phases and L _3 phases often seen in surfactant-alcohol -water systems. Lysophosphatidylcholine, the single-chain counterpart of the diacyl phospholipids, can also form bilayer phases when combined with long-chain fatty acids in water. The phase transition characteristics and appearance of the bilayers in equimolar mixtures of lysolipid and fatty acid are similar to those of the diacyl-PC. Electron microscopy reveals large extended multilayers in mixtures with excess lysolipid and multilamellar vesicles in mixtures with excess fatty acid.

Mitochondria mediate tumor necrosis factor-alpha/NF-kappaB signaling in skeletal muscle myotubes

NASA Technical Reports Server (NTRS)

Li, Y. P.; Atkins, C. M.; Sweatt, J. D.; Reid, M. B.; Hamilton, S. L. (Principal Investigator)

1999-01-01

Tumor necrosis factor-alpha (TNF-alpha) is implicated in muscle atrophy and weakness associated with a variety of chronic diseases. Recently, we reported that TNF-alpha directly induces muscle protein degradation in differentiated skeletal muscle myotubes, where it rapidly activates nuclear factor kappaB (NF-kappaB). We also have found that protein loss induced by TNF-alpha is NF-kappaB dependent. In the present study, we analyzed the signaling pathway by which TNF-alpha activates NF-kappaB in myotubes differentiated from C2C12 and rat primary myoblasts. We found that activation of NF-kappaB by TNF-alpha was blocked by rotenone or amytal, inhibitors of complex I of the mitochondrial respiratory chain. On the other hand, antimycin A, an inhibitor of complex III, enhanced TNF-alpha activation of NK-kappaB. These results suggest a key role of mitochondria-derived reactive oxygen species (ROS) in mediating NF-kappaB activation in muscle. In addition, we found that TNF-alpha stimulated protein kinase C (PKC) activity. However, other signal transduction mediators including ceramide, Ca2+, phospholipase A2 (PLA2), and nitric oxide (NO) do not appear to be involved in the activation of NF-kappaB.

Schindler disease: the molecular lesion in the alpha-N-acetylgalactosaminidase gene that causes an infantile neuroaxonal dystrophy.

PubMed Central

Wang, A M; Schindler, D; Desnick, R

1990-01-01

Schindler disease is a recently recognized infantile neuroaxonal dystrophy resulting from the deficient activity of the lysosomal hydrolase, alpha-N-acetylgalctosaminidase (alpha-GalNAc). The recent isolation and expression of the full-length cDNA encoding alpha-GalNAc facilitated the identification of the molecular lesions in the affected brothers from family D, the first cases described with this autosomal recessive disease. Southern and Northern hybridization analyses of DNA and RNA from the affected homozygotes revealed a grossly normal alpha-GalNAc gene structure and normal transcript sizes and amounts. Therefore, the alpha-GalNAc transcript from an affected homozygote was reverse-transcribed, amplified by the polymerase chain reaction (PCR), and sequenced. A single G to A transition at nucleotide 973 was detected in multiple subclones containing the PCR products. This point mutation resulted in a glutamic acid to lysine substitution in residue 325 (E325K) of the alpha-GalNAc polypeptide. The base substitution was confirmed by dot blot hybridization analyses of PCR-amplified genomic DNA from family members with allele-specific oligonucleotides. Furthermore, transient expression of an alpha-GalNAc construct containing the E325K mutation resulted in the expression of an immunoreactive polypeptide which had no detectable alpha-GalNAc activity. Images PMID:2243144

Pheromone induction of agglutination in Saccharomyces cerevisiae a cells

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

Terrance, K.; Lipke, P.N.

1987-10-01

a-Agglutinin, the cell surface sexual agglutinin of yeast a cells, was assayed by its ability to bind its complementary agglutinin, ..cap alpha..-agglutinin. The specific binding of /sup 125/I-..cap alpha..-agglutinin to a cells treated with the sex pheromone ..cap alpha..-factor was 2 to 2.5 times that of binding to a cells not treated with ..cap alpha..-factor. Competition with unlabeled ..cap alpha..-agglutinin revealed that the increased binding was due to increased cell surface expression of a-agglutinin, with no apparent change in the binding constant. The increase in site number was similar to the increase in cellular agglutinability. Increased expression of a-agglutinin followedmore » the same kinetics as the increase in cellular agglutinability, with a 10-min lag followed by a 15- to 20-min response time. Induction kinetics were similar in cells in phases G1 and G2 of the cell cycle. Maximal expression levels were similar in cells treated with excess pheromone and in cells exposed to pheromone after destruction of constitutively expressed a-agglutinin.« less

Characterization of the primary interaction between the mating pheromone, alpha-factor, and its receptor in Saccharomyces cerevisiae

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

Raths, S.K.

1987-01-01

Alpha-factor is a peptide of thirteen amino acids which is required for mating between the haploid mating types, a and ..cap alpha.., in Saccharomyces cerevisiae. An analogue of alpha-factor, DHP/sup 8/ DHP/sup 11/ Nle/sup 12/ tridecapeptide, was catalytically reduced in the presence of /sup 3/H gas for production of a radiolabeled pheromone suitable for use in binding studies. Incorporation of tritium resulted in /sup 3/H-alpha-factor with high specific activity, purity, biological activity and long shelf-life. Binding studies revealed that alpha-factor interacts with its receptor via a simple, reversible process which obeys the law of mass action. Association and dissociation kineticsmore » indicate values of 2.92 x 10/sup 6/ M/sup /minus/1/ min/sup -1/ for k/sub 1/ and between 4 and 7 x 10/sup /minus/2/ min/sup /minus/1/ for k/sub /minus/1/. Saturation binding studies reveal an equilibrium dissociation constant equal to 2.32 x 10/sup /minus/8/ M which approximate the kinetically-derived K/sub D/ of 2.12 x 10/sup /minus/8/ M. Scatchard and Hill analyses as well as dissociation behavior in the presence of excess unlabeled ligand indicate alpha-factor interacts with a homogeneous population of binding sites which do not interact and exhibit one affinity for the alpha-factor pheromone.« less

Commentary on Inhaled 239PuO 2 in Dogs — A Prophylaxis against Lung Cancer?

DOE PAGES

Cuttler, Jerry M.; Feinendegen, Ludwig E.

2015-01-01

Several studies on the effect of inhaled plutonium-dioxide particulates and the incidence of lung tumors in dogs reveal beneficial effects when the cumulative alpha-radiation dose is low. There is a threshold at an exposure level of about 100 cGy for excess tumor incidence and reduced lifespan. The observations conform to the expectations of the radiation hormesis dose-response model and contradict the predictions of the LNT hypothesis. These studies suggest investigating the possibility of employing low-dose alpha-radiation, such as from 239PuO 2 inhalation, as a prophylaxis against lung cancer.

Commentary on Inhaled 239PuO 2 in Dogs — A Prophylaxis against Lung Cancer?

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

Cuttler, Jerry M.; Feinendegen, Ludwig E.

Several studies on the effect of inhaled plutonium-dioxide particulates and the incidence of lung tumors in dogs reveal beneficial effects when the cumulative alpha-radiation dose is low. There is a threshold at an exposure level of about 100 cGy for excess tumor incidence and reduced lifespan. The observations conform to the expectations of the radiation hormesis dose-response model and contradict the predictions of the LNT hypothesis. These studies suggest investigating the possibility of employing low-dose alpha-radiation, such as from 239PuO 2 inhalation, as a prophylaxis against lung cancer.

SPECTROPHOTOMETRIC DETERMINATION OF ULTRA-SMALL QUANTITIES OF NICKEL IN INDIUM (in Russian)

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

Peshkova, V.M.; Bochkova, V.M.; Astakhova, E.K.

1961-09-01

alpha -Benzil doxime permits the determination of nickel by measuring optical density in the region of maximum absortption (at 275 m mu ), after the reagent excess is removed by washing the extract with alkali. Conditions were found for the spectrophotometric determination of ultra-small quantities (down to 0.005 gamma ) of nickel with alpha -benzil dioxime in the soultion of its pure salt, in the presence of cobalt and copper. A method was developed for the determination of traces of nickel down to 5 x 10 /sup -7%/ in metallic indium. The reproducibility of method is +25%. (auth)

Confirmation of radial velocity variability in Arcturus

NASA Technical Reports Server (NTRS)

Cochran, William D.

1988-01-01

The paper presents results of high-precision measurements of radial-velocity variations in Alpha Boo. Significant radial-velocity variability is detected well in excess of the random and systematic measurement errors. The radial velocity varies by an amount greater than 200 m/sec with a period of around 2 days.

An untargeted approach for the analysis of the urine peptidome of women with preeclampsia.

PubMed

Kononikhin, A S; Starodubtseva, N L; Bugrova, A E; Shirokova, V A; Chagovets, V V; Indeykina, M I; Popov, I A; Kostyukevich, Y I; Vavina, O V; Muminova, K T; Khodzhaeva, Z S; Kan, N E; Frankevich, V E; Nikolaev, E N; Sukhikh, G T

2016-10-21

Preeclampsia (PE) is a pregnancy complication characterized by high blood pressure and proteinuria. The disorder usually occurs after the 20th week of pregnancy and gets worse over time. PE increases the risk of poor outcomes for both the mother and the baby. In the study we applied LC-MS/MS method for the analysis of the urine peptidome of women with PE. Samples were prepared using size-exclusion chromatography method which gives more than twice peptides identities if compared with solid phase extraction. Thirty urine samples from women with mild and severe preeclampsia and the control group were analyzed. In total 1786 peptides were identified using complementary search engines (Mascot, MaxQuant and PEAKS). A high level of agreement in peptide identification was observed with previously published data. Label-free data comparison resulted in 35 peptides which reliably distinguished a particular PE group (severe or mild) from controls. Our results revealed unique identifications (correlate to alpha-1-antitrypsin, collagen alpha-1(I) chain, collagen alpha-1 (III) chain, and uromodulin, for instance) that can potentially serve as early indicators of PE. Copyright © 2016 Elsevier B.V. All rights reserved.

Isolation and biochemical characterisation of a novel collagen from Catostylus tagi.

PubMed

Calejo, M T; Morais, Z B; Fernandes, A I

2009-01-01

A preliminary biochemical approach to the study of collagen isolated from the medusa Catostylus tagi is reported and results are discussed in view of its use as a natural matrix for biomedical applications. Collagen from the jellyfish umbrella was isolated by pepsin digestion and purified by dialysis and salt precipitation. As expected, glycine represented almost one-third of the total amino acids. Aromatic amino-acid content was very low and imino acids were fewer than in collagens from fish and mammalian sources. Results from SDS-PAGE, ion-exchange chromatography and N-terminal amino-acid sequencing revealed an alpha1alpha2alpha3 heterotrimer, similar to vertebrate type V/XI. The molecular mass of two of the polypeptide chains was close to 85 kDa and 100 kDa for the third. However, the two chains presenting similar molecular mass, showed differences in charge and primary structure. Further characterisation showed a glycosylated protein with the carbohydrate moiety comprising almost 7% of the total mass, a denaturation temperature of 29.9 degrees C and multiple isoelectric points. Incubation with glutamyl endopeptidase resulted in significant digestion, in agreement with the protein's high content of Asp and Glu.

Cloning and characterization of the mouse alpha1C/A-adrenergic receptor gene and analysis of an alpha1C promoter in cardiac myocytes: role of an MCAT element that binds transcriptional enhancer factor-1 (TEF-1).

PubMed

O'Connell, T D; Rokosh, D G; Simpson, P C

2001-05-01

alpha1-Adrenergic receptor (AR) subtypes in the heart are expressed by myocytes but not by fibroblasts, a feature that distinguishes alpha1-ARs from beta-ARs. Here we studied myocyte-specific expression of alpha1-ARs, focusing on the subtype alpha1C (also called alpha1A), a subtype implicated in cardiac hypertrophic signaling in rat models. We first cloned the mouse alpha1C-AR gene, which consisted of two exons with an 18 kb intron, similar to the alpha1B-AR gene. The receptor coding sequence was >90% homologous to that of rat and human. alpha1C-AR transcription in mouse heart was initiated from a single Inr consensus sequence at -588 from the ATG; this and a putative polyadenylation sequence 8.5 kb 3' could account for the predominant 11 kb alpha1C mRNA in mouse heart. A 5'-nontranscribed fragment of 4.4 kb was active as a promoter in cardiac myocytes but not in fibroblasts. Promoter activity in myocytes required a single muscle CAT (MCAT) element, and this MCAT bound in vitro to recombinant and endogenous transcriptional enhancer factor-1. Thus, alpha1C-AR transcription in cardiac myocytes shares MCAT dependence with other cardiac-specific genes, including the alpha- and beta-myosin heavy chains, skeletal alpha-actin, and brain natriuretic peptide. However, the mouse alpha1C gene was not transcribed in the neonatal heart and was not activated by alpha1-AR and other hypertrophic agonists in rat myocytes, and thus differed from other MCAT-dependent genes and the rat alpha1C gene.

Crystal structure of alpha poly-p-xylylene.

NASA Technical Reports Server (NTRS)

Kubo, S.; Wunderlich, B.

1971-01-01

A crystal structure of alpha poly-p-xylylene is proposed with the help of data of oriented crystals grown during polymerization. The unit cell is monoclinic with the parameters a = 8.57 A, b = 10.62 A, c = 6.54 A (chain axis), and beta = 101.3 deg. Four repeating units per cell lead to a calculated density of 1.185 g/cu cm and a packing density of 0.71. The probable space group is P2 sub 1/m.

Involvement of Human Estrogen Related Receptor Alpha 1 (hERR 1) in Breast Cancer and Hormonally Insensitive Disease

DTIC Science & Technology

2000-08-01

SV40 early-to-late switch involves titration of cellular transcriptional repressors, Genes Dev. 7: 2206-19, 1993. 6. Bonnelye, E., Vanacker , J. M ...transcriptional regulator of the human medium-chain acyl coenzyme A dehydrogenase gene, Mol Cell Biol. 17: 5400-9, 1997. 8. Vanacker , J. M ., Bonnelye, E...related receptor-alpha), Mol Endocrinol. 13: 764-73, 1999. 9. Vanacker , J. M ., Pettersson, K., Gustafsson, J. A., and Laudet, V. Transcriptional

Quechua Amerindian population characterized by HLA-DQ alpha, YNZ22, 3'APO B, HUMTH01, and HUMVWA31A polymorphisms.

PubMed

Gené, M; Fuentes, M; Huguet, E; Piqué, E; Bert, F; Corella, A; Pérez-Pérez, A; Corbella, J; Moreno, P

1998-03-01

Allele and genotype frequencies of DNA polymorphisms were determined in a population sample of Quechua (n = 113) using the polymerase chain reaction (PCR). We report data on the frequencies of HLA-DQ alpha, YNZ22, 3'ApoB, HUMTH01 and HUMVWA31A alleles and the distribution of the different genotypes. No significant deviations between observed and expected numbers were found, thus assuming the Hardy-Weinberg equilibrium.

Molecular study of electron transfer flavoprotein alpha-subunit deficiency in two Japanese children with different phenotypes of glutaric acidemia type II.

PubMed

Purevjav, E; Kimura, M; Takusa, Y; Ohura, T; Tsuchiya, M; Hara, N; Fukao, T; Yamaguchi, S

2002-09-01

Electron transfer flavoprotein is a mitochondrial matrix protein composed of alpha- and beta-subunits (ETF alpha and ETF beta, respectively). This protein transfers electrons between several mitochondrial dehydrogenases and the main respiratory chain via ETF dehydrogenase (ETF-DH). Defects in ETF or ETF-DH cause glutaric acidemias type II (GAII). We investigated the molecular basis of ETF alpha deficiency in two Japanese children with different clinical phenotypes using expression study. Patient 1 had the severe form of GAII, a compound heterozygote of two mutations: 799G to A (alpha G267R) and nonsense 7C to T (alpha R3X). Patient 2 had the mild form and carried two heterozygous mutations: 764G to T (alpha G255V) and 478delG (frameshift). Both patients had one each of missense mutations in one allele; the others were either nonsense or truncated. Restriction enzyme digestion assay using genomic DNAs from 100 healthy Japanese revealed that these mutations were all novel. No signal for ETF alpha was detected by immunoblotting in cases of missense mutants, while wild-type cDNA resulted in expression of ETF alpha protein. Transfection with wild-type ETF alpha cDNA into cultured cells from both patients elevated incorporation of radioisotope-labelled fatty acids. These four mutations were pathogenic for GAII and missense mutations, alpha G255V and alpha G267R were considered anecdotal for mild and severe forms, respectively.

Thermodynamic confinement and alpha-helix persistence length in poly(gamma-benzyl-L-glutamate)-b-poly(dimethyl siloxane)-b-poly(gamma-benzyl-L-glutamate) triblock copolymers.

PubMed

Papadopoulos, P; Floudas, G; Schnell, I; Lieberwirth, I; Nguyen, T Q; Klok, H-A

2006-02-01

The structure and the associated dynamics of a series of poly(gamma-benzyl-L-glutamate)-b-poly(dimethyl siloxane)-b-poly(gamma-benzyl-L-glutamate) (PBLG-b-PDMS-b-PBLG) triblock copolymers were investigated using small- and wide-angle X-ray scattering, NMR, transmission electron microscopy, and dielectric spectroscopy, respectively. The structural analysis revealed phase separation in the case of the longer blocks with defected alpha-helical segments embedded within the block copolymer nanodomains. The alpha-helical persistence length was found to depend on the degree of segregation; thermodynamic confinement and chain stretching results in the partial annihilation of helical defects.

Integrating uncertainty propagation in GNSS radio occultation retrieval: from excess phase to atmospheric bending angle profiles

NASA Astrophysics Data System (ADS)

Schwarz, Jakob; Kirchengast, Gottfried; Schwaerz, Marc

2018-05-01

Global Navigation Satellite System (GNSS) radio occultation (RO) observations are highly accurate, long-term stable data sets and are globally available as a continuous record from 2001. Essential climate variables for the thermodynamic state of the free atmosphere - such as pressure, temperature, and tropospheric water vapor profiles (involving background information) - can be derived from these records, which therefore have the potential to serve as climate benchmark data. However, to exploit this potential, atmospheric profile retrievals need to be very accurate and the remaining uncertainties quantified and traced throughout the retrieval chain from raw observations to essential climate variables. The new Reference Occultation Processing System (rOPS) at the Wegener Center aims to deliver such an accurate RO retrieval chain with integrated uncertainty propagation. Here we introduce and demonstrate the algorithms implemented in the rOPS for uncertainty propagation from excess phase to atmospheric bending angle profiles, for estimated systematic and random uncertainties, including vertical error correlations and resolution estimates. We estimated systematic uncertainty profiles with the same operators as used for the basic state profiles retrieval. The random uncertainty is traced through covariance propagation and validated using Monte Carlo ensemble methods. The algorithm performance is demonstrated using test day ensembles of simulated data as well as real RO event data from the satellite missions CHAllenging Minisatellite Payload (CHAMP); Constellation Observing System for Meteorology, Ionosphere, and Climate (COSMIC); and Meteorological Operational Satellite A (MetOp). The results of the Monte Carlo validation show that our covariance propagation delivers correct uncertainty quantification from excess phase to bending angle profiles. The results from the real RO event ensembles demonstrate that the new uncertainty estimation chain performs robustly. Together with the other parts of the rOPS processing chain this part is thus ready to provide integrated uncertainty propagation through the whole RO retrieval chain for the benefit of climate monitoring and other applications.

Spiral chain structure of high pressure selenium-II{sup '} and sulfur-II from powder x-ray diffraction

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

Fujihisa, Hiroshi; Yamawaki, Hiroshi; Sakashita, Mami

2004-10-01

The structure of high pressure phases, selenium-II{sup '} (Se-II{sup '}) and sulfur-II (S-II), for {alpha}-Se{sub 8} (monoclinic Se-I) and {alpha}-S{sub 8} (orthorhombic S-I) was studied by powder x-ray diffraction experiments. Se-II{sup '} and S-II were found to be isostructural and to belong to the tetragonal space group I4{sub 1}/acd, which is made up of 16 atoms in the unit cell. The structure consisted of unique spiral chains with both 4{sub 1} and 4{sub 3} screws. The results confirmed that the structure sequence of the pressure-induced phase transitions for the group VIb elements depended on the initial molecular form. The chemicalmore » bonds of the phases are also discussed from the interatomic distances that were obtained.« less

Impact of fatty acyl composition and quantity of triglycerides on bioaccessibility of dietary carotenoids.

PubMed

Huo, Tianyao; Ferruzzi, Mario G; Schwartz, Steven J; Failla, Mark L

2007-10-31

A carotenoid-rich salad meal with varying amounts and types of triglycerides (TG) was digested using simulated gastric and small intestinal conditions. Xanthophylls (lutein and zeaxanthin) and carotenes (alpha-carotene, beta-carotene, and lycopene) in chyme and micelle fraction were quantified to determine digestive stability and efficiency of micellarization (bioaccessibility). Micellarization of lutein (+zeaxanthin) exceeded that of alpha- and beta-carotenes, which was greater than that of lycopene for all test conditions. Micellarization of carotenes, but not lutein (+zeaxanthin), was enhanced (P < 0.05) by addition of TG (2.5% v/w) to the meal and was dependent on fatty acyl chain length in structured TG (c18:1 > c8:0 > c4:0). The degree of unsaturation of c18 fatty acyl chains in TG added to the salad purée did not significantly alter the efficiency of micellarization of carotenoids. Relatively low amounts of triolein and canola oil (0.5-1%) were required for maximum micellarization of carotenes, but more oil (approximately 2.5%) was required when TG with medium chain saturated fatty acyl groups (e.g., trioctanoin and coconut oil) was added to the salad. Uptake of lutein and beta-carotene by Caco-2 cells also was examined by exposing cells to micelles generated during the simulated digestion of salad purée with either triolein or trioctanoin. Cell accumulation of beta-carotene was independent of fatty acyl composition of micelles, whereas lutein uptake was slightly, but significantly, increased from samples with digested triolein compared to trioctanoin. The results show that the in vitro transfer of alpha-carotene, beta-carotene, and lycopene from chyme to mixed micelles during digestion requires minimal (0.5-1%) lipid content in the meal and is affected by the length of fatty acyl chains but not the degree of unsaturation in TG. In contrast, fatty acyl chain length has limited if any impact on carotenoid uptake by small intestinal epithelial cells. These data suggest that the amount of TG in a typical meal does not limit the bioaccessibility of carotenoids.

Unraveling double stranded alpha-helical coiled coils: an x-ray diffraction study on hard alpha-keratin fibers.

PubMed

Kreplak, L; Doucet, J; Briki, F

2001-04-15

Transformations of proteins secondary and tertiary structures are generally studied in globular proteins in solution. In fibrous proteins, such as hard alpha-keratin, that contain long and well-defined double stranded alpha-helical coiled coil domains, such study can be directly done on the native fibrous tissue. In order to assess the structural behavior of the coiled coil domains under an axial mechanical stress, wide angle x-ray scattering and small angle x-ray scattering experiments have been carried out on stretched horse hair fibers at relative humidity around 30%. Our observations of the three major axial spacings as a function of the applied macroscopic strain have shown two rates. Up to 4% macroscopic strain the coiled coils were slightly distorted but retained their overall conformation. Above 4% the proportion of coiled coil domains progressively decreased. The main and new result of our study is the observation of the transition from alpha-helical coiled coils to disordered chains instead of the alpha-helical coiled coil to beta-sheet transition that occurs in wet fibers.

Cardiac stem cell genetic engineering using the alphaMHC promoter.

PubMed

Bailey, Brandi; Izarra, Alberto; Alvarez, Roberto; Fischer, Kimberlee M; Cottage, Christopher T; Quijada, Pearl; Díez-Juan, Antonio; Sussman, Mark A

2009-11-01

Cardiac stem cells (CSCs) show potential as a cellular therapeutic approach to blunt tissue damage and facilitate reparative and regenerative processes after myocardial infarction. Despite multiple published reports of improvement, functional benefits remain modest using normal stem cells delivered by adoptive transfer into damaged myocardium. The goal of this study is to enhance survival and proliferation of CSCs that have undergone lineage commitment in early phases as evidenced by expression of proteins driven by the alpha-myosin heavy chain (alphaMHC) promoter. The early increased expression of survival kinases augments expansion of the cardiogenic CSC pool and subsequent daughter progeny. Normal CSCs engineered with fluorescent reporter protein constructs under control of the alphaMHC promoter show transgene protein expression, confirming activity of the promoter in CSCs. Cultured CSCs from both nontransgenic and cardiac-specific transgenic mice expressing survival kinases driven by the alphaMHC promoter were analyzed to characterize transgene expression following treatments to promote differentiation in culture. Therapeutic genes controlled by the alphaMHC promoter can be engineered into and expressed in CSCs and cardiomyocyte progeny with the goal of improving the efficacy of cardiac stem cell therapy.

Tocotrienols suppress proinflammatory markers and cyclooxygenase-2 expression in RAW264.7 macrophages.

PubMed

Yam, Mun-Li; Abdul Hafid, Sitti Rahma; Cheng, Hwee-Ming; Nesaretnam, Kalanithi

2009-09-01

Tocotrienols are powerful chain breaking antioxidant. Moreover, they are now known to exhibit various non-antioxidant properties such as anti-cancer, neuroprotective and hypocholesterolemic functions. This study was undertaken to investigate the anti-inflammatory effects of tocotrienol-rich fraction (TRF) and individual tocotrienol isoforms namely delta-, gamma-, and alpha-tocotrienol on lipopolysaccharide-stimulated RAW264.7 macrophages. The widely studied vitamin E form, alpha-tocopherol, was used as comparison. Stimulation of RAW264.7 with lipopolysaccharide induced the release of various inflammatory markers. 10 mcirog/ml of TRF and all tocotrienol isoforms significantly inhibited the production of interleukin-6 and nitric oxide. However, only alpha-tocotrienol demonstrated a significant effect in lowering tumor necrosis factor-alpha production. Besides, TRF and all tocotrienol isoforms except gamma-tocotrienol reduced prostaglandin E(2) release. It was accompanied by the down-regulation of cyclooxygenase-2 gene expression by all vitamin E forms except alpha-tocopherol. Collectively, the data suggested that tocotrienols are better anti-inflammatory agents than alpha-tocopherol and the most effective form is delta-tocotrienol.

Nuclear receptor ERR alpha and coactivator PGC-1 beta are effectors of IFN-gamma-induced host defense.

PubMed

Sonoda, Junichiro; Laganière, Josée; Mehl, Isaac R; Barish, Grant D; Chong, Ling-Wa; Li, Xiangli; Scheffler, Immo E; Mock, Dennis C; Bataille, Alain R; Robert, Francois; Lee, Chih-Hao; Giguère, Vincent; Evans, Ronald M

2007-08-01

Macrophage activation by the proinflammatory cytokine interferon-gamma (IFN-gamma) is a critical component of the host innate response to bacterial pathogenesis. However, the precise nature of the IFN-gamma-induced activation pathway is not known. Here we show using genome-wide expression and chromatin-binding profiling that IFN-gamma induces the expression of many nuclear genes encoding mitochondrial respiratory chain machinery via activation of the nuclear receptor ERR alpha (estrogen-related receptor alpha, NR3B1). Studies with macrophages lacking ERR alpha demonstrate that it is required for induction of mitochondrial reactive oxygen species (ROS) production and efficient clearance of Listeria monocytogenes (LM) in response to IFN-gamma. As a result, mice lacking ERR alpha are susceptible to LM infection, a phenotype that is localized to bone marrow-derived cells. Furthermore, we found that IFN-gamma-induced activation of ERR alpha depends on coactivator PGC-1 beta (peroxisome proliferator-activated receptor gamma coactivator-1 beta), which appears to be a direct target for the IFN-gamma/STAT-1 signaling cascade. Thus, ERR alpha and PGC-1 beta act together as a key effector of IFN-gamma-induced mitochondrial ROS production and host defense.

Behavior of a cloned murine interferon alpha/beta receptor expressed in homospecific or heterospecific background.

PubMed Central

Uzé, G; Lutfalla, G; Bandu, M T; Proudhon, D; Mogensen, K E

1992-01-01

A murine interferon (IFN) alpha/beta receptor was cloned from the IFN-sensitive L1210 cell line on the basis of its homology with the human receptor. A combination of methods that includes the screening of random-primed and oligo(dT)-primed cDNA libraries and polymerase chain reactions with a single-side specificity was used. At the amino acid level, the murine IFN-alpha/beta shows 46% identity with its human counterpart. Both human WISH cells presenting a low sensitivity to mouse IFN and a murine L1210 mutant subline that does not express the receptor have been stably transfected with the murine IFN-alpha/beta receptor. Whereas transfected human cells became sensitive to a limited number of mouse IFN-alpha/beta subtypes, the transfected murine L1210 mutant was found to be fully complemented and became sensitive to all mouse IFN-alpha/beta subtypes tested, including those that were not active on transfected human cells. These results strongly suggest that the receptor described here is implicated in the mediation of the activities of all murine IFN-alpha/beta subtypes. Images PMID:1533935

Structural analysis of N-linked carbohydrate chains of funnel web spider (Agelenopsis aperta) venom peptide isomerase.

PubMed

Shikata, Y; Ohe, H; Mano, N; Kuwada, M; Asakawa, N

1998-06-01

The structure of the N-linked carbohydrate chains of peptide isomerase from the venom of the funnel web spider (Agelenopsis aperta) has been analyzed. Carbohydrates were released from peptide isomerase by hydrazinolysis and reductively aminated with 2-aminopyridine. The fluorescent derivatives were purified by phenol/chloroform extraction, followed by size-exclusion HPLC. The structure of the purified pyridylamino (PA-) carbohydrate chains were analyzed by a combination of two-dimensional HPLC mapping, sugar composition analysis, sequential exoglycosidase digestions, and mass spectrometry. The peptide isomerase contains six kinds of N-linked carbohydrate chains of truncated high-mannose type, with a fucose alpha 1-6 linked to the reducing N-acetylglucosamine in approximately 80% of them.

Polymorphism at the merozoite surface protein-3alpha locus of Plasmodium vivax: global and local diversity.

PubMed

Bruce, M C; Galinski, M R; Barnwell, J W; Snounou, G; Day, K P

1999-10-01

Allelic diversity at the Plasmodium vivax merozoite surface protein-3alpha (PvMsp-3alpha) locus was investigated using a combined polymerase chain reaction/restriction fragment length polymorphism (PCR/RFLP) protocol. Symptomatic patient isolates from global geographic origins showed a high level of polymorphism at the nucleotide level. These samples were used to validate the sensitivity, specificity, and reproducibility of the PCR/RFLP method. It was then used to investigate PvMsp3alpha diversity in field samples from children living in a single village in a malaria-endemic region of Papua New Guinea, with the aim of assessing the usefulness of this locus as an epidemiologic marker of P. vivax infections. Eleven PvMsp-3alpha alleles were distinguishable in 16 samples with single infections, revealing extensive parasite polymorphism within this restricted area. Multiple infections were easily detected and accounted for 5 (23%) of 22 positive samples. Pairs of samples from individual children provided preliminary evidence for high turnover of P. vivax populations.

Role of water molecules in structure and energetics of Pseudomonas aeruginosa lectin I interacting with disaccharides.

PubMed

Nurisso, Alessandra; Blanchard, Bertrand; Audfray, Aymeric; Rydner, Lina; Oscarson, Stefan; Varrot, Annabelle; Imberty, Anne

2010-06-25

Calcium-dependent lectin I from Pseudomonas aeruginosa (PA-IL) binds specifically to oligosaccharides presenting an alpha-galactose residue at their nonreducing end, such as the disaccharides alphaGal1-2betaGalOMe, alphaGal1-3betaGalOMe, and alphaGal1-4betaGalOMe. This provides a unique model for studying the effect of the glycosidic linkage of the ligands on structure and thermodynamics of the complexes by means of experimental and theoretical tools. The structural features of PA-IL in complex with the three disaccharides were established by docking and molecular dynamics simulations and compared with those observed in available crystal structures, including PA-IL.alphaGal1-2betaGalOMe complex, which was solved at 2.4 A resolution and reported herein. The role of a structural bridge water molecule in the binding site of PA-IL was also elucidated through molecular dynamics simulations and free energy calculations. This water molecule establishes three very stable hydrogen bonds with O6 of nonreducing galactose, oxygen from Pro-51 main chain, and nitrogen from Gln-53 main chain of the lectin binding site. Binding free energies for PA-IL in complex with the three disaccharides were investigated, and the results were compared with the experimental data determined by titration microcalorimetry. When the bridge water molecule was included in the free energy calculations, the simulations predicted the correct binding affinity trends with the 1-2-linked disaccharide presenting three times stronger affinity ligand than the other two. These results highlight the role of the water molecule in the binding site of PA-IL and indicate that it should be taken into account when designing glycoderivatives active against P. aeruginosa adhesion.

Effect of excessive acute-phase response on liver regeneration after partial hepatectomy in rats.

PubMed

Kusashio, Kimihiko; Shimizu, Hiroaki; Kimura, Fumio; Yoshidome, Hiroyuki; Ohtsuka, Masayuki; Kato, Atsushi; Yoshitomi, Hideyuki; Furukawa, Katsunori; Fukada, Tadaomi; Miyazaki, Masaru

2009-01-01

Acute-phase response (APR) is known to have a protective role against infection and tissue injury including partial hepatectomy (HTX). In this study, the effect of excessive APR on liver regeneration was investigated using a rat hepatectomized model. Experimental APR was induced by subcutaneous injection of turpentine oil, immediately, 24 h and 48 h after 50% HTX. Serum IL-6 and hepatic alpha2-macroglobulin mRNA was evaluated in rats receiving turpentine or saline as control after HTX. DNA synthesis and PCNA labeling index in the remnant liver was also investigated. Furthermore, in vitro total hepatic protein synthesis (TPS) and secretory protein synthesis (SPS) in the remnant liver were measured. Liver regeneration was significantly inhibited after HTX in the turpentine group, compared to the control. DNA synthesis and PCNA labeling index was also significantly decreased on days 1 and 2. A remarkable increase in the serum IL-6 and hepatic alpha2-macroglobulin mRNA was found in the turpentine group. Furthermore, SPS was significantly upregulated in the turpentine group, although TPS was almost similar between the groups. Induction of excessive APR after HTX inhibits liver regeneration. Acute phase protein synthesis is remarkably upregulated within the capacity of hepatic protein synthesis, most probably by a reduction in replicative protein synthesis, suggesting that APR may be more appropriate biological response for ensuring immediate survival.

Exploiting the CNC side chain in heterocyclic rearrangements: synthesis of 4(5)-acylamino-imidazoles.

PubMed

Piccionello, Antonio Palumbo; Buscemi, Silvestre; Vivona, Nicolò; Pace, Andrea

2010-08-06

A new variation on the Boulton-Katritzky reaction is reported, namely, involving use of a CNC side chain. A novel Montmorillonite-K10 catalyzed nonreductive transamination of a 3-benzoyl-1,2,4-oxadiazole afforded a 3-(alpha-aminobenzyl)-1,2,4-oxadiazole, which was condensed with benzaldehydes to afford the corresponding imines. In the presence of strong base, these imines underwent Boulton-Katritzky-type rearrangement to afford novel 4(5)-acylaminoimidazoles.

Crystal structure of the sweet-tasting protein thaumatin II at 1.27 A

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

Masuda, Tetsuya, E-mail: t2masuda@kais.kyoto-u.ac.jp; Department Natural Resources, Graduate School of Global Environmental Studies, Kyoto University, Uji, Kyoto 611-0011; Ohta, Keisuke

2011-07-08

Highlights: {yields} X-ray crystallographic structure of sweet-tasting protein, thaumatin II, was determined at a resolution of 1.27 A. {yields} The overall structure of thaumatin II is similar to that of thaumatin I, but a slight shift of the C{alpha} atom of G96 in thaumatin II was observed. {yields} The side chain of two critical residues, 67 and 82, for sweetness was modeled in two alternative conformations. {yields} The flexibility and fluctuation of side chains at 67 and 82 seems to be suitable for interaction of thaumatin molecules with sweet receptors. -- Abstract: Thaumatin, an intensely sweet-tasting protein, elicits a sweetmore » taste sensation at 50 nM. Here the X-ray crystallographic structure of one of its variants, thaumatin II, was determined at a resolution of 1.27 A. Overall structure of thaumatin II is similar to thaumatin I, but a slight shift of the C{alpha} atom of G96 in thaumatin II was observed. Furthermore, the side chain of residue 67 in thaumatin II is highly disordered. Since residue 67 is one of two residues critical to the sweetness of thaumatin, the present results suggested that the critical positive charges at positions 67 and 82 are disordered and the flexibility and fluctuation of these side chains would be suitable for interaction of thaumatin molecules with sweet receptors.« less

An N-terminal glycine to cysteine mutation in the collagen COL1A1 gene produces moderately severe osteogenesis imperfecta

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

Wilcox, W.; Scott, L.; Cohn, D.

Osteogenesis imperfecta (OI) is usually due to mutations in the type I procollagen genes COL1A1 and COL1A2. Point mutations close to the N-terminus are generally milder than those near the C-terminus of the molecule (the gradient hypothesis of collagen mutations). We describe a patient with moderately severe OI due to a mutation in the N-terminal portion of the triple helical domain of the {alpha}1(I) chain. Electrophoretic analysis of collagen isolated from fibroblast cultures suggested the abnormal presence of a cysteine in the N-terminal portion of the {alpha}1(I) chain. Five overlapping DNA fragments amplified from fibroblast RNA were screened for mutationsmore » using single strand conformational polymorphism (SSCP) and heteroduplex analyses. Direct DNA sequence analysis of the single positive fragment demonstrated a G to T transversion, corresponding to a glycine to cysteine substitution at position 226 of the triple helical domain of the {alpha}1(I) chain. The mutation was confirmed by restriction enzyme analysis of amplified genomic DNA. The mutation was not present in fibroblasts from either phenotypically normal parent. Combining this mutation with other reported mutations, glycine to cysteine substitutions at positions 205, 211, 223, and 226 produce a moderately severe phenotype whereas flanking mutations at positions 175 and 382 produce a mild phenotype. This data supports a regional rather than a gradient model of the relationship between the nature and location of type I collagen mutations and OI phenotype.« less

Excess thermodynamics of mixtures involving xenon and light linear alkanes by computer simulation.

PubMed

Carvalho, A J Palace; Ramalho, J P Prates; Martins, Luís F G

2007-06-14

Excess molar enthalpies and excess molar volumes as a function of composition for liquid mixtures of xenon + ethane (at 161.40 K), xenon + propane (at 161.40 K) and xenon + n-butane (at 182.34 K) have been obtained by Monte Carlo computer simulations and compared with available experimental data. Simulation conditions were chosen to closely match those of the corresponding experimental results. The TraPPE-UA force field was selected among other force fields to model all the alkanes studied, whereas the one-center Lennard-Jones potential from Bohn et al. was used for xenon. The calculated H(m)(E) and V(m)(E) for all systems are negative, increasing in magnitude as the alkane chain length increases. The results for these systems were compared with experimental data and with other theoretical calculations using the SAFT approach. An excellent agreement between simulation and experimental results was found for xenon + ethane system, whereas for the remaining two systems, some deviations that become progressively more significant as the alkane chain length increases were observed.

Stimulation of H(2)O(2) generation by calcium in brain mitochondria respiring on alpha-glycerophosphate.

PubMed

Tretter, Laszlo; Takacs, Katalin; Kövér, Kinga; Adam-Vizi, Vera

2007-11-15

It has been reported recently (Tretter et al., 2007b) that in isolated guinea pig brain mitochondria supported by alpha-glycerophosphate (alpha-GP) reactive oxygen species (ROS) are produced through the reverse electron transport (RET) in the respiratory chain and by alpha-glycerophosphate dehydrogenase (alpha-GPDH). We studied the effect of calcium on the generation of H(2)O(2) as measured by the Amplex Red fluorescent assay in this model. H(2)O(2) production in alpha-GP-supported mitochondria was increased significantly in the presence of 100, 250, and 500 nM Ca(2+), respectively. In addition, Ca(2+) enhanced the membrane potential, the rate of oxygen consumption, and the NAD(P)H autofluorescence in these mitochondria. Direct measurement of alpha-GPDH activity showed that Ca(2+) stimulated the enzyme by decreasing the Km for alpha-GP. In those mitochondria where RET was eliminated by the Complex I inhibitor rotenone (2 microM) or due to depolarization by ADP (1 mM), the rate of H(2)O(2) formation was smaller and the stimulation of H(2)O(2) generation by Ca(2+) was prevented partly, but the stimulatory effect of Ca(2+) was still significant. These data indicate that in alpha-GP-supported mitochondria activation of alpha-GPDH by Ca(2+) leads to an accelerated RET-mediated ROS generation as well as to a stimulated ROS production by alpha-GPDH.

Multimechanistic Monoclonal Antibodies (MAbs) Targeting Staphylococcus aureus Alpha-Toxin and Clumping Factor A: Activity and Efficacy Comparisons of a MAb Combination and an Engineered Bispecific Antibody Approach.

PubMed

Tkaczyk, C; Kasturirangan, S; Minola, A; Jones-Nelson, O; Gunter, V; Shi, Y Y; Rosenthal, K; Aleti, V; Semenova, E; Warrener, P; Tabor, D; Stover, C K; Corti, D; Rainey, G; Sellman, B R

2017-08-01

Secreted alpha-toxin and surface-localized clumping factor A (ClfA) are key virulence determinants in Staphylococcus aureus bloodstream infections. We previously demonstrated that prophylaxis with a multimechanistic monoclonal antibody (MAb) combination against alpha-toxin (MEDI4893*) and ClfA (11H10) provided greater strain coverage and improved efficacy in an S. aureus lethal bacteremia model. Subsequently, 11H10 was found to exhibit reduced affinity and impaired inhibition of fibrinogen binding to ClfA002 expressed by members of a predominant hospital-associated methicillin-resistant S. aureus (MRSA) clone, ST5. Consequently, we identified another anti-ClfA MAb (SAR114) from human tonsillar B cells with >100-fold increased affinity for three prominent ClfA variants, including ClfA002, and potent inhibition of bacterial agglutination by 112 diverse clinical isolates. We next constructed bispecific Abs (BiSAbs) comprised of 11H10 or SAR114 as IgG scaffolds and grafted anti-alpha-toxin (MEDI4893*) single-chain variable fragment to the amino or carboxy terminus of the anti-ClfA heavy chains. Although the BiSAbs exhibited in vitro potencies similar to those of the parental MAbs, only 11H10-BiSAb, but not SAR114-BiSAb, showed protective activity in murine infection models comparable to the respective MAb combination. In vivo activity with SAR114-BiSAb was observed in infection models with S. aureus lacking ClfA. Our data suggest that high-affinity binding to ClfA sequesters the SAR114-BiSAb to the bacterial surface, thereby reducing both alpha-toxin neutralization and protection in vivo These results indicate that a MAb combination targeting ClfA and alpha-toxin is more promising for future development than the corresponding BiSAb. Copyright © 2017 Tkaczyk et al.

In vitro and in vivo expression of interleukin-1alpha and tumor necrosis factor-alpha mRNA in pemphigus vulgaris: interleukin-1alpha and tumor necrosis factor-alpha are involved in acantholysis.

PubMed

Feliciani, C; Toto, P; Amerio, P; Pour, S M; Coscione, G; Shivji, G; Wang, B; Sauder, D N

2000-01-01

Keratinocyte-derived cytokines have been implicated in the pathogenesis of a number of skin diseases. In this study we examined the possible role of keratinocyte-derived cytokines in the development of acantholysis in pemphigus vulgaris. Nineteen patients with pemphigus vulgaris, demonstrating the characteristic clinical, pathologic, and immunopathologic findings were studied. In situ immunolabeling demonstrated the presence of two cytokines interleukin-1alpha and tumor necrosis factor-alpha, in lesional and perilesional areas. Results were confirmed by reverse transcriptase-polymerase chain reaction, demonstrating overexpression of both cytokines in vivo. To study the role of these cytokines in the pathogenesis of pemphigus vulgaris both in vitro and in vivo studies were performed. The results of the in vitro study demonstrated that pemphigus vulgaris IgG induced interleukin-1alpha and tumor necrosis factor-alpha mRNA in the skin. The potential pathogenic role of these mediators was demonstrated by a blocking study using antibodies against human interleukin-1alpha and tumor necrosis factor-alpha in keratinocytes cultures. A combination of anti-interleukin-1alpha and anti-tumor necrosis factor-alpha antibodies inhibited in vitro pemphigus vulgaris IgG induced acantholysis. To confirm the role of interleukin-1 and tumor necrosis factor-alpha in pemphigus, we utilized passive transfer studies using interleukin-1 deficient mice (ICE-/-, interleukin-1beta-/-) and tumor necrosis factor-alpha receptor deficient mice (TNFR1R2-/-). Both groups demonstrated a decreased susceptibility to the passive transfer of pemphigus. Our data support the role of cytokines interleukin-1 and tumor necrosis factor-alpha in the pathogenesis of pemphigus vulgaris.

Distinct expression pattern of IFN-alpha and TNF-alpha in juvenile idiopathic arthritis synovial tissue.

PubMed

Gattorno, M; Chicha, L; Gregorio, A; Ferlito, F; Rossi, F; Jarrossay, D; Lanzavecchia, A; Martini, A; Manz, M G

2007-04-01

Recent laboratory and clinical data suggest that two prototype autoimmune diseases, systemic lupus erythematosus and rheumatoid arthritis are mainly driven by distinct cytokines, interferon (IFN)-alpha and tumour necrosis factor (TNF)-alpha, respectively. We here investigated the presence and characteristics of natural type I IFN-producing cells (IPCs), as well as IFN-alpha and TNF-alpha expression at sites of inflammation in juvenile idiopathic arthritis (JIA). Peripheral blood (PB) and synovial fluid (SF) mononuclear cells (MNCs) (n = 25 each) from JIA patients with active disease were studied. IPCs were identified as BCDA-2(+)CD123(+)HLA-DR(+)CD45RA(+) cells, and dendritic cells (DCs) as CD11c(+)CD14(-/low)lin(-) cells by flow cytometry. IPCs and DCs were analysed for Toll-like receptor-7 and -9 mRNA expression by real-time polymerase chain reaction. IFN-alpha was measured by enzyme-linked immunosorbent assay in serum, SF and in supernatants of influenza virus-infected, cultured IPCs. Synovial tissues of n = 6 additional JIA patients were analysed by immunohistochemistry using mAbs against CD123, IFN-alpha, TNF-alpha, CD3, CD19 and CD138. IPCs were enriched in SF MNCs compared with PB MNCs in all JIA patients. Influenza-induced, but no spontaneous IFN-alpha release was detected from SF IPCs, and serum and SF IFN-alpha levels were not elevated. Nonetheless, in synovial tissue IFN-alpha producing cells accumulated at inflammatory lymph-follicular-like structures, while TNF-alpha producing cells were mostly found at the lining and sublining layers. These data suggest that besides TNF-alpha-expressing cells, IFN-alpha-producing IPCs are involved in initiation, maintenance or regulation of the inflammatory response in JIA.

Comprehensive analysis of all triple helical repeats in beta-spectrins reveals patterns of selective evolutionary conservation.

PubMed

Baines, Anthony J

2003-01-01

The spectrin superfamily (spectrin, alpha-actinin, utrophin and dystrophin) has in common a triple helical repeating unit of ~106 amino acid residues. In spectrin, alpha and beta chains contain multiple copies of this repeat. beta-spectrin chains contain the majority of binding activities in spectrin and are essential for animal life. Canonical beta-spectrins have 17 repeats; beta-heavy spectrins have 30. Here, the repeats of five human beta-spectrins, plus beta-spectrins from several other vertebrates and invertebrates, have been analysed. Repeats 1, 2, 14 and 17 in canonical beta are highly conserved between invertebrates and vertebrates, and repeat 8 in some isoforms. This is consistent with conservation of critical functions, since repeats 1, 2 and 17 bind alpha-spectrin. Repeats 1 of beta-spectrins are not always detected by SMART or Pfam tools. A profile hidden Markov model of beta-spectrin repeat 1 detects alpha-actinins, but not utrophin or dystrophin. Novel examples of repeat 1 were detected in the spectraplakins MACF1, BPAG1 and plectin close to the actin-binding domain. Ankyrin binds to the C-terminal portion of repeat 14; the high conservation of this entire repeat may point to additional, undiscovered ligand-binding activities. This analysis indicates that the basic triple helical repeat pattern was adapted early in the evolution of the spectrin superfamily to encompass essential binding activities, which characterise individual repeats in proteins extant today.

Biosynthesis and processing of platelet GPIIb-IIIa in human megakaryocytes.

PubMed

Duperray, A; Berthier, R; Chagnon, E; Ryckewaert, J J; Ginsberg, M; Plow, E; Marguerie, G

1987-06-01

Platelet membrane glycoprotein IIb-IIIa forms a calcium-dependent heterodimer and constitutes the fibrinogen receptor on stimulated platelets. GPIIb is a two-chain protein containing disulfide-linked alpha and beta subunits. GPIIIa is a single chain protein. These proteins are synthesized in the bone marrow by megakaryocytes, but the study of their synthesis has been hampered by the difficulty in obtaining enriched population of megakaryocytes in large numbers. To examine the biosynthesis and processing of GPIIb-IIIa, purified human megakaryocytes were isolated from liquid cultures of cryopreserved leukocytes stem cell concentrates from patients with chronic myelogenous leukemia. Immunoprecipitation of [35S]methionine pulse-chase-labeled cell extracts by antibodies specific for the alpha or beta subunits of GPIIb indicated that GPIIb was derived from a precursor of Mr 130,000 that contains the alpha and beta subunits. This precursor was converted to GPIIb with a half-life of 4-5 h. No precursor form of GPIIIa was detected. The glycosylation of GPIIb-IIIa was examined in megakaryocytes by metabolic labeling in the presence of tunicamycin, monensin, or treatment with endoglycosidase H. The polypeptide backbones of the GPIIb and the GPIIIa have molecular masses of 120 and 90 kD, respectively. High-mannose oligosaccharides are added to these polypeptide backbones co-translationally. The GPIIb precursor is then processed with conversion of high-mannose to complex type carbohydrates yielding the mature subunits GPIIb alpha (Mr 116,000) and GPIIb beta (Mr 25,000). No posttranslational processing of GPIIIa was detected.

Llama heavy-chain antibody fragments efficiently remove toxic shock syndrome toxin 1 from plasma in vitro but not in experimental porcine septic shock.

PubMed

Brummelhuis, Walter J; Joles, Jaap A; Stam, Jord C; Adams, Hendrik; Goldschmeding, Roel; Detmers, Frank J; El Khattabi, Mohamed; Maassen, Bram T; Verrips, C Theo; Braam, Branko

2010-08-01

Staphylococcus aureus produces the superantigen toxic shock syndrome toxin 1 (TSST-1). When the bacterium invades the human circulation, this toxin can induce life-threatening gram-positive sepsis. Current sepsis treatment does not remove bacterial toxins. Variable domains of llama heavy-chain antibodies (VHH) against toxic shock syndrome toxin 1 ([alpha]-TSST-1 VHH) were previously found to be effective in vitro. We hypothesized that removing TSST-1 with [alpha]-TSST-1 VHH hemofiltration filters would ameliorate experimental sepsis in pigs. After assessing in vitro whether timely removing TSST-1 interrupted TSST-1-induced mononuclear cell TNF-[alpha] production, VHH-coated filters were applied in a porcine sepsis model. Clinical course, survival, plasma interferon [gamma], and TSST-1 levels were similar with and without VHH-coated filters as were TSST-1 concentrations before and after the VHH filter. Plasma TSST-1 levels were much lower than anticipated from the distribution of the amount of infused TSST-1, suggesting compartmentalization to space or adhesion to surface not accessible to hemofiltration or pheresis techniques. Removing TSST-1 from plasma was feasible in vitro. However, the [alpha]-TSST-1 VHH adsorption filter-based technique was ineffective in vivo, indicating that improvement of VHH-based hemofiltration is required. Sequestration likely prevented the adequate removal of TSST-1. The latter warrants further investigation of TSST-1 distribution and clearance in vivo.

Acid-base titration of melanocortin peptides: evidence of Trp rotational conformers interconversion.

PubMed

Fernandez, Roberto M; Vieira, Renata F F; Nakaie, Clóvis R; Lamy, M Teresa; Ito, Amando S

2005-01-01

Tryptophantime-resolved fluorescence was used to monitor acid-base titration properties of alpha-melanocyte stimulating hormone (alpha-MSH) and the biologically more potent analog [Nle4, D-Phe7]alpha -MSH (NDP-MSH), labeled or not with the paramagnetic amino acid probe 2,2,6,6-tetramthylpiperidine-N-oxyl-4-amino-4-carboxylic acid (Toac). Global analysis of fluorescence decay profiles measured in the pH range between 2.0 and 11.0 showed that, for each peptide, the data could be well fitted to three lifetimes whose values remained constant. The less populated short lifetime component changed little with pH and was ascribed to Trp g+ chi1 rotamer, in which electron transfer deactivation predominates over fluorescence. The long and intermediate lifetime preexponential factors interconverted along that pH interval and the result was interpreted as due to interconversion between Trp g- and trans chi1 rotamers, driven by conformational changes promoted by modifications in the ionization state of side-chain residues. The differences in the extent of interconversion in alpha-MSH and NDP-MSH are indicative of structural differences between the peptides, while titration curves suggest structural similarities between each peptide and its Toac-labeled species, in aqueous solution. Though less sensitive than fluorescence, the Toac electron spin resonance (ESR) isotropic hyperfine splitting parameter can also monitor the titration of side-chain residues located relatively far from the probe. Copyright (c) 2005 Wiley Periodicals, Inc.

Combined Use of Residual Dipolar Couplings and Solution X-ray Scattering To Rapidly Probe Rigid-Body Conformational Transitions in a Non-phosphorylatable Active-Site Mutant of the 128 kDa Enzyme I Dimer

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

Takayama, Yuki; Schwieters, Charles D.; Grishaev, Alexander

2012-10-23

The first component of the bacterial phosphotransferase system, enzyme I (EI), is a multidomain 128 kDa dimer that undergoes large rigid-body conformational transitions during the course of its catalytic cycle. Here we investigate the solution structure of a non-phosphorylatable active-site mutant in which the active-site histidine is substituted by glutamine. We show that perturbations in the relative orientations and positions of the domains and subdomains can be rapidly and reliably determined by conjoined rigid-body/torsion angle/Cartesian simulated annealing calculations driven by orientational restraints from residual dipolar couplings and shape and translation information afforded by small- and wide-angle X-ray scattering. Although histidinemore » and glutamine are isosteric, the conformational space available to a Gln side chain is larger than that for the imidazole ring of His. An additional hydrogen bond between the side chain of Gln189 located on the EIN{sup {alpha}/{beta}} subdomain and an aspartate (Asp129) on the EIN{sup {alpha}} subdomain results in a small ({approx}9{sup o}) reorientation of the EIN{sup {alpha}} and EIN{sup {alpha}/{beta}} subdomains that is in turn propagated to a larger reorientation ({approx}26{sup o}) of the EIN domain relative to the EIC dimerization domain, illustrating the positional sensitivity of the EIN domain and its constituent subdomains to small structural perturbations.« less

Axolotl hemoglobin: cDNA-derived amino acid sequences of two alpha globins and a beta globin from an adult Ambystoma mexicanum.

PubMed

Shishikura, Fumio; Takeuchi, Hiro-aki; Nagai, Takatoshi

2005-11-01

Erythrocytes of the adult axolotl, Ambystoma mexicanum, have multiple hemoglobins. We separated and purified two kinds of hemoglobin, termed major hemoglobin (Hb M) and minor hemoglobin (Hb m), from a five-year-old male by hydrophobic interaction column chromatography on Alkyl Superose. The hemoglobins have two distinct alpha type globin polypeptides (alphaM and alpham) and a common beta globin polypeptide, all of which were purified in FPLC on a reversed-phase column after S-pyridylethylation. The complete amino acid sequences of the three globin chains were determined separately using nucleotide sequencing with the assistance of protein sequencing. The mature globin molecules were composed of 141 amino acid residues for alphaM globin, 143 for alpham globin and 146 for beta globin. Comparing primary structures of the five kinds of axolotl globins, including two previously established alpha type globins from the same species, with other known globins of amphibians and representatives of other vertebrates, we constructed phylogenetic trees for amphibian hemoglobins and tetrapod hemoglobins. The molecular trees indicated that alphaM, alpham, beta and the previously known alpha major globin were adult types of globins and the other known alpha globin was a larval type. The existence of two to four more globins in the axolotl erythrocyte is predicted.

Leptin potentiates Prevotella intermedia lipopolysaccharide-induced production of TNF-alpha in monocyte-derived macrophages.

PubMed

Kim, Sung-Jo

2010-06-01

In addition to regulating body weight, leptin is also recognized for its role in the regulation of immune function and inflammation. The purpose of this study was to investigate the effect of leptin on Prevotella (P.) intermedia lipopolysaccharide (LPS)-induced tumor necrosis factor (TNF)-alpha production in differentiated THP-1 cells, a human monocytic cell line. LPS from P. intermedia ATCC 25611 was prepared by the standard hot phenol-water method. THP-1 cells were incubated in the medium supplemented with phorbol myristate acetate to induce differentiation into macrophage-like cells. The amount of TNF-alpha and interleukin-8 secreted into the culture medium was determined by enzyme-linked immunosorbent assay (ELISA). TNF-alpha and Ob-R mRNA expression levels were determined by semi-quantitative reverse transcription-polymerase chain reaction analysis. Leptin enhanced P. intermedia LPS-induced TNF-alpha production in a dose-dependent manner. Leptin modulated P. intermedia LPS-induced TNF-alpha expression predominantly at the transcriptional level. Effect of leptin on P. intermedia LPS-induced TNF-alpha production was not mediated by the leptin receptor. The ability of leptin to enhance P. intermedia LPS-induced TNF-alpha production may be important in the establishment of chronic lesion accompanied by osseous tissue destruction observed in inflammatory periodontal disease.

Differences in Beef Quality between Angus (Bos taurus taurus) and Nellore (Bos taurus indicus) Cattle through a Proteomic and Phosphoproteomic Approach.

PubMed

Rodrigues, Rafael Torres de Souza; Chizzotti, Mario Luiz; Vital, Camilo Elber; Baracat-Pereira, Maria Cristina; Barros, Edvaldo; Busato, Karina Costa; Gomes, Rafael Aparecido; Ladeira, Márcio Machado; Martins, Taiane da Silva

2017-01-01

Proteins are the major constituents of muscle and are key molecules regulating the metabolic changes during conversion of muscle to meat. Brazil is one of the largest exporters of beef and most Brazilian cattle are composed by zebu (Nellore) genotype. Bos indicus beef is generally leaner and tougher than Bos taurus such as Angus. The aim of this study was to compare the muscle proteomic and phosphoproteomic profile of Angus and Nellore. Seven animals of each breed previously subjected the same growth management were confined for 84 days. Proteins were extracted from Longissimus lumborum samples collected immediately after slaughter and separated by two-dimensional electrophoresis. Pro-Q Diamond stain was used in phosphoproteomics. Proteins identification was performed using matrix assisted laser desorption/ionization time-of-flight mass spectrometry. Tropomyosin alpha-1 chain, troponin-T, myosin light chain-1 fragment, cytoplasmic malate dehydrogenase, alpha-enolase and 78 kDa glucose-regulated protein were more abundant in Nellore, while myosin light chain 3, prohibitin, mitochondrial stress-70 protein and heat shock 70 kDa protein 6 were more abundant in Angus (P<0.05). Nellore had higher phosphorylation of myosin regulatory light chain-2, alpha actin-1, triosephosphate isomerase and 14-3-3 protein epsilon. However, Angus had greater phosphorylation of phosphoglucomutase-1 and troponin-T (P<0.05). Therefore, proteins involved in contraction and muscle organization, myofilaments expressed in fast or slow-twitch fibers and heat shock proteins localized in mitochondria or sarcoplasmic reticulum and involved in cell flux of calcium and apoptosis might be associated with differences in beef quality between Angus and Nellore. Furthermore, prohibitin appears to be a potential biomarker of intramuscular fat in cattle. Additionally, differences in phosphorylation of myofilaments and glycolytic enzymes could be involved with differences in muscle contraction force, susceptibility to calpain, apoptosis and postmortem glycolysis, which might also be related to differences in beef quality among Angus and Nellore.

Disruption of the processing alpha-mannosidase gene does not prevent outer chain synthesis in Saccharomyces cerevisiae.

PubMed Central

Puccia, R; Grondin, B; Herscovics, A

1993-01-01

Processing of N-linked oligosaccharides in Saccharomyces cerevisiae begins with the removal of glucose and mannose residues from Glc3Man9GlcNAc2 to form a single isomer of Man8GlcNAc2. The importance of mannose removal for subsequent outer chain synthesis was examined in strains of S. cerevisiae disrupted in the MNS1 gene encoding a specific alpha 1,2-mannosidase responsible for Man8GlcNAc2 synthesis [Camirand, Heysen, Grondin and Herscovics (1991) J. Biol. Chem. 266, 15120-15127]. Both MNS1 transcripts of 1.85 kb and 1.7 kb were not observed in Northern blots of mns1 cells (i.e. cells containing the disrupted gene). Analysis on Bio-Gel P-6 of endo-beta-N-acetylglucosaminidase-H-sensitive oligosaccharides following a 10 min pulse with [2-3H]mannose revealed similar amounts of labelled outer chains excluded from the gel in both control and mns1 cells. H.p.l.c. of the included oligosaccharides showed that a Man9GlcNAc, rather than a Man8GlcNAc, intermediate was formed in mns1 cells. Analysis of [3H]mannose-labelled core oligosaccharides from immunoprecipitated CPY and invertase by h.p.l.c. showed a similar size distribution in mns1 and control cells. Invertase immunoprecipitated from [35S]methionine-labelled mns1 cells was highly glycosylated, but migrated slightly faster than that from control cells on denaturing PAGE, indicating a small difference in glycosylation. A similar difference in mobility was observed for invertase activity stain following non-denaturing gel electrophoresis. It is concluded that the alpha-mannosidase encoded by MNS1 is the only enzyme responsible for mannose removal in vivo, and that this processing step is not essential for outer chain synthesis. Images Figure 1 Figure 4 PMID:8439291

Shotgun proteomics reveals specific modulated protein patterns in tears of patients with primary open angle glaucoma naïve to therapy.

PubMed

Pieragostino, Damiana; Agnifili, Luca; Fasanella, Vincenzo; D'Aguanno, Simona; Mastropasqua, Rodolfo; Di Ilio, Carmine; Sacchetta, Paolo; Urbani, Andrea; Del Boccio, Piero

2013-06-01

Primary open angle glaucoma (POAG) is one of the main causes of irreversible blindness worldwide. The pathogenesis of POAG is still unclear. Alteration and sclerosis of trabecular meshwork with changes in aqueous humor molecular composition seem to play the key role. Increased intraocular pressure is widely known to be the main risk factor for the onset and progression of the disease. Unfortunately, the early diagnosis of POAG still remains the main challenge. In order to provide insight into the patho-physiology of glaucoma, here we report a shotgun proteomics approach to tears of patients with POAG naïve to therapy. Our proteomics results showed 27 differential tear proteins in POAG vs. CTRL comparison (25 up regulated proteins in the POAG group and two unique proteins in the CTRL group), 16 of which were associated with inflammatory response, free radical scavenging, cell-to-cell signaling and interaction. Overall the protein modulation shown in POAG tears proves the involvement of biochemical networks linked to inflammation. Among all regulated proteins, a sub-group of 12 up-regulated proteins in naïve POAG patients were found to be down-regulated in medically controlled POAG patients treated with prostanoid analogues (PGA), as reported in our previous work (i.e., lipocalin-1, lysozyme C, lactotransferrin, proline-rich-protein 4, prolactin-inducible protein, zinc-alpha-2-glycoprotein, polymeric immunoglobulin receptor, cystatin S, Ig kappa chain C region, Ig alpha-2 chain C region, immunoglobulin J chain, Ig alpha-1 chain C region). In summary, our findings indicate that the POAG tears protein expression is a mixture of increased inflammatory proteins that could be potential biomarkers of the disease, and their regulation may be involved in the mechanism by which PGA are able to decrease the intraocular pressure in glaucoma patients.

A Single-Domain Llama Antibody Potently Inhibits the Enzymatic Activity of Botulinum Neurotoxin by Binding to the Non-Catalytic [alpha]-Exosite Binding Region

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

Dong, Jianbo; Thompson, Aaron A.; Fan, Yongfeng

2010-08-13

Ingestion or inhalation of botulinum neurotoxin (BoNT) results in botulism, a severe and frequently fatal disease. Current treatments rely on antitoxins, which, while effective, cannot reverse symptoms once BoNT has entered the neuron. For treatments that can reverse intoxication, interest has focused on developing inhibitors of the enzymatic BoNT light chain (BoNT Lc). Such inhibitors typically mimic substrate and bind in or around the substrate cleavage pocket. To explore the full range of binding sites for serotype A light chain (BoNT/A Lc) inhibitors, we created a library of non-immune llama single-domain VHH (camelid heavy-chain variable region derived from heavy-chain-only antibody)more » antibodies displayed on the surface of the yeast Saccharomyces cerevisiae. Library selection on BoNT/A Lc yielded 15 yeast-displayed VHH with equilibrium dissociation constants (K{sub d}) from 230 to 0.03 nM measured by flow cytometry. Eight of 15 VHH inhibited the cleavage of substrate SNAP25 (synaptosome-associated protein of 25,000 Da) by BoNT/A Lc. The most potent VHH (Aa1) had a solution K{sub d} for BoNT/A Lc of 1.47 x 10{sup -10} M and an IC{sub 50} (50% inhibitory concentration) of 4.7 x 10{sup -10} M and was resistant to heat denaturation and reducing conditions. To understand the mechanism by which Aa1 inhibited catalysis, we solved the X-ray crystal structure of the BoNT/A Lc-Aa1 VHH complex at 2.6 {angstrom} resolution. The structure reveals that the Aa1 VHH binds in the {alpha}-exosite of the BoNT/A Lc, far from the active site for catalysis. The study validates the utility of non-immune llama VHH libraries as a source of enzyme inhibitors and identifies the BoNT/A Lc {alpha}-exosite as a target for inhibitor development.« less

Biochemistry of snake venom neurotoxins and their application to the study of synapse. [Neurotoxins isolated from venom of the Formosan banded krait

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

Hanley, M.R.

1978-11-01

The crude venom of the Formosan banded krait, Bungarus multicinctus, was separated into eleven lethal protein fractions. Nine fractions were purified to final homogeneous toxins, designated ..cap alpha..-bungarotoxin, ..beta..-bungarotoxin, and toxins 7, 8, 9A, 11, 12, 13, and 14. Three of the toxins, ..cap alpha..-bungarotoxin, 7, and 8, were identified as post-synaptic curarimimetic neurotoxins. The remaining toxins were identified as pre-synaptic neurotoxins. ..cap alpha..-Bungarotoxin, toxin 7, and toxin 8 are all highly stable basic polypeptides of approx. 8000 daltons molecular weight. The pre-synaptic toxins fell into two structural groups: toxin 9A and 14 which were single basic chains of approx.more » 14,000 daltons, and ..beta..-bungarotoxin, and toxins 11 thru 13 which were composed of two chains of approx. 8000 and approx. 13,000 daltons covalently linked by disulfides. All the pre-synaptic neurotoxins were shown to have intrinsic calcium-dependent phospholipase A activities. Under certain conditions, intact synaptic membranes were hydrolyzed more rapidly than protein-free extracted synaptic-lipid liposomes which, in turn, were hydrolyzed more rapidly than any other tested liposomes. It was speculated that cell-surface arrays of phosphatidyl serine/glycolipids created high affinity target sites for ..beta..-bungarotoxin. Single-chain toxins were found to be qualitatively different from the two-chain toxins in their ability to block the functioning of acetylcholine receptors, and were quantitatively different in their enzymatic and membrane disruptive activities. ..beta..-Bungarotoxin was shown to be an extremely potent neuronal lesioning agent. There was no apparent selectivity for cholinergic over non-cholinergic neurons, nor for nerve terminals over cell bodies. It was suggested that ..beta..-bungarotoxin can be considered a useful new histological tool, which may exhibit some regional selectivity.« less

Differences in Beef Quality between Angus (Bos taurus taurus) and Nellore (Bos taurus indicus) Cattle through a Proteomic and Phosphoproteomic Approach

PubMed Central

Chizzotti, Mario Luiz; Vital, Camilo Elber; Baracat-Pereira, Maria Cristina; Barros, Edvaldo; Busato, Karina Costa; Gomes, Rafael Aparecido; Ladeira, Márcio Machado; Martins, Taiane da Silva

2017-01-01

Proteins are the major constituents of muscle and are key molecules regulating the metabolic changes during conversion of muscle to meat. Brazil is one of the largest exporters of beef and most Brazilian cattle are composed by zebu (Nellore) genotype. Bos indicus beef is generally leaner and tougher than Bos taurus such as Angus. The aim of this study was to compare the muscle proteomic and phosphoproteomic profile of Angus and Nellore. Seven animals of each breed previously subjected the same growth management were confined for 84 days. Proteins were extracted from Longissimus lumborum samples collected immediately after slaughter and separated by two-dimensional electrophoresis. Pro-Q Diamond stain was used in phosphoproteomics. Proteins identification was performed using matrix assisted laser desorption/ionization time-of-flight mass spectrometry. Tropomyosin alpha-1 chain, troponin-T, myosin light chain-1 fragment, cytoplasmic malate dehydrogenase, alpha-enolase and 78 kDa glucose-regulated protein were more abundant in Nellore, while myosin light chain 3, prohibitin, mitochondrial stress-70 protein and heat shock 70 kDa protein 6 were more abundant in Angus (P<0.05). Nellore had higher phosphorylation of myosin regulatory light chain-2, alpha actin-1, triosephosphate isomerase and 14-3-3 protein epsilon. However, Angus had greater phosphorylation of phosphoglucomutase-1 and troponin-T (P<0.05). Therefore, proteins involved in contraction and muscle organization, myofilaments expressed in fast or slow-twitch fibers and heat shock proteins localized in mitochondria or sarcoplasmic reticulum and involved in cell flux of calcium and apoptosis might be associated with differences in beef quality between Angus and Nellore. Furthermore, prohibitin appears to be a potential biomarker of intramuscular fat in cattle. Additionally, differences in phosphorylation of myofilaments and glycolytic enzymes could be involved with differences in muscle contraction force, susceptibility to calpain, apoptosis and postmortem glycolysis, which might also be related to differences in beef quality among Angus and Nellore. PMID:28103301

Decorin causes autophagy in endothelial cells via Peg3

PubMed Central

Buraschi, Simone; Neill, Thomas; Goyal, Atul; Poluzzi, Chiara; Smythies, James; Owens, Rick T.; Schaefer, Liliana; Torres, Annabel; Iozzo, Renato V.

2013-01-01

Soluble decorin affects the biology of several receptor tyrosine kinases by triggering receptor internalization and degradation. We found that decorin induced paternally expressed gene 3 (Peg3), an imprinted tumor suppressor gene, and that Peg3 relocated into autophagosomes labeled by Beclin 1 and microtubule-associated light chain 3. Decorin evoked Peg3-dependent autophagy in both microvascular and macrovascular endothelial cells leading to suppression of angiogenesis. Peg3 coimmunoprecipitated with Beclin 1 and LC3 and was required for maintaining basal levels of Beclin 1. Decorin, via Peg3, induced transcription of Beclin 1 and microtubule-associated protein 1 light chain 3 alpha genes, thereby leading to a protracted autophagic program. Mechanistically, decorin interacted with VEGF receptor 2 (VEGFR2) in a region overlapping with its natural ligand VEGFA, and VEGFR2 was required for decorin-evoked Beclin 1 and microtubule-associated protein 1 light chain 3 alpha expression as well as for Peg3 induction in endothelial cells. Moreover, decorin induced VEGFR2-dependent mitochondrial fragmentation and loss of mitochondrial membrane potential. Thus, we have unveiled a mechanism for a secreted proteoglycan in inducing Peg3, a master regulator of macroautophagy in endothelial cells. PMID:23798385

Breathers and rogue waves in a Heisenberg ferromagnetic spin chain or an alpha helical protein

NASA Astrophysics Data System (ADS)

Yang, Jin-Wei; Gao, Yi-Tian; Su, Chuan-Qi; Wang, Qi-Min; Lan, Zhong-Zhou

2017-07-01

In this paper, a fourth-order variable-coefficient nonlinear Schrödinger equation for a one-dimensional continuum anisotropic Heisenberg ferromagnetic spin chain or an alpha helical protein has been investigated. Breathers and rogue waves are constructed via the Darboux transformation and generalized Darboux transformation, respectively. Results of the breathers and rogue waves are presented: (1) The first- and second-order Akhmediev breathers and Kuznetsov-Ma solitons are presented with different values of variable coefficients which are related to the energy transfer or higher-order excitations and interactions in the helical protein, or related to the spin excitations resulting from the lowest order continuum approximation and octupole-dipole interaction in a Heisenberg ferromagnetic spin chain, and the nonlinear periodic breathers resulting from the Akhmediev breathers are studied as well; (2) For the first- and second-order rogue waves, we find that they can be split into many similar components when the variable coefficients are polynomial functions of time; (3) Rogue waves can also be split when the variable coefficients are hyperbolic secant functions of time, but the profile of each component in such a case is different.

A novel anti-alpha-fetoprotein single-chain variable fragment displays anti-tumor effects in HepG2 cells as a single agent or in combination with paclitaxel.

PubMed

Ji, Xiaonan; Shen, Yanli; Sun, Hao; Gao, Xiangdong

2016-08-01

Human hepatocellular carcinoma (HCC) has a high rate of tumor recurrence and metastasis, resulting in shortened survival time. The function of alpha-fetoprotein (AFP) as a regulatory factor in the growth of HCC cells has been well defined. The aim of this study was to investigate the use of a novel AFP-specific single-chain variable fragment that blocked AFP and inhibited HCC cell growth. The results indicated that the anti-AFP single-chain variable fragment (scFv) induced growth inhibition of AFP-expressing HCC cell lines in vitro through induction of G1 cell cycle arrest and apoptosis. The mechanism of apoptosis probably involved with blocking AFP internalization and regulation of the PTEN/PI3K/Akt signaling network. Moreover, the anti-AFP-scFv also effectively sensitized the HepG2 cells to paclitaxel (PTX) at a lower concentration. The combination effect of PTX and anti-AFP-scFv displayed a synergistic effect on HepG2 cells both in vitro and in vivo. Our results demonstrated that targeting AFP by specific antibodies has potential immunotherapeutic efficacy in human HCC.

Evidence for cross-linking in tomato cutin using HR-MAS NMR spectroscopy.

PubMed

Deshmukh, Ashish P; Simpson, André J; Hatcher, Patrick G

2003-11-01

Cutin is a polyester biopolymer component of plant leaf and fruit cuticles, most often associated with waxes and cuticular polysaccharides, and sometimes with another aliphatic biopolymer called cutan. Insolubility of these cuticular biopolymers has made it difficult to apply traditional analytical techniques for structure determination, because most techniques providing molecular level details require solubility. By using the relatively new technique of one and two-dimensional high-resolution magic angle spinning (HR-MAS) NMR spectroscopy, with added information from solid-state 13C NMR spectroscopy, detailed through-bond connectivities and assignments are made for cutin from Lycopersicon esculentum (tomato) fruit. Based on the data obtained, tomato cutin is found to be predominantly an aliphatic polyester with some olefinic and aromatic moieties, consistent with previous studies that employed various degradative approaches. Aside from esters, there are free primary and secondary alcohol groups, as well as free fatty acids. A significant finding is the presence of alpha-branched fatty acids/esters. Mid-chain hydroxyls appear to be generally unesterified, but esters of mid-chain hydroxyls have been identified. The alpha-branched fatty acids/esters and esters of mid-chain hydroxyls could point towards cross-linking.

Dust Attenuation and H(alpha) Star Formation Rates of Z Approx. 0.5 Galaxies

NASA Technical Reports Server (NTRS)

Ly, Chun; Malkan, Matthew A.; Kashikawa, Nobunari; Ota, Kazuaki; Shimasaku, Kazuhiro; Iye, Masanori; Currie, Thayne

2012-01-01

Using deep narrow-band and broad-band imaging, we identify 401 z approximately 0.40 and 249 z approximately 0.49 H-alpha line-emitting galaxies in the Subaru Deep Field. Compared to other H-alpha surveys at similar redshifts, our samples are unique since they probe lower H-alpha luminosities, are augmented with multi-wavelength (rest-frame 1000AA--1.5 microns) coverage, and a large fraction (20%) of our samples has already been spectroscopically confirmed. Our spectra allow us to measure the Balmer decrement for nearly 60 galaxies with H-beta detected above 5-sigma. The Balmer decrements indicate an average extinction of A(H-alpha)=0.7(uparrow){+1.4}_{-0.7} mag. We find that the Balmer decrement systematically increases with higher H-alpha luminosities and with larger stellar masses, in agreement with previous studies with sparser samples. We find that the SFRs estimated from modeling the spectral energy distribution (SED) is reliable---we derived an "intrinsic" H-alpha luminosity which is then reddened assuming the color excess from SED modeling. The SED-predicted H-alpha luminosity agrees with H-alpha narrow-band measurements over 3 dex (rms of 0.25 dex). We then use the SED SFRs to test different statistically-based dust corrections for H-alpha and find that adopting one magnitude of extinction is inappropriate: galaxies with lower luminosities are less reddened. We find that the luminosity-dependent dust correction of Hopkins et al. yields consistent results over 3 dex (rms of 0.3 dex). Our comparisons are only possible by assuming that stellar reddening is roughly half of nebular reddening. The strong correspondence argue that with SED modeling, we can derive reliable intrinsic SFRs even in the absence of H-alpha measurements at z approximately 0.5.

A SUZAKU OBSERVATION OF Mkn 590 REVEALS A VANISHING SOFT EXCESS

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

Rivers, Elizabeth; Markowitz, Alex; Rothschild, Richard

2012-11-01

We have analyzed a long-look Suzaku observation of the Seyfert 1.2 Mkn 590. We aimed to measure the Compton reflection strength, Fe K complex properties, and soft excess emission as had been observed previously in this source. The Compton reflection strength was measured to be in the range 0.2-1.0 depending on the model used. A moderately strong Fe K{alpha} emission line was detected with an equivalent width of {approx}120 {+-} 25 eV and an Fe K{beta} line was identified with an equivalent width of {approx}30 {+-} 20 eV, although we could not rule out contribution from ionized Fe emission atmore » this energy. Surprisingly, we found no evidence for soft excess emission. Comparing our results with a 2004 observation from XMM-Newton we found that either the soft excess has decreased by a factor of 20-30 in 7 years or the photon index has steepened by 0.10 (with no soft excess present) while the continuum flux in the range 2-10 keV has varied only minimally (10%). This result could support recent claims that the soft excess is independent of the X-ray continuum.« less

Optical Absorption in Liquid Semiconductors

NASA Astrophysics Data System (ADS)

Bell, Florian Gene

An infrared absorption cell has been developed which is suitable for high temperature liquids which have absorptions in the range .1-10('3) cm('-1). The cell is constructed by clamping a gasket between two flat optical windows. This unique design allows the use of any optical windows chemically compatible with the liquid. The long -wavelength limit of the measurements is therefore limited only by the choice of the optical windows. The thickness of the cell can easily be set during assembly, and can be varied from 50 (mu)m to .5 cm. Measurements of the optical absorption edge were performed on the liquid alloy Se(,1-x)Tl(,x) for x = 0, .001, .002, .003, .005, .007, and .009, from the melting point up to 475(DEGREES)C. The absorption was found to be exponential in the photon energy over the experimental range from 0.3 eV to 1.2 eV. The absorption increased linearly with concentration according to the empirical relation (alpha)(,T)(h(nu)) = (alpha)(,1) + (alpha)(,2)x, and the absorption (alpha)(,1) was interpreted as the absorption in the absence of T1. (alpha)(,1) also agreed with the measured absorption in 100% Se at corresponding temperatures and energies. The excess absorption defined by (DELTA)(alpha) = (alpha)(,T)(h(nu))-(alpha)(,1) was interpreted as the absorption associated with Tl and was found to be thermally activated with an activation energy E(,t) = 0.5 eV. The exponential edge is explained as absorption on atoms immersed in strong electric fields surrounding ions. The strong fields give rise to an absorption tail similar to the Franz-Keldysh effect. A simple calculation is performed which is based on the Dow-Redfield theory of absorption in an electric field with excitonic effects included. The excess absorption at low photon energies is proportional to the square of the concentration of ions, which are proposed to exist in the liquid according to the relation C(,i) (PROPORTIONAL) x(' 1/2)(.)e('-E)t('/kT), which is the origin of the thermal activation and the proportionality to Tl concentration. The ionic model satisfactorily explains the observed concentration and temperature dependence of the absorption. It also provides for the first time, a universal explanation of the exponential edge in liquid semiconductors where charged defects are present, and provides a means of measuring the concentration of ions when the absorption can be calibrated.

Topological side-chain classification of beta-turns: ideal motifs for peptidomimetic development.

PubMed

Tran, Tran Trung; McKie, Jim; Meutermans, Wim D F; Bourne, Gregory T; Andrews, Peter R; Smythe, Mark L

2005-08-01

Beta-turns are important topological motifs for biological recognition of proteins and peptides. Organic molecules that sample the side chain positions of beta-turns have shown broad binding capacity to multiple different receptors, for example benzodiazepines. Beta-turns have traditionally been classified into various types based on the backbone dihedral angles (phi2, psi2, phi3 and psi3). Indeed, 57-68% of beta-turns are currently classified into 8 different backbone families (Type I, Type II, Type I', Type II', Type VIII, Type VIa1, Type VIa2 and Type VIb and Type IV which represents unclassified beta-turns). Although this classification of beta-turns has been useful, the resulting beta-turn types are not ideal for the design of beta-turn mimetics as they do not reflect topological features of the recognition elements, the side chains. To overcome this, we have extracted beta-turns from a data set of non-homologous and high-resolution protein crystal structures. The side chain positions, as defined by C(alpha)-C(beta) vectors, of these turns have been clustered using the kth nearest neighbor clustering and filtered nearest centroid sorting algorithms. Nine clusters were obtained that cluster 90% of the data, and the average intra-cluster RMSD of the four C(alpha)-C(beta) vectors is 0.36. The nine clusters therefore represent the topology of the side chain scaffold architecture of the vast majority of beta-turns. The mean structures of the nine clusters are useful for the development of beta-turn mimetics and as biological descriptors for focusing combinatorial chemistry towards biologically relevant topological space.

Detection limit of a VCO based detection chain dedicated to particles recognition and tracking

NASA Astrophysics Data System (ADS)

Coulié, K.; Rahajandraibe, W.; Aziza, H.; Micolau, G.; Vauché, R.

2018-01-01

A particle detection chain based on CMOS-SOI VCO circuit is presented. The solution is used for the recognition and the tracking of a given particle at circuit level. TCAD simulation of the detector has been performed on a 3×3 matrix of diodes based detector for particles recognition and tracking. The current response of the detector has been used for a case study in order to determine the ability of the chain to recognize an alpha particle crossing a 3×3 detection cell. The detection limit of the proposed solution is investigated and discussed in this paper.

Preferential V beta gene usage and lack of junctional sequence conservation among human T cell receptors specific for a tetanus toxin- derived peptide: evidence for a dominant role of a germline-encoded V region in antigen/major histocompatibility complex recognition

PubMed Central

1992-01-01

To investigate the structural and genetic basis of the T cell response to defined peptide/major histocompatibility (MHC) class II complexes in humans, we established a large panel of T cell clones (61) from donors of different HLA-DR haplotypes and reactive with a tetanus toxin- derived peptide (tt830-844) recognized in association with most DR molecules (universal peptide). By using a bacterial enterotoxin-based proliferation assay and cDNA sequencing, we found preferential use of a particular V beta region gene segment, V beta 2.1, in three of the individuals studied (64%, n = 58), irrespective of whether the peptide was presented by the DR6wcI, DR4w4, or DRw11.1 and DRw11.2 alleles, demonstrating that shared MHC class II antigens are not required for shared V beta gene use by T cell receptors (TCRs) specific for this peptide. V alpha gene use was more heterogeneous, with at least seven different V alpha segments derived from five distinct families encoding alpha chains able to pair with V beta 2.1 chains to form a tt830-844/DR- specific binding site. Several cases were found of clones restricted to different DR alleles that expressed identical V beta and (or very closely related) V alpha gene segments and that differed only in their junctional sequences. Thus, changes in the putative complementary determining region 3 (CDR3) of the TCR may, in certain cases, alter MHC specificity and maintain peptide reactivity. Finally, in contrast to what has been observed in other defined peptide/MHC systems, a striking heterogeneity was found in the junctional regions of both alpha and beta chains, even for TCRs with identical V alpha and/or V beta gene segments and the same restriction. Among 14 anti-tt830-844 clones using the V beta 2.1 gene segment, 14 unique V beta-D-J beta junctions were found, with no evident conservation in length and/or amino acid composition. One interpretation for this apparent lack of coselection of specific junctional sequences in the context of a common V element, V beta 2.1, is that this V region plays a dominant role in the recognition of the tt830-844/DR complex. PMID:1371303

Innate immune reactivity of the liver in rats fed a choline-deficient L-amino-acid-defined diet.

PubMed

Kawaratani, Hideto; Tsujimoto, Tatsuhiro; Kitazawa, Toshiyuki; Kitade, Mitsuteru; Yoshiji, Hitoshi; Uemura, Masahito; Fukui, Hiroshi

2008-11-21

To investigate the innate immune reactivity of tumor necrosis factor-alpha (TNF-alpha), Toll-like receptor 4 (TLR4), and CD14 in the liver of non-alcoholic steatohepatitis (NASH) model rats. Male F344 rats were fed a choline-deficient L-amino-acid-defined (CDAA) diet. The rats were killed after 4 or 8 wk of the diet, and their livers were removed for immunohistochemical investigation and RNA extraction. The liver specimens were immunostained for TNF-alpha, TLR4, and CD14. The gene expressions of TNF-alpha, TLR4, and CD14 were determined by reverse-transcriptase polymerase chain reaction (RT-PCR). Kupffer cells were isolated from the liver by Percoll gradient centrifugation, and were then cultured to measure TNF-alpha production. The serum and liver levels of TNF-alpha in the CDAA-fed rats increased significantly as compared with the control group, as did the immunohistochemical values and gene expressions of TNF-alpha, TLR4, and CD14 with the progression of steatohepatitis. TNF-alpha production from the isolated Kupffer cells of the CDAA-fed rats was elevated by lipopolysaccharide stimulation. The expressions of TNF-alpha, TLR4, and CD14 increased in the NASH model, suggesting that TLR4 and CD14-mediated endotoxin liver damage may also occur in NASH.

[Effect of TNF-alpha gene polymorphism on outcome of thalidomide-based regimens for multiple myeloma].

PubMed

DU, Juan; Yuan, Zhen-Gang; Zhang, Chun-Yang; Fu, Wei-Jun; Jiang, Hua; Chen, Bao-An; Hou, Jian

2009-10-01

To evaluate the effect of polymorphism at the -238 and -308 position of the TNF-alpha promotor region on the clinical outcome of thalidomide (Thal)-based regimens for the treatment of multiple myeloma (MM). The polymorphism at the -238 and -308 position of the TNF-alpha promotor region of 168 MM patients treated with Thal-based regimens were determined by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Genotypes were tested for association with overall response by logistic regression, and survival was evaluated by univariate and multivariate analysis. In TNF-alpha -238 position, 11 (6.5%) patients had GA genotype and 1 (0.6%) AA genotype. In TNF-alpha -308 position, 19 (11.3%) had GA genotype and 1 (0.6%) AA genotype. In univariate analysis, the TNF-alpha -238 GA + AA genotypes were associated with a significantly prolonged progression free survival (PFS) (P = 0.017), and a better overall survival (OS) (P = 0.150). Multivariate COX regression analysis showed that TNF-alpha -238 polymorphic status was an independent prognostic factor for prolonged PFS (P = 0.049). The TNF-alpha -238 polymorphic status is associated with a favorable clinical outcome in MM patients treated with thalidomide-based regimen. The polymorphism status of TNF-alpha gene might be of promise for developing a more informative stratification system for MM.

Stable, inducible thermoacidophilic alpha-amylase from Bacillus acidocaldarius.

PubMed Central

Buonocore, V; Caporale, C; De Rosa, M; Gambacorta, A

1976-01-01

Bacillus acidocaldarius Agnano 101 produces an inducible thermoacidophilic alpha-amylase. The enzyme production occurs during the stationary phase of growth in the presence of compounds with alpha-1,4-glucosidic linkages. The enzymatic activity is both present in the culture medium and associated with the cells; the enzymes purified from both sources show identical molecular and catalytic properties. The purified amylase has a single polypeptide chain of molecular weight 68,000 and behaves like an alpha-amylase with affinity constants for starch and related substances of 0.8 to 0.9 mg/ml. The pH and temperature optima for activity are 3.5 and 75degreesC, respectively. The amylase is stable at acidic pH (below 4.5). Its thermal stability is strictly dependent upon protein concentration; the half-life at 60degreesC of the amylase in a 70-mug/ml solution is about 5 days. PMID:10276

Energy-dependent etching-related impacts on CR-39 alpha detection efficiency for the Rn-222 and Rn-220 decay chains

NASA Astrophysics Data System (ADS)

Tan, Y.; Yuan, H.; Kearfott, K. J.

2018-04-01

CR-39 detectors are widely used to measure environmental levels of Rn-222, Rn-220 and their progeny. Prior research reported the CR-39 detection efficiency for alpha particles from Rn-222, Rn-220 and their progeny under a variety of etching conditions. This paper provides an explanation for interesting observations included in that work, namely that the critical incidence angle decreases with the increasing particle energy and the detection efficiency for 8.78 MeV alpha particles is zero. This paper explains these phenomena from a consideration of the interaction of alpha particles with the CR-39 detectors and the physics of etching dynamics. The proposed theory provides a rationale for an approach to optimizing the etching conditions of CR-39 detector for measuring Rn-222, Rn-220 and their progenies.

[Atopy and interleukin-4 receptor].

PubMed

Izuhara, K

1999-06-01

Both IL-4 and IL-13 induce IgE synthesis in B cells by binding to their functional receptors on target cells. These receptors are considered to be composed of heterodimers and both share the IL-4R alpha chain (IL-4R alpha) as a component. Atopy is an inherited tendency, underlying asthma, rhinitis and eczema, and generating high nonspecific IgE and/or high specific IgE against common antigens. Based on findings concerning the molecular mechanism of the signal transduction of IL-4 and IL-13, IL-4R alpha was considered a gene that gave rise to atopy. One polymorphism in the IL-4R alpha gene, Ile50Val, has been correlated with atopy by both genetic and functional assessment. The strategy used in these studies should lead to identification of other genes involved in atopy. Furthermore, these studies should be useful for gene diagnosis of atopy and development of new therapies for atopy in the future.

N-substituted imidazolines and ethylenediamines and their action on alpha- and beta-adrenergic receptors.

PubMed

Hamada, A; Yaden, E L; Horng, J S; Ruffolo, R R; Patil, P N; Miller, D D

1985-09-01

A series of N-substituted imidazolines and ethylenediamines were synthesized and examined for their activity in alpha- and beta-adrenergic systems. The length of the intermediate side chain between the catechol and imidazoline ring or the amine of the ethylenediamine segment was shown to affect the adrenergic activity. N-[2-(3,4-Dihydroxyphenyl)ethyl]imidazoline hydrochloride (2) and N-[2-(3,4-dihydroxyphenyl)ethyl]ethylenediamine dihydrochloride (4), both with two methylene groups between the catechol and amine segment, were found to be somewhat selective for alpha 2-adrenergic receptors while 1-(3,4-dihydroxybenzyl)imidazoline hydrochloride (1) and N-2-(3,4-dihydroxybenzyl)ethylenediamine dihydrochloride (3), both with one methylene group between the catechol and amine segment, were more selective for alpha1-adrenergic receptors in a pithed rat model. Of the four compounds examined, only compound 2 showed significant direct activity on beta1- and beta2-adrenergic receptors.

Characterization of salivary alpha-amylase binding to Streptococcus sanguis

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

Scannapieco, F.A.; Bergey, E.J.; Reddy, M.S.

1989-09-01

The purpose of this study was to identify the major salivary components which interact with oral bacteria and to determine the mechanism(s) responsible for their binding to the bacterial surface. Strains of Streptococcus sanguis, Streptococcus mitis, Streptococcus mutans, and Actinomyces viscosus were incubated for 2 h in freshly collected human submandibular-sublingual saliva (HSMSL) or parotid saliva (HPS), and bound salivary components were eluted with 2% sodium dodecyl sulfate. By sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western transfer, alpha-amylase was the prominent salivary component eluted from S. sanguis. Studies with {sup 125}I-labeled HSMSL or {sup 125}I-labeled HPS also demonstrated a componentmore » with an electrophoretic mobility identical to that of alpha-amylase which bound to S. sanguis. Purified alpha-amylase from human parotid saliva was radiolabeled and found to bind to strains of S. sanguis genotypes 1 and 3 and S. mitis genotype 2, but not to strains of other species of oral bacteria. Binding of ({sup 125}I)alpha-amylase to streptococci was saturable, calcium independent, and inhibitable by excess unlabeled alpha-amylases from a variety of sources, but not by secretory immunoglobulin A and the proline-rich glycoprotein from HPS. Reduced and alkylated alpha-amylase lost enzymatic and bacterial binding activities. Binding was inhibited by incubation with maltotriose, maltooligosaccharides, limit dextrins, and starch.« less

No need to be HAMLET or BAMLET to interact with histones: binding of monomeric alpha-lactalbumin to histones and basic poly-amino acids.

PubMed

Permyakov, Serge E; Pershikova, Irina V; Khokhlova, Tatyana I; Uversky, Vladimir N; Permyakov, Eugene A

2004-05-18

The ability of a specific complex of human alpha-lactalbumin with oleic acid (HAMLET) to induce cell death with selectivity for tumor and undifferentiated cells was shown recently to be mediated by interaction of HAMLET with histone proteins irreversibly disrupting chromatin structure [Duringer, C., et al. (2003) J. Biol. Chem. 278, 42131-42135]. Here we show that monomeric alpha-lactalbumin (alpha-LA) in the absence of fatty acids is also able to bind efficiently to the primary target of HAMLET, histone HIII, regardless of Ca(2+) content. Thus, the modification of alpha-LA by oleic acid is not required for binding to histones. We suggest that interaction of negatively charged alpha-LA with the basic histone stabilizes apo-alpha-LA and destabilizes the Ca(2+)-bound protein due to compensation for excess negative charge of alpha-LA's Ca(2+)-binding loop by positively charged residues of the histone. Spectrofluorimetric curves of titration of alpha-LA by histone H3 were well approximated by a scheme of cooperative binding of four alpha-LA molecules per molecule of histone, with an equilibrium dissociation constant of 1.0 microM. Such a stoichiometry of binding implies that the binding process is not site-specific with respect to histone and likely is driven by just electrostatic interactions. Co-incubation of positively charged poly-amino acids (poly-Lys and poly-Arg) with alpha-LA resulted in effects which were similar to those caused by histone HIII, confirming the electrostatic nature of the alpha-LA-histone interaction. In all cases that were studied, the binding was accompanied by aggregation. The data indicate that alpha-lactalbumin can be used as a basis for the design of antitumor agents, acting through disorganization of chromatin structure due to interaction between alpha-LA and histone proteins.

Antiapoptotic property of human alpha-synuclein in neuronal cell lines is associated with the inhibition of caspase-3 but not caspase-9 activity.

PubMed

Li, Wenxue; Lee, Michael K

2005-06-01

Abnormalities of alpha-synuclein (alpha-Syn) are mechanistically linked to Parkinson's disease (PD) and other alpha-synucleinopathies. To gain additional insights into the relationships between alpha-Syn expression and cell death, we examined the effects of expressing human alpha-Syn (Hualpha-Syn) variants on the cellular vulnerability to apoptotic stimuli. We show that the expression of wild-type (WT) and A30P mutant, but not A53T mutant, Hualpha-Syn leads to the protection of neuronal cell lines from apoptosis but not necrosis. Significantly, Hualpha-Syn did not protect non-neuronal cell lines from apoptosis. We also show that A53T mutant is a loss of function in regards to the antiapoptotic property since the expression of WT Hualpha-Syn with an excess of A53T mutant Hualpha-Syn leads to protection of the cells from apoptosis. The antiapoptotic property is specific to human alpha-Syn as neither beta-Syn nor mouse alpha-Syn protected cells from apoptosis, and the carboxy-terminal 20 amino acids are required for the antiapoptotic property. Analyses of capase-3 and caspase-9 activation reveal that the antiapoptotic property of Hualpha-Syn in neuronal cell lines is associated with the attenuation of caspase-3 activity without affecting the caspase-9 activity or the levels of cleaved, active caspase-3. We conclude that Hualpha-Syn modulates the activity of cleaved caspase-3 product in neuronal cell lines.

Characterization of alpha-ketobutyrate metabolism in rat tissues: effects of dietary protein and fasting.

PubMed

Steele, R D; Weber, H; Patterson, J I

1984-04-01

The oxidative decarboxylation of alpha-ketobutyrate was studied in rat tissue preparations. Decarboxylation was confined to the mitochondrial fraction and required coenzyme A, NAD, TPP and FAD for optimal activity in solubilized preparations. The pH optimum for this reaction in liver was 7.8, somewhat higher than that reported for other alpha-keto acid dehydrogenases. An apparent Km of 0.63 mM for alpha-ketobutyrate was determined for the rat liver system. Competition by other alpha-keto acids at 10 mM concentrations inhibited enzyme activity up to 75%. Tissue distribution of alpha-ketobutyrate dehydrogenase activity relative to liver activity was (in percent): liver, 100; heart, 127; brain, 63; kidney, 57; skeletal muscle, 38; and small intestine, 7. Total liver alpha-ketobutyrate dehydrogenase was decreased by 40% after a 24-hour fast. Similar results were found for kidney and heart activity. alpha-Aminobutyrate-pyruvate aminotransferase activity in liver or kidney was not affected by fasting; however, it was induced in liver by 50% after feeding a 40% casein diet for 10 days compared to rats fed a 20% casein diet. Increasing the dietary casein content from 6 through 40% of the diet resulted in about a fivefold increase in liver alpha-ketobutyrate dehydrogenase activity. The substantial extrahepatic capacity for alpha-ketobutyrate metabolism makes it unlikely that a loss of liver function results in an inability to metabolize alpha-ketobutyrate. Whether alpha-ketobutyrate is decarboxylated by a specific enzyme or by an already characterized complex such as pyruvate dehydrogenase or the branched-chain keto acid dehydrogenase remains to be established.

Magneto-structural correlation in a series of bimetallic alternating chain complexes of [CrIIIL(CN)4]n[MnIII(salpn)]n.nsolvents (L=2,2'-bipy or 9,10-phen, salpn=substituted salicyldehyde, solvents=water and methanol).

PubMed

Pan, Feng; Wang, Zhe-Ming; Gao, Song

2007-11-26

Five chain compounds based on the building block of [Cr(L)(CN)4]- (L=2,2'-bipy, 1-4; L=9,10-phen, 5) and [Mn(salpn)]+ (salpn=substituted salicyldehyde-type Schiff base in Scheme 1) have been prepared and characterized structurally and magnetically. The four compounds (1-4) consisting of [Cr(bipy)(CN)4]- units possess straight bimetallic chains as the [Cr(bpy)(CN)4]- unit links the two neighbor [Mn(salpn)]+ units with the two trans-cyanide ligands, while in 5 the chain is zigzag because the [Cr(phen)(CN)4]- unit connects the [Mn(salpn)]+ units with its two cis-cyanide ligands. The bond angles of Mn-N-C-Cr are adjusted by different coligands of salpn and bipy/phen. The chains are stacking via mainly the aromatic pi-pi-type interactions. All compounds show 3D antiferromagnetic ordering with Néel temperatures ranging from 3.7 to 8.1 K, and they are metamagnets displaying antiferromagnetic to ferrimagnetic transition at critical fields of 4.0-13.1 kOe at 1.9 K. This is due to weak interchain antiferromagnetic interactions between the ferrimagnetic bimetallic chains in the materials. The intrachain couplings (J, in cm(-1)) in the materials, between cyanide-bridged CrIII and MnIII ions, from -1.84 to -5.35 cm(-1), follow a linear relationship (J=-33+0.18alpha) to the Mn-N-C angles (alpha, in deg). In addition, the weak interchain antiferromagnetic interactions and critical fields for antiferromagnetic-ferrimagnetic transition are closely related to some of their structural factors, which were studied very superficially only referring to the separations of nearest chains in each material.

The determination of antibody to Streptococcus mutans serotypes in saliva for children ages 3 to 7 years.

PubMed

Everhart, D L; Rothenberg, K; Carter, W H; Klapper, B

1978-04-01

The saliva of 29 children ages 3 to 7 years was followed by indirect immunoflourescence to determine the antibody reacting with the 5 different serotypes of S mutans. Fluorescent antisera specific for alpha chain and gamma chain were used. Statistical analysis of the data demonstrated a significant negative correlation between antibody of immunoglobin class (IgA) to S mutans type b and the decayed, extracted and filled surfaces of deciduous teeth.

Morphology of poly-p-xylylene crystallized during polymerization.

NASA Technical Reports Server (NTRS)

Kubo, S.; Wunderlich, B.

1971-01-01

The morphology of as-polymerized poly-p-xylylene grown between -17 and 30 C is found to consist of lame llar alpha crystals oriented with the (010) plane parallel to the support surface. The crystallinity decreases with decreasing polymerization temperature. Spherulitic and nonspherulitic portions of the polymer film consist of folded chain lamellas with the chain axis parallel to the support surface. The results were obtained by small- and wide-angle X-ray measurements, electron and optical microscopy, and differential thermal analysis.

Treatment with agalsidase beta during pregnancy in Fabry disease.

PubMed

Politei, Juan M

2010-04-01

Fabry disease is an X-linked lysosomal storage disease caused by a deficiency of alpha-galactosidase A, which leads to excessive accumulation of glycosphingolipids in most tissues in the body, with life-threatening clinical consequences in the kidney, heart, and cerebrovascular system. Enzyme replacement therapy using exogenously produced alpha-galactosidase has been available for treatment of this multisystem progressive disease since 2001. Two different preparations of enzyme replacement therapy for Fabry disease are available outside of the USA: agalsidase alfa and agalsidase beta. Despite being X-linked, Fabry disease affects many female patients, and this report presents a successful pregnancy of a female patient receiving agalsidase beta.

Coaching the alpha male.

PubMed

Ludeman, Kate; Erlandson, Eddie

2004-05-01

Highly intelligent, confident, and successful, alpha males represent about 70% of all senior executives. Natural leaders, they willingly take on levels of responsibility most rational people would find overwhelming. But many of their quintessential strengths can also make alphas difficult to work with. Their self-confidence can appear domineering. Their high expectations can make them excessively critical. Their unemotional style can keep them from inspiring their teams. That's why alphas need coaching to broaden their interpersonal tool kits while preserving their strengths. Drawing from their experience coaching more than 1,000 senior executives, the authors outline an approach tailored specifically for the alpha. Coaches get the alpha's attention by inundating him with data from 360-degree feedback presented in ways he will find compelling--both hard-boiled metrics and vivid verbatim comments from colleagues about his strengths and weaknesses. A 360-degree assessment is a wake-up call for most alphas, providing undeniable proof that their behavior doesn't work nearly as well as they think it does. That paves the way for a genuine commitment to change. In order to change, the alpha must venture into unfamiliar--and often uncomfortable--psychological territory. He must admit vulnerability, accept accountability not just for his own work for others', connect with his underlying emotions, learn to motivate through a balance of criticism and validation, and become aware of unproductive behavior patterns. The goal of executive coaching is not simply to treat the alpha as an individual problem but to improve the entire team dynamic. Initial success creates an incentive to persevere, and the virtuous cycle reverberates throughout the entire organization.

Hyperthyroidism caused by a pituitary thyrotrophin-secreting tumour with excessive secretion of thyrotrophin-releasing hormone and subsequently followed by Graves' disease in a middle-aged woman.

PubMed

Kamoi, K; Mitsuma, T; Sato, H; Yokoyama, M; Washiyama, K; Tanaka, R; Arai, O; Takasu, N; Yamada, T

1985-11-01

A 46-year-old woman had signs of thyrotoxicosis and galactorrhoea. Serum immunoreactive TSH and its alpha-subunit increased in the presence of high serum triiodothyronine (T3), thyroxine (T4), and free T4 concentrations, whereas beta-subunit TSH was undetectable. Exogenous TRH failed to increase serum TSH. Serum TSH was markedly suppressed by glucocorticoid, but was increased by antithyroid drug. L-Dopa or bromocriptine partially suppressed, but nomifensine had no influence on serum TSH. Serum prolactin (Prl) was above normal and markedly increased by TRH, but depressed by bromocriptine and not suppressed by nomifensine. Plasma TRH was normal in the hyperthyroid state, but was increased by glucocorticoid and antithyroid drug. Excess thyroid hormone depressed plasma TRH concentrations. Basal serum GH levels were constantly low. Transsphenoidal removal of the tumour normalized serum hormones (T3, T4 free T4, TSH, alpha-subunit and Prl), and eradicated the clinical signs of hyperthyroidism and galactorrhoea. Histological study of the tumour tissue demonstrated both thyrotrophes and somatotrophes. A reciprocal relationship between serum TSH and T4 concentrations shifted to a higher level before but was normalized after removal of the tumour. Ten months later, the clinical signs of thyrotoxicosis and the increase in serum thyroid hormone recurred without a concomitant increase in serum TSH and its alpha-subunit. Thyroidal auto-antibodies were slightly positive, but thyrotrophin-binding inhibitor immunoglobulin (TBII) was negative. Administration of antithyroid drug produced a euthyroid state, but 3 years later, discontinuation of the treatment resulted in recurrent hyperthyroidism without suppressed plasma TRH and with no evidence of regrowth of the pituitary tumour. It is suggested that the patient initially had hyperthyroidism owing to excessive TSH secretion from the tumour caused by abnormal TRH secretion, and subsequently had hyperthyroidism owing to Graves' disease.

Sequence similarities and evolutionary relationships of microbial, plant and animal alpha-amylases.

PubMed

Janecek, S

1994-09-01

Amino acid sequence comparison of 37 alpha-amylases from microbial, plant and animal sources was performed to identify their mutual sequence similarities in addition to the five already described conserved regions. These sequence regions were examined from structure/function and evolutionary perspectives. An unrooted evolutionary tree of alpha-amylases was constructed on a subset of 55 residues from the alignment of sequence similarities along with conserved regions. The most important new information extracted from the tree was as follows: (a) the close evolutionary relationship of Alteromonas haloplanctis alpha-amylase (thermolabile enzyme from an antarctic psychrotroph) with the already known group of homologous alpha-amylases from streptomycetes, Thermomonospora curvata, insects and mammals, and (b) the remarkable 40.1% identity between starch-saccharifying Bacillus subtilis alpha-amylase and the enzyme from the ruminal bacterium Butyrivibrio fibrisolvens, an alpha-amylase with an unusually large polypeptide chain (943 residues in the mature enzyme). Due to a very high degree of similarity, the whole amino acid sequences of three groups of alpha-amylases, namely (a) fungi and yeasts, (b) plants, and (c) A. haloplanctis, streptomycetes, T. curvata, insects and mammals, were aligned independently and their unrooted distance trees were calculated using these alignments. Possible rooting of the trees was also discussed. Based on the knowledge of the location of the five disulfide bonds in the structure of pig pancreatic alpha-amylase, the possible disulfide bridges were established for each of these groups of homologous alpha-amylases.

IDENTIFYING NEARBY, YOUNG, LATE-TYPE STARS BY MEANS OF THEIR CIRCUMSTELLAR DISKS

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

Schneider, Adam; Song, Inseok; Melis, Carl

2012-10-01

It has recently been shown that a significant fraction of late-type members of nearby, very young associations (age {approx}<10 Myr) display excess emission at mid-IR wavelengths indicative of dusty circumstellar disks. We demonstrate that the detection of mid-IR excess emission can be utilized to identify new nearby, young, late-type stars including two definite new members ('TWA 33' and 'TWA 34') of the TW Hydrae Association (TWA). Both new TWA members display mid-IR excess emission in the Wide-field Infrared Survey Explorer catalog and they show proper motion and youthful spectroscopic characteristics-namely, H{alpha} emission, strong lithium absorption, and low surface gravity featuresmore » consistent with known TWA members. We also detect mid-IR excess-the first unambiguous evidence of a dusty circumstellar disk-around a previously identified UV-bright, young, accreting star (2M1337) that is a likely member of the Lower-Centaurus Crux region of the Scorpius-Centaurus Complex.« less

Radiation-induced myelomatosis.

PubMed

Cuzick, J

1981-01-22

It is well known that radiation can cause myeloid leukemia. However, no excess of chronic lymphocytic leukemia has been observed. Myelomatosis, like chronic lymphocytic leukemia, is a tumor of B lymphocytes. To determine whether this disease has a radiogenic origin, we surveyed all cohorts of persons exposed to radiation for which data on cancer-related mortality are available. An excess of myeloma was found in most cohorts. However, a striking deficit was found in two groups irradiated intensely for uterine neoplasms (three cases observed, 10.71 expected; P = 0.012). All other groups combined had a highly significant excess (50 observed, 22.21 expected; P = 2 X 10(-7)). The largest relative risk appeared among persons receiving internal doses of alpha-particles (14 observed, 3.24 expected; P = 2 X 10(-5)), but a significant excess (13 observed, 6.33 expected; P = 0.026) was also found in patients receiving only therapeutic or diagnostic gamma-rays or x-rays. Most cases occurred 15 to 25 years after exposure.

Species differences in the effects of prostanoids on MAP kinase phosphorylation, myosin light chain phosphorylation and contraction in bovine and cat iris sphincter smooth muscle.

PubMed

Kaddour-Djebbar, I; Ansari, H R; Akhtar, R A; Abdel-Latif, A A

2005-01-01

There is evidence from our own laboratory and that of others that EP-receptor ligands are strong contractile agonists in bovine iris sphincter and that FP-receptor agonists are strong contractile agonists in cat iris sphincter. Here, we have investigated the effects of prostaglandin (PG) receptor agonists of the FP-, EP-, TP- and DP-class on myosin light chain (MLC) phosphorylation, p42/p44 MAP kinase phosphorylation and contraction in the iris sphincter of bovine and cat. Using three signal transduction mechanism assays, namely MLC phosphorylation, MAP kinase phosphorylation and contraction, we demonstrated that in bovine iris sphincter the rank order of potency of the PG agonists in the contractile and MLC phosphorylation assays is as follows: E2>U46619>F2alpha>D2, and in cat F2alpha>D2>E2>U46619. In the MAP kinase assay, in bovine iris sphincter the rank order of potency is E2>F2alpha and in cat F2alpha>E2. These conclusions are supported by the following findings: (1) In the contractile assay, in the bovine sphincter the EC50s for PGF2alpha, PGE2, U46619 and PGD2 were found to be 1.4x10(-7), 5.0x10(-9), 9.0x10(-9) and 1.3x10(-6)M, respectively, and the corresponding values in the cat were 1.9x10(-8), 2.3x10(-7), 1.5x10(-6) and 6.9x10(-8)M, respectively. (2) In the MLC phophorylation assay, in the bovine sphincter PGF2alpha, PGE2, U46619 and PGD2 increased MLC phophorylation by 118%, 165%, 153% and 72%, respectively, and the corresponding values in cat were 175%, 99%, 90% and 95%, respectively. (3) In the MAP kinase assay, in the bovine iris sphincter PGF2alpha and PGE2, increased MAP kinase phosphorylation by 276% and 328%, respectively, and the corresponding values in cat were 308% and 245%, respectively. The data presented demonstrate pronounced species differences in the effects of the prostanoids on the MLC kinase signaling pathway in bovine and cat irides and furthermore confirm the existence of FP-receptors in that of the bovine.

Thermodynamic studies of drug-alpha-cyclodextrin interactions in water at 298.15 K: promazine hydrochloride/chlorpromazine hydrochloride + alpha-cyclodextrin + H(2)O systems.

PubMed

Terdale, Santosh S; Dagade, Dilip H; Patil, Kesharsingh J

2007-12-06

Data on osmotic coefficients have been obtained for a binary aqueous solution of two drugs, namely, promazine hydrochloride (PZ) and chlorpromazine hydrochloride (CPZ) using a vapor pressure osmometer at 298.15 K. The observed critical micelle concentration (cmc) agrees excellently with the available literature data. The measurements are extended to aqueous ternary solutions containing fixed a concentration of alpha-cyclodextrin (alpha-CD) of 0.1 mol kg(-1) and varied concentrations (approximately 0.005-0.2 mol kg(-1)) of drugs at 298.15 K. It has been found that the cmc values increase by the addition of alpha-CD. The mean molal activity coefficients of the ions and the activity coefficient of alpha-CD in binary as well as ternary solutions were obtained, which have been further used to calculate the excess Gibbs free energies and transfer Gibbs free energies. The lowering of the activity coefficients of ions and of alpha-CD is attributed to the existence of host-guest (inclusion)-type complex equilibria. It is suggested that CPZ forms 2:1 and 1:1 complexed species with alpha-CD, while PZ forms only 1:1 complexed species. The salting constant (ks) values are determined at 298.15 K for promazine-alpha-CD and chlorpromazine-alpha-CD complexes, respectively, by following the method based on the application of the McMillan-Mayer theory of virial coefficients to transfer free energy data. It is noted that the presence of chlorine in the drug molecule imparts better complexing capacity, the effect of which gets attenuated as a result of hydrophobic interaction. The results are discussed from the point of view of associative equilibria before the cmc and complexed equilibria for binary and ternary solutions, respectively.

TNF-alpha infusion impairs corpora cavernosa reactivity.

PubMed

Carneiro, Fernando S; Zemse, Saiprazad; Giachini, Fernanda R C; Carneiro, Zidonia N; Lima, Victor V; Webb, R Clinton; Tostes, Rita C

2009-03-01

Erectile dysfunction (ED), as well as cardiovascular diseases (CVDs), is associated with endothelial dysfunction and increased levels of proinflammatory cytokines, such as tumor necrosis factor-alpha (TNF-alpha). We hypothesized that increased TNF-alpha levels impair cavernosal function. In vitro organ bath studies were used to measure cavernosal reactivity in mice infused with vehicle or TNF-alpha (220 ng/kg/min) for 14 days. Gene expression of nitric oxide synthase isoforms was evaluated by real-time polymerase chain reaction. Corpora cavernosa from TNF-alpha-infused mice exhibited decreased nitric oxide (NO)-dependent relaxation, which was associated with decreased endothelial nitric oxide synthase (eNOS) and neuronal nitric oxide synthase (nNOS) cavernosal expression. Cavernosal strips from the TNF-alpha-infused mice displayed decreased nonadrenergic-noncholinergic (NANC)-induced relaxation (59.4 +/- 6.2 vs. control: 76.2 +/- 4.7; 16 Hz) compared with the control animals. These responses were associated with decreased gene expression of eNOS and nNOS (P < 0.05). Sympathetic-mediated, as well as phenylephrine (PE)-induced, contractile responses (PE-induced contraction; 1.32 +/- 0.06 vs. control: 0.9 +/- 0.09, mN) were increased in cavernosal strips from TNF-alpha-infused mice. Additionally, infusion of TNF-alpha increased cavernosal responses to endothelin-1 and endothelin receptor A subtype (ET(A)) receptor expression (P < 0.05) and slightly decreased tumor necrosis factor-alpha receptor 1 (TNFR1) expression (P = 0.063). Corpora cavernosa from TNF-alpha-infused mice display increased contractile responses and decreased NANC nerve-mediated relaxation associated with decreased eNOS and nNOS gene expression. These changes may trigger ED and indicate that TNF-alpha plays a detrimental role in erectile function. Blockade of TNF-alpha actions may represent an alternative therapeutic approach for ED, especially in pathologic conditions associated with increased levels of this cytokine.

Properties of the branched-chain 2-hydroxy acid/2-oxo acid shuttle in mouse spermatozoa.

PubMed

Coronel, C E; Gallina, F G; Gerez de Burgos, N M; Burgos, C; Blanco, A

1986-05-01

Operation of the branched-chain 2-hydroxy acid/2-oxo acid shuttle for the transfer of reducing equivalents in mitochondria of mouse spermatozoa was studied in vitro in reconstituted systems. Results show that the branched-chain 2-oxo acids within the mitochondria are offered several metabolic pathways. (a) Decarboxylation: mouse sperm mitochondria possess high branched-chain 2-oxo acid decarboxylase activity. (b) Recycling to the cytosol by using a transport system which can be inhibited by alpha-cyano-3-hydroxycinnamate and pH 6.8. (c) Transamination to the corresponding amino acids: experiments presented indicate that leucine formed from 4-methyl-2-oxopentanoate may pass to the external phase, re-initiating the cycle. These two last possibilities would allow autocatalytic operation of the shuttle. The branched-chain 2-hydroxy acids apparently do not utilize the monocarboxylate carrier to penetrate the mitochondria.

Decay properties of {sup 265}Sg(Z=106) and {sup 266}Sg(Z=106)

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

Tuerler, A.; Dressler, R.; Eichler, B.

1998-04-01

The presently known most neutron-rich isotopes of element 106 (seaborgium, Sg), {sup 265}Sg and {sup 266}Sg, were produced in the fusion reaction {sup 22}Ne+{sup 248}Cm at beam energies of 121 and 123 MeV. Using the On-Line Gas chemistry Apparatus OLGA, a continuous separation of Sg was achieved within a few seconds. Final products were assayed by {alpha}-particle and spontaneous fission (SF) spectrometry. {sup 265}Sg and {sup 266}Sg were identified by observing time correlated {alpha}-{alpha}-({alpha}) and {alpha}-SF decay chains. A total of 13 correlated decay chains of {sup 265}Sg (with an estimated number of 2.8 random correlations) and 3 decay chainsmore » of {sup 266}Sg (0.6 random correlations) were identified. Deduced decay properties were T{sub 1/2}=7.4{sub {minus}2.7}{sup +3.3} s (68{percent} c.i.) and E{sub {alpha}}=8.69 MeV (8{percent}), 8.76 MeV (23{percent}), 8.84 MeV (46{percent}), and 8.94 MeV (23{percent}) for {sup 265}Sg; and T{sub 1/2}=21{sub {minus}12}{sup +20} s (68{percent} c.i.) and E{sub {alpha}}=8.52 MeV (33{percent}) and 8.77 MeV (66{percent}) for {sup 266}Sg. The resolution of the detectors was between 50{endash}100 keV (full width at half maximum). Upper limits for SF of {le}35{percent} and {le}82{percent} were established for {sup 265}Sg and {sup 266}Sg, respectively. The upper limits for SF are given with a 16{percent} error probability. Using the lower error limits of the half-lives of {sup 265}Sg and {sup 266}Sg, the resulting lower limits for the partial SF half-lives are T{sub 1/2}{sup SF}({sup 265}Sg){ge}13 s and T{sub 1/2}{sup SF}({sup 266}Sg){ge}11 s. Correspondingly, the partial {alpha}-decay half-lives are between T{sub 1/2}{sup {alpha}}({sup 265}Sg)=4.7{endash}16.5 s (68{percent} c.i.) and T{sub 1/2}{sup {alpha}}({sup 266}Sg)=9{endash}228 s (68{percent} c.i.), using the upper and lower error limits of the half-lives of {sup 265}Sg and {sup 266}Sg. The lower limit on the partial SF half-life of {sup 266}Sg is in good agreement with theoretical predictions. Production cross sections of about 240 pb and 25 pb for the {alpha}-decay branch in {sup 265}Sg and {sup 266}Sg were estimated, respectively. {copyright} {ital 1998} {ital The American Physical Society}« less

Heterocyclic alpha-alkylidene cyclopentenones obtained via a Pauson-Khand reaction of amino acid derived allenynes. A scope and limitation study directed toward the preparation of a tricyclic pyrrole library.

PubMed

Brummond, Kay M; Curran, Dennis P; Mitasev, Branko; Fischer, Stefan

2005-03-04

The synthesis of a novel class of tricyclic pyrroles has been accomplished by using a Pauson-Khand/Stetter/Paal-Knorr reaction sequence. Full details of the Pauson-Khand reaction of amino acid tethered allenynes 4a-e and 9a-d are disclosed. The study of this reaction led to the discovery of an unprecedented substituent effect on the diastereoselectivity of the Mo(CO)6 mediated allenic Pauson-Khand reaction. It was found that amino acid tethered allenynes with aromatic side chains afford alpha-alkylidene cyclopentenones with the opposite diastereoselectivity compared to those with aliphatic side chains. This effect has been attributed to complexation of the metal mediator to the aromatic ring in the substrate. Furthermore, an isomerization of one of the diastereomers of the alpha-alkylidene cyclopentenones was encountered, leading to eventual decomposition. The stable diastereomers were found to react well in the Stetter reaction leading to 1,4-diketones that were converted to pyrroles. The observation that the first generation of 2-alkyl-substituted pyrroles was unstable led to a second generation of 2-carboxamide pyrroles with sufficient stability for biological tests which are in progress.

Exercise promotes alpha7 integrin gene transcription and protection of skeletal muscle.

PubMed

Boppart, Marni D; Volker, Sonja E; Alexander, Nicole; Burkin, Dean J; Kaufman, Stephen J

2008-11-01

The alpha7beta1 integrin is increased in skeletal muscle in response to injury-producing exercise, and transgenic overexpression of this integrin in mice protects against exercise-induced muscle damage. The present study investigates whether the increase in the alpha7beta1 integrin observed in wild-type mice in response to exercise is due to transcriptional regulation and examines whether mobilization of the integrin at the myotendinous junction (MTJ) is a key determinant in its protection against damage. A single bout of downhill running exercise selectively increased transcription of the alpha7 integrin gene in 5-wk-old wild-type mice 3 h postexercise, and an increased alpha7 chain was detected in muscle sarcolemma adjacent to tendinous tissue immediately following exercise. The alpha7B, but not alpha7A isoform, was found concentrated and colocalized with tenascin-C in muscle fibers lining the MTJ. To further validate the importance of the integrin in the protection against muscle damage following exercise, muscle injury was quantified in alpha7(-/-) mice. Muscle damage was extensive in alpha7(-/-) mice in response to both a single and repeated bouts of exercise and was largely restricted to areas of high MTJ concentration and high mechanical force near the Achilles tendon. These results suggest that exercise-induced muscle injury selectively increases transcription of the alpha7 integrin gene and promotes a rapid change in the alpha7beta integrin at the MTJ. These combined molecular and cellular alterations are likely responsible for integrin-mediated attenuation of exercise-induced muscle damage.

Cloning, sequence, and expression of a blood group B active recombinant alpha-D-galactosidase from pinto bean (Phaseolus vulgaris).

PubMed

Davis, M O; Hata, D J; Johnson, S A; Jones, D E; Harmata, M A; Evans, M L; Walker, J C; Smith, D S

1997-07-01

A cDNA encoding pinto bean alpha-D-galactosidase [E.C. 3.2.1.22] was obtained by amplification of cDNA using highly conserved sequences found in eucaryotic alpha-D-galactosidases. Subsequently a full length Phaseolus cDNA clone was obtained that is 1537 nt long and contains untranslated 5' and 3' sequences. The nucleotide sequence of the cDNA has a high degree of homology with other eucaryotic alpha-D-galactosidase genes. The recombinant alpha-D-galactosidase (rGal) was expressed in Escherichia coli and purified by ion exchange and affinity chromatography. Purified rGal was homogeneous by SDS-PAGE and had relative masses of 40.1 and 45.4 kDa under nonreducing and reducing conditions, respectively. The N-terminal sequence of the expressed protein contained the sequence GNGLGQTPPMG corresponding to that deduced from the cDNA sequence. The native molecular weight for rGal was determined to be 32.18 kDa by Sephacryl S-200 chromatography. The specific activity of the rGal was 349 mu moles of PNP-alpha-D-galactopyranoside hydrolyzed per mg of pure rGal per min. rGal was highly specific for alpha-D-galactosyl residues and degraded B oligosaccharide. No detectable hemagglutinin or protease activity was present in the preparations. Furthermore, rGal was active against the blood group B antigen on native human erythrocytes in cell suspension assays. The only detectable RBC phenotypic change was loss of the B and P1 epitopes. Recombinant Phaseolus vulgaris alpha-D-galactosidase may have useful biotechnical applications in the potential mass production of enzymatically converted, universally transfusable type O RBCs. alpha-D-galactosidase [E.C. 3.2.1.22] has been purified from a variety of procaryotic and eucaryotic species. Most alpha-D-galactosidases have similar low molecular weight substrate specificities, but activity against high molecular weight substrates is variable. Terminal alpha-D-galactoside residues are present in glycoproteins and glycolipids. Some alpha-D-galactosidases have activity against alpha-D-galactosyl residues on cell membrane glycoconjugates. Glycosidases with this property are useful for carbohydrate structural studies and biotechnical applications. Enzymes free of other glycosidase activities with activity near neutral pH are particularly useful for membrane modification studies on native cells. Complex sugar chains in glycolipids and glycoproteins have often been implicated in the growth and development of eucaryotes. In particular, complex sugar chains play an important role in the recognition of self in the immune system. Some alpha-D-galactosidases can modify certain carbohydrate membrane epitopes, thereby modulating the immune response. For example, the blood group B epitope expressed on erythrocytes contains a terminal alpha-D-galactosyl residue. Individuals lacking this antigen produce naturally occurring complement fixing antibodies to the B epitope. Hydrolysis of this terminal saccharide destroys the antigenic activity of the B determinant producing H antigen (blood type O) on erythrocytes. Only rare individuals produce clinically significant antibodies to the H antigen, and therefore, type O red blood cells are "universally" compatible and in great demand. Dhar purified alpha-D-galactosidase isozymes from Phaseolus vulgaris and characterized their activity. To our knowledge, our laboratory, in a brief report, is the first to describe the cloning of the gene and the use of recombinant enzyme for seroconverting blood type B to O cells. This paper describes the cloning, sequence, expression, purification, and characterization of recombinant alpha-D-galactosidase. Activity of the recombinant enzyme on the native human erythrocyte blood group B epitope is shown.

Haptoglobin polymorphism among Saharian and West African groups. Haptoglobin phenotype determination by radioimmunoelectrophoresis on Hp O samples.

PubMed Central

Constans, J; Viau, M; Gouaillard, C; Clerc, A

1981-01-01

The haptoglobin (Hp) polymorphism is investigated in 11 African groups living in an area from the Algerian Sahara to Central Africa. More than 4,000 samples were examined. In the Saharian samples, the Hp1 gene frequency is higher than in any other African group. From north to south, a decrease in the Hp1 gene frequency is observed; in the Pygmy sample only, this frequency is lower than the frequency of the Hp2 gene. By means of a sensitive radioimmunoelectrophoresis, the presence of a residual Hp in Hp O sera in which the Hp polymorphism can also be determined can be revealed. Absence of Hp 1-1 and significant excess of Hp 2-2 individuals were observed. More Hp 2-1M phenotypes were detected in the Hp O population than in the non-Hp O population examined. In the Hp O samples, the influence of the phenotype distribution on the Hp gene frequencies is discussed. The heavy polymers of the Hp related to the presence of the alpha 2 chain (Hp2 gene product) are involved only in the biological mechanisms responsible for the presence of Hp O and Hp 2-1 M phenotypes among African groups. Images Fig. 1 PMID:7258189

Ancient roots for polymorphism at the HLA-DQ. alpha. locus in primates

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

Gyllensten, U.B.; Erlich, H.A.

1989-12-01

The genes encoding the human histocompatibility antigens (HLA) exhibit a remarkable degree of polymorphism as revealed by immunologic and molecular analyses. This extensive sequence polymorphism either may have been generated during the lifetime of the human species or could have arisen before speciation and been maintained in the contemporary human population by selection or, possibly, by genetic drift. These two hypotheses were examined using the polymerase chain reaction method to amplify polymorphic sequences from the DQ{alpha} locus, as well as the DX{alpha} locus, an homologous but nonexpressed locus, in a series of primates that diverged at known times. In general,more » the amino acid sequence of a specific human DQ{alpha} allelic type is more closely related to its chimpanzee or gorilla counterpart than to other human DQ{alpha} alleles. Phylogenetic analysis of the silent nucleotide position changes shows that the similarity of allelic types between species is due to common ancestry rather than convergent evolution. Thus, most of the polymorphism at the DQ{alpha} locus in the human species was already present at least 5 million years ago in the ancestral species that gave rise to the chimpanzee, gorilla, and human lineages. However, one of the DQ{alpha} alleles may have arisen after speciation by recombination between two ancestral alleles.« less

Modulating Vascular Hemodynamics With an Alpha Globin Mimetic Peptide (HbαX).

PubMed

Keller, T C Stevenson; Butcher, Joshua T; Broseghini-Filho, Gilson Brás; Marziano, Corina; DeLalio, Leon J; Rogers, Stephen; Ning, Bo; Martin, Jennifer N; Chechova, Sylvia; Cabot, Maya; Shu, Xiahong; Best, Angela K; Good, Miranda E; Simão Padilha, Alessandra; Purdy, Michael; Yeager, Mark; Peirce, Shayn M; Hu, Song; Doctor, Allan; Barrett, Eugene; Le, Thu H; Columbus, Linda; Isakson, Brant E

2016-12-01

The ability of hemoglobin to scavenge the potent vasodilator nitric oxide (NO) in the blood has been well established as a mechanism of vascular tone homeostasis. In endothelial cells, the alpha chain of hemoglobin (hereafter, alpha globin) and endothelial NO synthase form a macromolecular complex, providing a sink for NO directly adjacent to the production source. We have developed an alpha globin mimetic peptide (named HbαX) that displaces endogenous alpha globin and increases bioavailable NO for vasodilation. Here we show that, in vivo, HbαX administration increases capillary oxygenation and blood flow in arterioles acutely and produces a sustained decrease in systolic blood pressure in normal and angiotensin II-induced hypertensive states. HbαX acts with high specificity and affinity to endothelial NO synthase, without toxicity to liver and kidney and no effect on p50 of O 2 binding in red blood cells. In human vasculature, HbαX blunts vasoconstrictive response to cumulative doses of phenylephrine, a potent constricting agent. By binding to endothelial NO synthase and displacing endogenous alpha globin, HbαX modulates important metrics of vascular function, increasing vasodilation and flow in the resistance vasculature. © 2016 American Heart Association, Inc.

Intracellular activation of the fibrinolytic cascade in the Quebec Platelet Disorder.

PubMed

Sheth, Prameet M; Kahr, Walter H A; Haq, M Anwar; Veljkovic, Dragoslava Kika; Rivard, Georges E; Hayward, Catherine P M

2003-08-01

The Quebec Platelet Disorder (QPD) is an unusual bleeding disorder associated with increased platelet stores of urokinase-type plasminogen activator (u-PA) and proteolysis of platelet alpha-granule proteins. The increased u-PA and proteolyzed plasminogen in QPD platelets led us to investigate possible contributions of intracellular plasmin generation to QPD alpha-granule proteolysis. ELISA indicated there were normal amounts of plasminogen and plasmin-alpha(2)-antiplasmin (PAP) complexes in QPD plasmas. Like normal platelets, QPD platelets contained only a small proportion of the blood plasminogen, however, they contained an increased amount of PAP complexes compared to normal platelets (P < 0.005). The quantities of plasminogen stored in platelets were important to induce QPD-like proteolysis of normal alpha-granule proteins by two chain u-PA (tcu-PA) in vitro. Moreover, adding supplemental plasminogen to QPD, but not to control, platelet lysates, triggered further alpha-granule protein proteolysis to forms that comigrated with plasmin degraded proteins. These data suggest the generation of increased but limiting amounts of plasmin within platelets is involved in producing the unique phenotypic changes to alpha-granule proteins in QPD platelets. The QPD is the only known bleeding disorder associated with chronic, intracellular activation of the fibrinolytic cascade.

Coronal X-ray enhancements associated with H-alpha filament disappearances

NASA Technical Reports Server (NTRS)

Webb, D. F.; Krieger, A. S.; Rust, D. M.

1976-01-01

The X-ray telescope experiment aboard the Skylab/ATM mission provided high-resolution soft X-ray images of the lower corona away from active regions, revealing frequent large-scale transient X-ray enhancements which could often be associated with the disappearance of H-alpha filaments. The X-ray emitting structures featured peak brightnesses similar to those of nonflaring active region structures, durations from 3 to 40 hours, shapes that in general outlined the preexisting H-alpha filaments to as large as 400,000 km, apparent expansion velocities of the order of tens of km/sec, and observed peak temperatures of the order of a few million degrees. One such event is described and analyzed in detail. Two explanations of the events are discussed: (1) excess cool material is present in the filament cavity, contributing to the X-ray enhancement, and (2) the enhancement is due to the compression of preexisting material by a changing magnetic field.

Molecular modeling of calmodulin: a comparison with crystallographic data

NASA Technical Reports Server (NTRS)

McDonald, J. J.; Rein, R.

1989-01-01

Two methods of side-chain placement on a modeled protein have been examined. Two molecular models of calmodulin were constructed that differ in the treatment of side chains prior to optimization of the molecule. A virtual bond analysis program developed by Purisima and Scheraga was used to determine the backbone conformation based on 2.2 angstroms resolution C alpha coordinates for the molecules. In the first model, side chains were initially constructed in an extended conformation. In the second model, a conformational grid search technique was employed. Calcium ions were treated explicitly during energy optimization using CHARMM. The models are compared to a recently published refined crystal structure of calmodulin. The results indicate that the initial choices for side-chains, but also significant effects on the main-chain conformation and supersecondary structure. The conformational differences are discussed. Analysis of these and other methods makes possible the formulation of a methodology for more appropriate side-chain placement in modeled proteins.

Dietary medium-chain triglycerides prevent chemically induced experimental colitis in rats.

PubMed

Kono, Hiroshi; Fujii, Hideki; Ishii, Kenichi; Hosomura, Naohiro; Ogiku, Masahito

2010-03-01

The effects of dietary medium-chain triglycerides (MCTs) on experimental colitis induced by 2,4,6-trinitrobenzene sulphonic acid (TNBS) were investigated in rats. Male Wistar rats were given an intracolonic injection of TNBS and were then fed liquid diets containing MCTs or corn oil (AIN93) as controls. Serum and tissue samples were collected 1 week after TNBS enema. The severity of colitis was evaluated pathologically, and tissue myeloperoxidase (MPO) activity was measured. Furthermore, messenger RNA (mRNA) and protein levels for inflammatory cytokines and a chemokine were assessed by reverse-transcription polymerase chain reaction and enzyme-linked immunosorbent assay, respectively. In another set of experiments, the protein expression of Toll-like receptor (TLR)-4 in the colon was measured 1 week after feeding of liquid diets. To investigate the effects of MCTs on macrophages, RAW246.7 macrophages were incubated with media containing albumin conjugated with MCT or linoleic acid, which is the major component of corn oil. Then, the production of tumor necrosis factor-alpha (TNF-alpha) was measured. Dietary MCTs blunted significantly the protein levels of TLR-4 in the colon. Furthermore, the expression of TLR-4 was significantly blunted in RAW264.7 cells incubated with MCTs compared with cells incubated with linoleic acid. Induction of interleukin 1beta (IL-1beta), TNF-alpha, and macrophage inflammatory protein-2 (MIP-2) in the colon was attenuated by dietary MCT. Furthermore, MPO activities in the colonic tissue were significantly blunted in animals fed the MCT diets compared with those fed the control diets. As a result, dietary MCTs improved chemically induced colitis significantly. MCTs most likely are useful for the therapy of inflammatory bowel disease as an anti-inflammatory immunomodulating nutrient. Copyright 2010 Mosby, Inc. All rights reserved.

Structure-activity relationships of the melanocortin tetrapeptide Ac-His-DPhe-Arg-Trp-NH2 at the mouse melanocortin receptors. Part 3: modifications at the Arg position.

PubMed

Holder, Jerry Ryan; Xiang, Zhimin; Bauzo, Rayna M; Haskell-Luevano, Carrie

2003-01-01

The melanocortin pathway is involved in the regulation of several physiological functions including skin pigmentation, steroidogenesis, obesity, energy homeostasis, and exocrine gland function. This melanocortin pathway consists of five known G-protein coupled receptors, endogenous agonists derived from the proopiomelanocortin (POMC) gene transcript, the endogenous antagonists Agouti and the Agouti-related protein (AGRP) and signals through the intracellular cAMP signal transduction pathway. The melanocortin-3 receptor (MC3R) and melanocortin-4 receptor (MC4R) located in the brain are implicated as participating in the metabolic and food intake aspects of energy homeostasis and are stimulated by melanocortin agonists such as alpha-melanocyte stimulation hormone (alpha-MSH). All the endogenous (POMC-derived) melanocortin agonists contain the putative message sequence "His-Phe-Arg-Trp." Herein, we report 12 tetrapeptides, based upon the template Ac-His(6)-DPhe(7)-Arg(8)-Trp(9)-NH(2) (alpha-MSH numbering) that have been modified at the Arg(8) position by neutral, basic, or acidic amino acid side chains. These peptides have been pharmacologically characterized for agonist activity at the mouse melanocortin receptors MC1R, MC3R, MC4R, and MC5R. The most notable results of this study include the observation that removal of the guanidinyl side chain moiety results in decreased melanocortin receptor potency, but that this Arg(8) side chain is not critical for melanocortin receptor agonist activity. Additionally, incorporation of the homoArg(8) residue results in 56-fold MC4R versus MC3R selectivity, and the Orn(8) residue results in 123-fold MC4R versus MC5R and 63-fold MC5R versus MC3R selectivity. Copyright 2002 Elsevier Science Inc.

KM+, a mannose-binding lectin from Artocarpus integrifolia: amino acid sequence, predicted tertiary structure, carbohydrate recognition, and analysis of the beta-prism fold.

PubMed Central

Rosa, J. C.; De Oliveira, P. S.; Garratt, R.; Beltramini, L.; Resing, K.; Roque-Barreira, M. C.; Greene, L. J.

1999-01-01

The complete amino acid sequence of the lectin KM+ from Artocarpus integrifolia (jackfruit), which contains 149 residues/mol, is reported and compared to those of other members of the Moraceae family, particularly that of jacalin, also from jackfruit, with which it shares 52% sequence identity. KM+ presents an acetyl-blocked N-terminus and is not posttranslationally modified by proteolytic cleavage as is the case for jacalin. Rather, it possesses a short, glycine-rich linker that unites the regions homologous to the alpha- and beta-chains of jacalin. The results of homology modeling implicate the linker sequence in sterically impeding rotation of the side chain of Asp141 within the binding site pocket. As a consequence, the aspartic acid is locked into a conformation adequate only for the recognition of equatorial hydroxyl groups on the C4 epimeric center (alpha-D-mannose, alpha-D-glucose, and their derivatives). In contrast, the internal cleavage of the jacalin chain permits free rotation of the homologous aspartic acid, rendering it capable of accepting hydrogen bonds from both possible hydroxyl configurations on C4. We suggest that, together with direct recognition of epimeric hydroxyls and the steric exclusion of disfavored ligands, conformational restriction of the lectin should be considered to be a new mechanism by which selectivity may be built into carbohydrate binding sites. Jacalin and KM+ adopt the beta-prism fold already observed in two unrelated protein families. Despite presenting little or no sequence similarity, an analysis of the beta-prism reveals a canonical feature repeatedly present in all such structures, which is based on six largely hydrophobic residues within a beta-hairpin containing two classic-type beta-bulges. We suggest the term beta-prism motif to describe this feature. PMID:10210179

Antimycobacterial action of thiolactomycin: an inhibitor of fatty acid and mycolic acid synthesis.

PubMed Central

Slayden, R A; Lee, R E; Armour, J W; Cooper, A M; Orme, I M; Brennan, P J; Besra, G S

1996-01-01

Thiolactomycin (TLM) possesses in vivo antimycobacterial activity against the saprophytic strain Mycobacterium smegmatis mc2155 and the virulent strain M. tuberculosis Erdman, resulting in complete inhibition of growth on solid media at 75 and 25 micrograms/ml, respectively. Use of an in vitro murine macrophage model also demonstrated the killing of viable intracellular M. tuberculosis in a dose-dependent manner. Through the use of in vivo [1,2-14C]acetate labeling of M. smegmatis, TLM was shown to inhibit the synthesis of both fatty acids and mycolic acids. However, synthesis of the shorter-chain alpha'-mycolates of M. smegmatis was not inhibited by TLM, whereas synthesis of the characteristic longer-chain alpha-mycolates and epoxymycolates was almost completely inhibited at 75 micrograms/ml. The use of M. smegmatis cell extracts demonstrated that TLM specifically inhibited the mycobacterial acyl carrier protein-dependent type II fatty acid synthase (FAS-II) but not the multifunctional type I fatty acid synthase (FAS-I). In addition, selective inhibition of long-chain mycolate synthesis by TLM was demonstrated in a dose-response manner in purified, cell wall-containing extracts of M. smegmatis cells. The in vivo and in vitro data and knowledge of the mechanism of TLM resistance in Escherichia coli suggest that two distinct TLM targets exist in mycobacteria, the beta-ketoacyl-acyl carrier protein synthases involved in FAS-II and the elongation steps leading to the synthesis of the alpha-mycolates and oxygenated mycolates. The efficacy of TLM against M. smegmatis and M. tuberculosis provides the prospects of identifying fatty acid and mycolic acid biosynthetic genes and revealing a novel range of chemotherapeutic agents directed against M. tuberculosis. PMID:9124847

Inhibition of hair follicle growth by a laminin-1 G-domain peptide, RKRLQVQLSIRT, in an organ culture of isolated vibrissa rudiment.

PubMed

Hayashi, Kazuhiro; Mochizuki, Mayumi; Nomizu, Motoyoshi; Uchinuma, Eijyu; Yamashina, Shohei; Kadoya, Yuichi

2002-04-01

We established a serum-free organ culture system of isolated single vibrissa rudiments taken from embryonic day 13 mice. This system allowed us to test more than 30 laminin-derived cell adhesive peptides to determine their roles on the growth and differentiation of vibrissa hair follicles. We found that the RKRLQVQLSIRT sequence (designated AG-73), which mapped to the LG-4 module of the laminin-alpha1 chain carboxyl-terminal G domain, perturbed the growth of hair follicles in vitro. AG-73 is one of the cell-binding peptides identified from more than 600 systematically synthesized 12 amino acid peptides covering the whole amino acid sequence of the laminin-alpha1, -beta1, and -gamma1 chains, by cell adhesion assay. Other cell-adhesive laminin peptides and a control scrambled peptide, LQQRRSVLRTKI, however, failed to show any significant effects on the growth of hair follicles. The AG-73 peptide binds to syndecan-1, a transmembrane heparan-sulfate proteoglycan. Syndecan-1 was expressed in both the mesenchymal condensation and the epithelial hair peg of developing vibrissa, suggesting that AG-73 binding to the cell surface syndecan-1 perturbed the epithelial-mesenchymal interactions of developing vibrissa. The formation of hair bulbs was aberrant in the explants treated with AG-73. In addition, impaired basement membrane formation, an abnormal cytoplasmic bleb formation, and an unusual basal formation of actin bundles were noted in the AG-73-treated-hair matrix epithelium, indicating that AG-73 binding perturbs various steps of epithelial morphogenesis, including the basement membrane remodeling. We also found a region-specific loss of the laminin-alpha1 chain in the basement membrane at the distal region of the invading hair follicle epithelium, indicating that laminins play a part in hair morphogenesis.

Preparation of fluorescent tocopherols for use in protein binding and localization with the alpha-tocopherol transfer protein.

PubMed

Nava, Phillip; Cecchini, Matt; Chirico, Sara; Gordon, Heather; Morley, Samantha; Manor, Danny; Atkinson, Jeffrey

2006-06-01

Sixteen fluorescent analogues of the lipid-soluble antioxidant vitamin alpha-tocopherol were prepared incorporating fluorophores at the terminus of omega-functionalized 2-n-alkyl-substituted chromanols (1a-d and 4a-d) that match the methylation pattern of alpha-tocopherol, the most biologically active form of vitamin E. The fluorophores used include 9-anthroyloxy (AO), 7-nitrobenz-2-oxa-1,3-diazole (NBD), N-methyl anthranilamide (NMA), and dansyl (DAN). The compounds were designed to function as fluorescent reporter ligands for protein-binding and lipid transfer assays. The fluorophores were chosen to maximize the fluorescence changes observed upon moving from an aqueous environment (low fluorescence intensity) to an hydrophobic environment such as a protein's binding site (high fluorescence intensity). Compounds 9d (anthroyloxy) and 10d (nitrobenzoxadiazole), having a C9-carbon chain between the chromanol and the fluorophore, were shown to bind specifically and reversibly to recombinant human tocopherol transfer protein (alpha-TTP) with dissociation constants of approximately 280 and 60 nM, respectively, as compared to 25 nM for the natural ligand 2R,4'R,8'R-alpha-tocopherol. Thus, compounds have been prepared that allow the investigation of the rate of alpha-TTP-mediated inter-membrane transfer of alpha-tocopherol and to investigate the mechanism of alpha-TTP function at membranes of different composition.

Structural re-alignment in an immunologic surface region of ricin A chain

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

Zemla, A T; Zhou, C E

2007-07-24

We compared structure alignments generated by several protein structure comparison programs to determine whether existing methods would satisfactorily align residues at a highly conserved position within an immunogenic loop in ribosome inactivating proteins (RIPs). Using default settings, structure alignments generated by several programs (CE, DaliLite, FATCAT, LGA, MAMMOTH, MATRAS, SHEBA, SSM) failed to align the respective conserved residues, although LGA reported correct residue-residue (R-R) correspondences when the beta-carbon (Cb) position was used as the point of reference in the alignment calculations. Further tests using variable points of reference indicated that points distal from the beta carbon along a vector connectingmore » the alpha and beta carbons yielded rigid structural alignments in which residues known to be highly conserved in RIPs were reported as corresponding residues in structural comparisons between ricin A chain, abrin-A, and other RIPs. Results suggest that approaches to structure alignment employing alternate point representations corresponding to side chain position may yield structure alignments that are more consistent with observed conservation of functional surface residues than do standard alignment programs, which apply uniform criteria for alignment (i.e., alpha carbon (Ca) as point of reference) along the entirety of the peptide chain. We present the results of tests that suggest the utility of allowing user-specified points of reference in generating alternate structural alignments, and we present a web server for automatically generating such alignments.« less

Evolution and Function of the TCR Vgamma9 Chain Repertoire: It’s Good to be Public

PubMed Central

Pauza, C. David; Cairo, Cristiana

2015-01-01

Lymphocytes expressing a T cell receptor (TCR) composed of Vgamma9 and Vdelta2 chains represent a minor fraction of human thymocytes. Extrathymic selection throughout post-natal life causes the proportion of cells with a Vgamma9-JP rearrangement to increase and elevates the capacity for responding to non-peptidic phosphoantigens. Extrathymic selection is so powerful that phosphoantigen-reactive cells comprise about 1 in 40 circulating memory T cells from healthy adults and the subset can be expanded rapidly upon infection or in response to malignancy. Skewing of the gamma delta TCR repertoire is accompanied by selection for public gamma chain sequences such that many unrelated individuals overlap extensive in their circulating repertoire. This type of selection implies the presence of a monomorphic antigen-presenting molecule that is an object of current research but remains incompletely defined. While selection on a monomorphic presenting molecule may seem unusual, similar mechanisms shape the alpha beta T cell repertoire including the extreme examples of NKT or mucosal-associated invariant T cells (MAIT) and the less dramatic amplification of public Vbeta chain rearrangements driven by individual MHC molecules and associated with resistance to viral pathogens. Selecting and amplifying public T cell receptors whether alpha beta or gamma delta, are important steps in developing an anticipatory TCR repertoire. Cell clones expressing public TCR can accelerate the kinetics of response to pathogens and impact host survival. PMID:25769734

Regulation of acetylcholine receptor alpha subunit variants in human myasthenia gravis. Quantification of steady-state levels of messenger RNA in muscle biopsy using the polymerase chain reaction.

PubMed Central

Guyon, T; Levasseur, P; Truffault, F; Cottin, C; Gaud, C; Berrih-Aknin, S

1994-01-01

Myasthenia gravis (MG) is an autoimmune disease mediated by auto-antibodies that attack the nicotinic acetylcholine receptor (AChR). To elucidate the molecular mechanisms underlying the decrease in AChR levels at the neuromuscular junction, we investigated the regulation of AChR expression by analyzing mRNA of the two AChR alpha subunit isoforms (P3A+ and P3A-) in muscle samples from myasthenic patients relative to controls. We applied a quantitative method based on reverse transcription of total RNA followed by polymerase chain reaction (PCR), using an internal standard we constructed by site-directed mutagenesis. An increased expression of mRNA coding for the alpha subunit of the AChR isoforms was observed in severely affected patients (P < 0.003 versus controls) but not in moderately affected patients, independently of the anti-AChR antibody titer. Study of mRNA precursor levels indicates a higher expression in severely affected patients compared to controls, suggesting an enhanced rate of transcription of the message coding for the alpha subunit isoforms in these patients. We have also reported that mRNA encoding both isoforms are expressed at an approximate 1:1 ratio in controls and in patients. We have thus identified a new biological parameter correlated with disease severity, and provide evidence of a compensatory mechanism to balance the loss of AChR in human myasthenia gravis, which is probably triggered only above a certain degree of AChR loss. Images PMID:8040257

Proteolytic processing of the pro beta chain of beta-hexosaminidase occurs at basic residues contained within an exposed disulfide loop structure.

PubMed

Sagherian, C; Poroszlay, S; Vavougios, G; Mahuran, D

1993-01-01

Lysosomal beta-hexosaminidase (EC 3.2.1.52) occurs as two major isozymes, Hex A (alpha beta) and Hex B (beta beta). The alpha and beta subunits are encoded by the HEXA and HEXB genes, respectively. Extensive homology in both the gene structures and deduced primary sequences demonstrate their common evolutionary origin. While undergoing similar proteolytic modifications in the lysosome, the pro beta polypeptide is additionally cleaved internally to produce the mature 24-30 kilodalton beta b and beta a chains. Previous data have suggested that this processing event occurs somewhere between residues Ser311 and Lys315. In this report we demonstrate that this area is located in a hydrophilic disulfide-loop structure (between Cys309 and Cys360). The cleavage event is prevented by the deletion through in vitro mutagenesis of the Arg312-Gln-Asn-Lys tetrapeptide or by its substitution with the aligned alpha residues (Gly-Ser-Glu-Pro). Reintroduction of either Arg312 or Lys315 reinstates the processing. Furthermore, we show that this area is not involved in lysosomal targeting of pro-Hex B, or in the increased stability or the variation in substrate specificity of the beta as compared with the alpha subunit. Our data suggest the presence of a novel lysosomal endoprotease. Like other endoproteases it is specific for basic amino acids; however, it cleaves on the amino-terminal side rather than the conventional carboxy-terminal side of such residues and then only if they are fully exposed to the lysosomal environment.(ABSTRACT TRUNCATED AT 250 WORDS)

Influence of major structural features of tocopherols and tocotrienols on their omega-oxidation by tocopherol-omega-hydroxylase.

PubMed

Sontag, Timothy J; Parker, Robert S

2007-05-01

Human cytochrome P450 4F2 (CYP4F2) catalyzes the initial omega-hydroxylation reaction in the metabolism of tocopherols and tocotrienols to carboxychromanols and is, to date, the only enzyme shown to metabolize vitamin E. The objective of this study was to characterize this activity, particularly the influence of key features of tocochromanol substrate structure. The influence of the number and positions of methyl groups on the chromanol ring, and of stereochemistry and saturation of the side chain, were explored using HepG2 cultures and microsomal reaction systems. Human liver microsomes and microsomes selectively expressing recombinant human CYP4F2 exhibited substrate activity patterns similar to those of HepG2 cells. Although activity was strongly associated with substrate accumulation by cells or microsomes, substantial differences in specific activities between substrates remained under conditions of similar microsomal membrane substrate concentration. Methylation at C5 of the chromanol ring was associated with markedly low activity. Tocotrienols exhibited much higher Vmax values than their tocopherol counterparts. Side chain stereochemistry had no effect on omega-hydroxylation of alpha-tocopherol (alpha-TOH) by any system. Kinetic analysis of microsomal CYP4F2 activity revealed Michaelis-Menten kinetics for alpha-TOH but allosteric cooperativity for other vitamers, especially tocotrienols. Additionally, alpha-TOH was a positive effector of omega-hydroxylation of other vitamers. These results indicate that CYP4F2-mediated tocopherol-omega-hydroxylation is a central feature underlying the different biological half-lives, and therefore biopotencies, of the tocopherols and tocotrienols.

Removing peroxide impurities from ether improves the quality of globin chains for biosynthetic studies.

PubMed

Mirzazadeh, Roghieh; Khatami, Shohreh; Bayat, Parastoo; Zamani, Zahra; Sadeghi, Sedigheh; Roohi, Soghra; Saidi, Parinaz

2005-01-01

The diagnosis of the different forms of thalassemia is one of the important applications of analysis of globin chains. These analyses are done by high performance liquid chromatography (HPLC) using a MONO-S cation exchange column and ether is used for washing the globin powder in the final step. The presence of peroxide impurities in ether could change the structure of the globin chains. In the chromatograms, these modified globins appear as separated peaks next to the unmodified globin peaks. In these cases, the alpha/beta ratio exceed by artifact the correct value. Our study demonstrates that diagnostic centers should ensure that the ether they use is pure.

Partial rescue of glomerular laminin alpha5 mutations by wild-type endothelia produce hybrid glomeruli.

PubMed

Abrahamson, Dale R; St John, Patricia L; Isom, Kathryn; Robert, Barry; Miner, Jeffrey H

2007-08-01

Both endothelial cells and podocytes are sources for laminin alpha1 at the inception of glomerulogenesis and then for laminin alpha5 during glomerular maturation. Why glomerular basement membranes (GBM) undergo laminin transitions is unknown, but this may dictate glomerular morphogenesis. In mice that genetically lack laminin alpha5, laminin alpha5beta2gamma1 is not assembled, vascularized glomeruli fail to form, and animals die at midgestation with neural tube closure and placental deficits. It was previously shown that renal cortices of newborn mice contain endothelial progenitors (angioblasts) and that when embryonic day 12 kidneys are transplanted into newborn kidney, hybrid glomeruli (host-derived endothelium and donor-derived podocytes) result. Reasoning that host endothelium may correct the glomerular phenotype that is seen in laminin alpha5 mutants, alpha5 null embryonic day 12 metanephroi were grafted into wild-type newborn kidney. Hybrid glomeruli were identified in grafts by expression of a host-specific LacZ lineage marker. Labeling of glomerular hybrid GBM with chain-specific antibodies showed a markedly stratified distribution of laminins: alpha5 was found only on the inner endothelial half of GBM, whereas alpha1 located to outer layers beneath mutant podocytes. For measurement of the contribution of host endothelium to hybrid GBM, immunofluorescent signals for laminin alpha5 were quantified: Hybrid GBM contained approximately 50% the normal alpha5 complement as wild-type GBM. Electron microscopy of glomerular hybrids showed vascularization, but podocyte foot processes were absent. It was concluded that (1) endothelial and podocyte-derived laminins remain tethered to their cellular origin, (2) developing endothelial cells contribute large amounts of GBM laminins, and (3) podocyte foot process differentiation may require direct exposure to laminin alpha5.

Identification of high-mannose and multiantennary complex-type N-linked glycans containing alpha-galactose epitopes from Nurse shark IgM heavy chain.

PubMed

Harvey, David J; Crispin, Max; Moffatt, Beryl E; Smith, Sylvia L; Sim, Robert B; Rudd, Pauline M; Dwek, Raymond A

2009-11-01

MALDI-TOF mass spectrometry, negative ion nano-electrospray MS/MS and exoglycosidase digestion were used to identify 36 N-linked glycans from 19S IgM heavy chain derived from the nurse shark (Ginglymostoma cirratum). The major glycan was the high-mannose compound, Man(6)GlcNAc(2) accompanied by small amounts of Man(5)GlcNAc(2), Man(7)GlcNAc(2) and Man(8)GlcNAc(2). Bi- and tri-antennary (isomer with a branched 3-antenna) complex-type glycans were also abundant, most contained a bisecting GlcNAc residue (beta1-->4-linked to the central mannose) and with varying numbers of alpha-galactose residues capping the antennae. Small amounts of monosialylated glycans were also found. This appears to be the first comprehensive study of glycosylation in this species of animal. The glycosylation pattern has implications for the mechanism of activation of the complement system by nurse shark IgM.

Nonionic diethanolamide amphiphiles with isoprenoid-type hydrocarbon chains: thermotropic and lyotropic liquid crystalline phase behaviour

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

Sagnella, Sharon M.; Conn, Charlotte E.; Krodkiewska, Irena

2014-09-24

The thermotropic and lyotropic liquid crystalline phase behaviour of a series of diethanolamide amphiphiles with isoprenoid-type hydrocarbon chains (geranoyl, H-farnesoyl, and phytanoyl) has been investigated. When neat, both H-farnesoyl and phytanoyl diethanolamide form a smectic liquid crystalline structure at sub-zero temperatures. In addition, all three diethanolamides exhibit a glass transition temperature at around -73 C. Geranoyl diethanolamide forms a lamellar crystalline phase with a lattice parameter of 17.4 {angstrom} following long term storage accompanied by the loss of the glass transition. In the presence of water, H-farnesoyl and phytanoyl diethanolamide form lyotropic liquid crystalline phases, whilst geranoyl diethanolamide forms anmore » L{sub 2} phase. H-farnesoyl diethanolamide forms a fluid lamellar phase (L{sub {alpha}}) at room temperature and up to {approx} 40 C. Phytanoyl diethanolamide displays a rich mesomorphism forming the inverse diamond (Q{sub II}{sup D}) and gyroid (Q{sub II}{sup G}) bicontinuous cubic phases in addition to an L{sub {alpha}} phase.« less

Pregnancy complicated with Alport syndrome: a good obstetric outcome and failure to diagnose an infant born to a mother with Alport syndrome by umbilical cord immunofluorescence staining.

PubMed

Matsubara, Shigeki; Ueda, Yoshihiko; Takahashi, Hisako; Nagai, Takashi; Kuwata, Tomoyuki; Muto, Shigeaki; Yamaguchi, Takehiko; Takizawa, Toshihiro; Suzuki, Mitsuaki

2009-12-01

Alport syndrome is a familial progressive nephritis. The most frequent type is X-linked Alport syndrome, caused by genetic abnormalities in the alpha 5 chain of type IV collagen. Skin biopsy is a useful tool for diagnosing this disease. It is not well known how this syndrome affects pregnancy and how it is affected by pregnancy, or whether the umbilical cord may provide material for detecting this collagen abnormality. We report a primigravida with Alport syndrome with mild proteinuria who gave birth abdominally to a term male infant without deteriorating renal function during pregnancy. The umbilical cord from not only this infant but also from an Alport (-) control infant showed negative immunofluorescence staining for the alpha 5 chain of type IV collagen. Women with Alport syndrome without renal dysfunction may follow an uneventful obstetrical course until term. The cord may not be suitable for diagnosing Alport syndrome with immunofluorescence staining.

PG 0308 + 096 and PG 1026 + 002 - Two new short period binary stars resulting from common-envelope evolution

NASA Technical Reports Server (NTRS)

Saffer, Rex A.; Wade, Richard A.; Liebert, James; Green, Richard F.; Sion, Edward M.; Bechtold, J.; Foss, Diana; Kidder, K.

1993-01-01

Ultraviolet spectroscopy, optical spectroscopy, and spectrophotometry have been used to study the excess UV stars PG 0308 + 096 and PG 1026 + 002. Both objects are short-period binary systems, each containing a DA white dwarf star and a dM star. Orbital periods of approximately 0.284 day for PG 0308 + 096, and aproximately 0.597 day for PG 1026, have been found by spectroscopic analysis of the H-alpha emission line. Ly-alpha and Balmer line profile fitting were used to estimate the mass of white dwarf stars; mass estimates for the dM stars are based on their spectral types. The orbital inclinations are derived from these masses, the periods, and amplitudes of the H-alpha radial velocity curves. The equivalent width of the H-alpha emission line, in each binary system, varies with the orbital phase in such a manner as to imply that it arises, in large part at least, from the hemisphere of the M star that faces the white dwarf star.

Determination of (alpha)-dialkylamino acids and their Enantiomers in Geological Samples by High-Performance Liquid Chromatography after Dervatization with a Chiral Adduct of (omicron)-Phthaldialdehyde

NASA Technical Reports Server (NTRS)

Zhoa, Meixun; Bada, Jeffrey L.

1995-01-01

Derivatization with (omicron)-phthaldialdehyde (OPA) and the chiral thiol N-acetyl-L-cysteine (NAC) is a convenient and sensitive technique for the HPLC detection and resolution of protein amino acid enantiomers. The kinetics of the reaction of OPA-NAC with (alpha)-dialkylamino acids was investigated. The fluorescence yield of (alpha)-dialkylamino acids was only about 10% of that of protein amino acids when the derivatization was carried out at room temperature for 1-2 min, which is the procedure generally used for protein amino acid analyses. The fluorescence yield of (alpha)-dialkylamino acids can be enhanced by up to ten-fold when the derivatization reaction time is increased to 15 min at room temperature. The OPA-NAC technique was optimized for the detection and enantiomeric resolution of a-dialkylamino acids in geological samples which contain a large excess of protein amino acids. The estimated detection limit for a-dialkylamino acids is 1-2 pmol, comparable to that for protein amino acids.

Molecular structure of the pyruvate dehydrogenase complex from Escherichia coli K-12.

PubMed

Vogel, O; Hoehn, B; Henning, U

1972-06-01

The pyruvate dehydrogenase core complex from E. coli K-12, defined as the multienzyme complex that can be obtained with a unique polypeptide chain composition, has a molecular weight of 3.75 x 10(6). All results obtained agree with the following numerology. The core complex consists of 48 polypeptide chains. There are 16 chains (molecular weight = 100,000) of the pyruvate dehydrogenase component, 16 chains (molecular weight = 80,000) of the dihydrolipoamide dehydrogenase component, and 16 chains (molecular weight = 56,000) of the dihydrolipoamide dehydrogenase component. Usually, but not always, pyruvate dehydrogenase complex is produced in vivo containing at least 2-3 mol more of dimers of the pyruvate dehydrogenase component than the stoichiometric ratio with respect to the core complex. This "excess" component is bound differently than are the eight dimers in the core complex.

Evaluation of strategies to control Fab light chain dimer during mammalian expression and purification: A universal one-step process for purification of correctly assembled Fab.

PubMed

Spooner, Jennifer; Keen, Jenny; Nayyar, Kalpana; Birkett, Neil; Bond, Nicholas; Bannister, David; Tigue, Natalie; Higazi, Daniel; Kemp, Benjamin; Vaughan, Tristan; Kippen, Alistair; Buchanan, Andrew

2015-07-01

Fabs are an important class of antibody fragment as both research reagents and therapeutic agents. There are a plethora of methods described for their recombinant expression and purification. However, these do not address the issue of excessive light chain production that forms light chain dimers nor do they describe a universal purification strategy. Light chain dimer impurities and the absence of a universal Fab purification strategy present persistent challenges for biotechnology applications using Fabs, particularly around the need for bespoke purification strategies. This study describes methods to address light chain dimer formation during Fab expression and identifies a novel CH 1 affinity resin as a simple and efficient one-step purification for correctly assembled Fab. © 2015 Wiley Periodicals, Inc.

Transepithelial transport of alpha-lipoic acid across human intestinal Caco-2 cell monolayers.

PubMed

Takaishi, Naoki; Yoshida, Kazutaka; Satsu, Hideo; Shimizu, Makoto

2007-06-27

Alpha-lipoic acid (LA) is used in dietary supplements or food with antioxidative functions. The mechanism for the intestinal absorption of alpha-lipoic acid was investigated in this study by using human intestinal Caco-2 cell monolayers. LA was rapidly transported across the Caco-2 cell monolayers, this transport being energy-dependent, suggesting transporter-mediated transport to be the mechanism involved. The LA transport was strongly dependent on the pH value, being accelerated in the acidic pH range. Furthermore, such monocarboxylic acids as benzoic acid and medium-chain fatty acids significantly inhibited LA transport, suggesting that a proton-linked monocarboxylic acid transporter (MCT) was involved in the intestinal transport of LA. The conversion of LA to the more antioxidative dihydrolipoic acid was also apparent during the transport process.

Purification of alpha-glucosidae and invertase from bakers' yeast on modified polymeric supports.

PubMed

Lothe, R R; Purohit, S S; Shaikh, S S; Malshe, V C; Pandit, A B

1999-01-01

In the present work Amberlite XAD-16 and Indion NPA-1, Polystyrene Divinylbenzene macroreticular spherical resins, have been evaluated quantitatively as supports for the adsorption and isolation of the yeast proteins and the enzymes, invertase and alpha-glucosidase. Modification of these supports has been carried out by surface grafting using acrylate polymers to reduce the hydrophobicity and nonspecific adsorption of proteins. Good grafting efficiency, in excess of 90%, has been obtained using ultrasonic irradiation for the surface activation of polystyrene resins. XAD-16 has higher adsorption capacities for the total yeast proteins as well as for both the enzymes, alpha-glucosidase and invertase, than NPA-1 in its respective native and grafted form. Adsorption capacities of XAD-16 and NPA-1 in their respective native and grafted forms for alpha-glucosidase are higher than the capacities for invertase. Nonspecific adsorption of total proteins has been reduced considerably after the grafting of acrylate polymers on hydrophobic supports. At the same time selectivity for the adsorption of both the enzymes has been enhanced on grafted supports. The overall solid-liquid adsorption mass transfer coefficient values (Kla) estimated for adsorption of invertase on XAD are lower than those for alpha-glucosidase. Native and grafted resins could be regenerated and reused for adsorption of alpha-glucosidase for two regeneration cycles studied. Storage stability of invertase and alpha-glucosidase is the same on native and grafted form of XAD-16 and is more than the enzymes in the free form.

Dithiothreitol-based protein equalization technology to unravel biomarkers for bladder cancer.

PubMed

Araújo, J E; López-Fernández, H; Diniz, M S; Baltazar, Pedro M; Pinheiro, Luís Campos; da Silva, Fernando Calais; Carrascal, Mylène; Videira, Paula; Santos, H M; Capelo, J L

2018-04-01

This study aimed to assess the benefits of dithiothreitol (DTT)-based sample treatment for protein equalization to assess potential biomarkers for bladder cancer. The proteome of plasma samples of patients with bladder carcinoma, patients with lower urinary tract symptoms (LUTS) and healthy volunteers, was equalized with dithiothreitol (DTT) and compared. The equalized proteomes were interrogated using two-dimensional gel electrophoresis and matrix assisted laser desorption ionization time of flight mass spectrometry. Six proteins, namely serum albumin, gelsolin, fibrinogen gamma chain, Ig alpha-1 chain C region, Ig alpha-2 chain C region and haptoglobin, were found dysregulated in at least 70% of bladder cancer patients when compared with a pool of healthy individuals. One protein, serum albumin, was found overexpressed in 70% of the patients when the equalized proteome of the healthy pool was compared with the equalized proteome of the LUTS patients. The pathways modified by the proteins differentially expressed were analyzed using Cytoscape. The method here presented is fast, cheap, of easy application and it matches the analytical minimalism rules as outlined by Halls. Orthogonal validation was done using western-blot. Overall, DTT-based protein equalization is a promising methodology in bladder cancer research. Copyright © 2017 Elsevier B.V. All rights reserved.

16 CFR 801.90 - Transactions or devices for avoidance.

Code of Federal Regulations, 2010 CFR

2010-01-01

... observe the waiting period. 2. Suppose “A” wholly owns and operates a chain of twenty retail hardware... fair market value of the assets of the twenty store corporations is in excess of $50 million (as...