bluuebunny/crossref_metadata_2025_split huggingface dataset:
Cloning, expression and purification of the coat protein of encephalitis virus (DIEV) infecting Dicentrarchus labrax
Authors: Sideris Diamantis C. 
Published: July 2007

AbstractThe coat protein gene from encephalitis virus infecting Dicentrarhuslabrax (DIEV) has been cloned by gene amplification, sequenced and expressed in Escherichia coli. DNA sequencing has revealed an open reading frame of 1017 bases encoding a polypeptide of 338 amino acids. The sequence similarities between the DIEV coat protein gene and the same gene in five encephalitis viruses infected other fish species were over 71.5% at the nucleotide level and over 79.5% at the amino acid level. These results indicate that the nodaviruses that cause encephalopathy and retinopathy in fishes are very closed related. E. coli cells harbouring the plasmid containing the DIEV gene can produce the viral coat protein. An efficient purification scheme using a Sepharore‐Ni+2 column is presented. This, gives approx. 10 mg of more than 95% pure protein per gr of E. coli culture.
Recombinant CTFR detection in CF tracheal epithelial cells following in vitro liposomeme‐mediated gene transfer
Authors: Colosimo Alessia , Scarpino Stefania , Sangiuolo Federica , Sario Sabrina Di , Mossa Giuseppe , Novelli Giuseppe , Dallapiccola Bruno 
Published: July 2007

AbstractThe efficacy of CFTR gene transfer mediated by cationic liposomes Dc‐Chol/DOPE into cystic fibrosis (CF) tracheal epithelial cells carrying defective processing mutations (S549N/N1303K), was assessed by studying mRNA and protein expression of the recombinant product. Appreciable levels of mRNA transcripts were detected 48 h after transfection, while complete translocation of the recombinant CFTR to the apical membrane of epithelial cells was observed after 72 h following transfection. Our results suggest that in vitro restoration of a normal CFTR processing and migration to the cell plasmalemma requires 72 h at least as demonstrated by immunocyto‐fluorescence using the monoclonal antibody MATG 1016. These findings are relevant onto gene transfer phase I clinical studies.
Efficient production of active TNF‐α by albumin signal peptide
Authors: Maeda Yuu , Soda Mariko , Ito Keizo , Sato Kenzo 
Published: July 2007

AbstractTNF‐α is initially synthesized as a membrane‐anchored precursor protein and processed proteolytically by a matrix metalloproteinase (MMP)‐like enzyme. In order to establish an efficient expression system of TNF‐α in mammalian cells without involvement of the extracellular enzyme, an expression plasmid (pCN‐alb‐TNF) was constructed with a signal sequence of the rat albumin gene as a module for secretion. The highest level of production of TNF‐α was observed in the clone CT‐3 by SDS‐PAGE and Western blot analysis. Biological activity of the secretion was revealed by repression of catalase gene expression in hepatoma cells and cytotoxity to L929 cells. Attachment of the signal peptide to mature form resulted in the enhancement of production of the cytokine.
Isoform‐specific monoclonal antibodies against HSP90
Authors: Nemoto Takayuki , Roi Rina , Matsusaka Tomonori , Iwanari Hiroko , Yamashita Hisahiko , Kyakumoto Seiko , Sato Nobuko 
Published: July 2007

AbstractThe purpose of this study was to develop monoclonal antibodies (mAbs) that distinguish between the two isoforms of human 90‐kDa heat shock protein (HSP90), i.e HSP90α and β. Human HSP90α and β isoforms expressed in Escherichia coil were separately used as antigens for developing the mAbs. Twenty‐three and ten mAbs were obtained by immunization of mice with HSP90α and β, respectively. Among them, ten and three mAbs specifically recognized HSP90α and β isoforms, respectively, on the criteria of both enzyme‐linked immunosorbent and immunoblotting analyses. Immunochemical analysis by use of the mAbs revealed that both of the HSP90 isoforms were present in human cells even under unstressed conditions and that the expression of HSP90α was more strongly induced when the cells were exposed to arsenate. This is the first report of the development of the mAbs discriminating between the two isoforms of HSP90. The mAbs specific for HSP90 isoforms should be useful for the regulational and functional analyses of HSP90 isoforms.
A recombinant human fibronectin fragment facilitates retroviral mediated gene transfer into human hematopoietic progenitor cells
Authors: Dutt Parmesh , Hanenberg Helmut , Vik Terry , Williams David A. , Yoder Mervin C. 
Published: July 2007

AbstractGene transfer into hematopoietic progenitor cells is increased when these cells are adherent to the carboxy‐terminal chymotryptic fragment of human plasma fibronectin (FN30/35) containing the heparin binding domain (HBD) and the alternatively spliced type three connecting segment (IIICS) region. We report herein the production of a recombinant human fibronectin fragment comprised of the HBD and IIICS regions expressed by Sf9 insect cells following recombinant baculovirus infection. The resulting isolated peptide, HuBacFN, facilitated gene transfer into human hematopoietic cell lines and primary human hematopoietic progenitors to a level achieved with the purified human plasma FN30/35 peptide.
Nonenzymatic reduction of brucine N‐oxide by the heme group of cytochrome P450
Authors: Takekawa Koji , Sugihara Kazumi , Kitamura Shigeyuki , Tatsumi Kiyoshi 
Published: July 2007

AbstractEvidence showing that cytochrome P450‐mediated reduction of brucine N‐oxide to brucine by rat liver microsomes proceeds nonenzymatically in the presence of both a reduced pyridine nucleotide and FAD is presented. The microsomal N‐oxide reduction appears to proceed in two steps: The first step is reduction of FAD by NADPH or NADH either enzymatically or nonenzymatically. The second step is nonenzymatic reduction of the tertiary amine N‐oxide by the reduced flavin and is nonenzymatically catalyzed by the heme group of cytochrome P450.
Effects of vitamin E and selenium on antioxidant defense in rat heart
Authors: Scholz R. W. , Minicucci L. A. , Reddy C. C. 
Published: July 2007

AbstractHeart mitochondria, isolated from rats fed diets deficient or supplemented with vitamin E (E) and/or selenium (Se), were subjected to time‐course assays of lipid peroxidation stimulated by ascorbate/ADP/Fe3+. Mitochondria depleted of α‐tocopherol (α‐TH) peroxided rapidly as assessed by formation of thiobarbituric acid reactive substances (TBARS). Formation of TBARS was strongly inhibited in mitochondria from rats fed diets supplemented with E. Selenium deficiency, reduced glutathione (GSH), glutathione disulfide (GSSG) or GSH + GSSG did not affect the course of lipid peroxidation in mitochondria from rats supplemented or deficient in E. Combined E and Se deficiency resulted in significantly lower total (oxidized + reduced) mitochondrial coenzyme Q‐9 (CoQ‐9) concentration compared with control rats supplemented with dietary E and Se. Time‐course changes in mitochondrial α‐TH and total CoQ‐9 during oxidizing conditions were minor in +E rats. Total CoQ‐9 was reduced substantially, however, during the course of lipid peroxidation in mitochondria depleted of α‐TH. Selenium‐dependent glutathione peroxidase (Se‐GSHPx) activity was reduced by approximately 96% in heart cytosol, and to a somewhat lesser extent in mitochondria, by dietary Se deficiency. Non‐Se GSHPx activity was not detected in heart cytosol but was detected in very small amounts in heart mitochondria. Glutathione S‐transferase activity of heart cytosol was decreased in E and/or Se deficiency. The results of these experiments indicate that membrane α‐TH was most effective in inhibiting lipid peroxidation in heart mitochondria.
Purification and biochemical characterisation of human placental acid α‐glucosidase
Authors: Chadalavada Durga M. , Sivakami S. 
Published: July 2007

AbstractAn acid α‐glucosidase (EC 3.2.1.3) has been purified to electrophoretic homogeneity from the soluble fraction of the human term placenta. In the presence of SDS, two doublets of 79 and 67 kDa were seen in addition to other bands of extremely low intensity. Each of these bands was seen to cross‐react with polyclonal antiserum raised to the purified enzyme, thus confirming the homogeneity of the preparation. The purified enzyme exhibited a broad substrate specificity. The kinetic data revealed the possible presence of multiple substrate binding sites. Chemical modification using group specific reagents indicated the presence of a carboxyl group and tryptophan at the active site. Based on these results a possible structure for the active site of the human term placental acid α‐glucosidase has been proposed.
S and G2 phase histone biosynthesis of HEp‐2 cells after the influence of the bisalkylating agent, chlorambucil
Authors: Sourlingas Thomae G. , Sekeri‐Pataryas Kalliope E. 
Published: July 2007

AbstractChlorambucil was previously found to be a specific inhibitor of total histone synthesis without affecting total cellular protein synthesis. In this study, we used S and G2 phase HEp‐2 cancer cells to further analyze and specifically localize this effect. One hour (S phase), 4 hours (S phase) and 9 hours (G2 phase) after release from an aphidicolin double block synchronization procedure, cells were preincubated for 60 min with 30 μM chlorambucil and then radiolabeled for another 60 min in the continued presence of the agent. At the end of each of these time intervals, cells in almost mid‐S phase, late S phase and toward the end of the G2 phase were obtained for analysis. It was found that chlorambucil partially inhibits total histone synthesis nonspecifically as to the variants being synthesized (S phase and basal variants) but only during the first half of the S phase. DNA synthesis is also inhibited partially, but during the second half of the S phase. The position during the S phase where chlorambucil exerts its effect on total histone synthesis, the degree of this effect and its uncoupling with DNA synthesis inhibition, indicate that it is temporally linked with the onset of S phase histone
Antioxidants and cataract: (Cataract induction in space environment and application to terrestrial aging cataract)
Authors: Bantseev Vladimir , Bhardwaj Ratan , Rathbun W. , Nagasawa H. , Trevithick John R. 
Published: July 2007

AbstractThe effect of several antioxidants and cysteine‐elevating precursor drugs (prodrugs) was tested on lens damage occurring after in vitro exposure to low levels of 60Co‐y‐irradiation, to simulate in vitro the exposure to radiation in vivo of (1) astronauts (2) jet crews (3) military radiation accident personnel. Tocopherol (100 μM), ascorbic acid (1mM), R‐α‐lipoic acid (1mM), and taurine (0.5 mM) protected against radiation‐ associated protein leakage. MTCA and ribocysteine protected lenses against opacification, LDH and protein leakage, indicating that antioxidants and prodrugs of cysteine appear to offer protection against lens damage caused by low level radiation.
Expression of progesterone receptor mRNA in the first trimester human placenta
Authors: Shanker Y. Gopi , Sharma S. C. , Rao A. Jagannadha 
Published: July 2007

AbstractThe human placenta produces large quantities of progesterone, the function and target tissues of action of which during pregnancy are not completely understood. Although it has been suggested that placenta itself could be a potential target tissue for progesterone action, there is no conclusive evidence for the presence of progesterone receptors in the human placenta. We provide evidence for the expression of progesterone receptor mRNA in the human placenta by reverse transcription coupled to polymerase chain reaction (RT‐PCR). This was further confirmed by Southern hybridization, restriction analysis and sequencing of the PCR amplified fragment.
A comparison of the hydroxyl radical scavenging properties of the shark bile steroid 5β‐scymnol and plant pycnogenols
Authors: Macrides Theodore A. , Shihata Amal , Kalafatis Nicolette , Wright Paul F. A. 
Published: July 2007

AbstractThe hydroxyl radical (OH·) quenching abilities of the following compounds were compared in the deoxyribose degradation system (initiated by the ferrous‐ascorbate Fenton reaction): (a) 5β‐scymnol, the hepatoprotective shark bile sterol, and its mono‐ and di‐sulfate esters; (b) three marketed pycnogenol preparations (syn: proanthocyanidin ‐ natural plant‐derived polyphenolic bioflavonoids) extracted from pine tree (Pinus maritima) bark and grape (Vitis vinifera) seeds; and (c) two known hydroxyl radical scavengers, dimethyl sulfoxide and mannitol, and the peroxyl radical scavenger TroloxTM (the α‐tocopherol analogue). 5β‐scymnol was a more potent OH· quencher than dimethyl sulfoxide, mannitol and Trolox, and markedly more potent than the pycnogenol preparations. Increased sulfation of 5β‐scymnol progressively reduced its free radical scavenging activity, thus clearly attributing the potent OH· quenching properties to its novel tri‐alcohol‐substituted aliphatic side chain. The favourable interaction of these bile steroids with reactive oxygen species in an aqueous environment, makes them attractive candidates for evaluation as protective agents against disorders in which oxidative stress is implicated.
Developmental study of hepatic glutamine synthetase in a mouse model of congenital hyperammonemia
Authors: Skarpetas Andrew , Mawal Yogesh , Qureshi Ijaz A. 
Published: July 2007

AbstractThe development of hepatic glutamine synthetase (GS; EC 6.3.1.2) activity and expression was studied in 1 to 112 day old sparse‐fur (spf) mutant mice, with X‐linked ornithine transcarbamylase (OTC, EC 2.1.3.3.) deficiency. The spf/Y mutant mice were found to have a smaller body weight (p<0.01) yet possessed a larger liver (p<0.01‐0.05) in comparison to normal male mice (+/Y). The neonatal hepatic GS activity was retarded in the spf/Y mice (p<0.01) but reached normal values by the 28th day of age, after which it increased as compared to the control CD‐I mice (p<0.01). The spf GS activity remained constant from 28 to 56 days, whereas the CD‐I GS activity decreased. A further significant increase in the spf GS activity was observed from 56 day to 112 day indicating its adaptation. The decrease of GS mRNA in the spflY mice from 28 to 112 days of age (3.72±0.25 vs 1.68±0.32, p<0.01) suggests translational and post‐translational modifications in the regulation of GS activity. The changes in the activity and expression patterns of GS could be due to an effect of the OTC mutation on the hepatic ammonia metabolism. This may be indicative of the adaptational processes in the spf mutant mice, which may play a specific role in this animal model to help it to survive with its hyperammonemia.
cAMP‐dependent phosphorylation activates phosphofructokinase from mantle tissue of the mollusc Mytilus galloprovincialis. Identification of the phosphorylated site
Authors: Fernández Montserrat , Cao Jesús , Vega Félix V. , Hellman Ulf , Wernstedt Christer , Villamarín J. Antonio 
Published: July 2007

AbstractPhosphofructokinase from mantle tissue of the sea mussel Mytilus galloprovincialis was phosphorylated in vitro by a protein kinase isolated from the same tissue, homologous to mammalian cAMP‐dependent protein kinase; the maximal level of phosphorylation achieved was around 1 mol of Pi/mol of phosphofructokinase subunit. The covalent incorporation of phosphate leads to a notable increase in the enzyme activity assayed at near‐physiological concentrations of substrates and allosteric modulators and neutral pH. Tryptic digestion of labeled phosphofructokinase released a phosphopeptide whose sequence was Lys‐Asp‐Ser(P)‐Ile‐Trp‐Ile‐Gln‐Thr‐Gly‐Arg. This sequence showed high homology with the phosphopeptides from other invertebrates whose phosphofructokinase is also activated by cAMP‐dependent phosphorylation.
The nitric oxide donor S‐nitroso‐N‐acetyl‐D,L‐penicillamine degrades heparan sulfate and heparin
Authors: Ghael Dineshchandra , Mileva Mayia , Dweck Harry S. , Rosenfeld Louis 
Published: July 2007

AbstractNitric oxide (NO) is a powerful vascular and neural regulator. One of the breakdown products of nitric oxide is nitrite which converts to nitrous acid, a reagent routinely used for the degradation of heparin and heparan sulfate. We have recently shown that nitric oxide gas degrades heparin and heparan sulfate through a nitrous acid mechanism (Vilar et al, 1997, Biochemical Journal, 324, 473‐479). The purpose of the present study is to confirm these findings using the nitric oxide donor S‐nitroso‐N‐acetyl‐D,L‐penicillamine (SNAP) under conditions that are close to those found in vivo. The results show that 2 mM SNAP releases a steady‐state level of nitrite of over 200 μM. This level substantially degrades heparin and heparan sulfate at a pH of up to 5.0. This reaction may be important in breakdown of the glycosaminoglycan components of the extracellular matrix during normal and pathological conditions.
Iron as a possible mediator of the oxic‐to‐anoxic transition in Paracoccus denitrificans
Authors: Kucera Igor , Mat'chová Ivana 
Published: July 2007

AbstractExpression of the lacZ gene from the Fnr‐regulated FF‐melR promoter on a plasmid in iron‐deprived Paracoccus denitrificans cells required not only a decreased oxygen tension but also supplementation with iron. The levels of β‐galactosidase and 5‐aminolevulinate synthase showed comparable responses to changes in iron availability. The presence of soluble and particulate enzymes catalyzing the reduction of Fe(III) by NADH suggests a hypothesis in which the redox state of the cytoplasmic NAD‐couple determines the concentration of free Fe(II) and thereby modulates the activity of a common regulator of the Fnr type.
cDNA cloning and genomic organization of enhancer of split groucho gene from nematode Caenorhabditis elegans
Authors: Sharief Farida S. , Tsoi Stephen C. ‐M. , Li Steven S. ‐L. 
Published: July 2007

AbstractThis first genomic Enhancer of split groucho (ESG) gene and its full length complementary DNA (cDNA) from nematode C. elegans were cloned and sequenced via homology with the corresponding Drosophila groucho cDNA. The cDNA of 2.1‐Kb encodes a protein of 612 amino acids, and the nematode ESG protein is the smallest and most different in structure compared to all ESG related proteins. The gene isolated is 4,246‐bp in size, including 1,219‐bp promoter region. A putative TATA‐box at position ‐1166, two consensus sequence of ACTGG, characteristic of leader binding protein‐1 (LBP‐1) binding motifs at position ‐563 and ‐211 and nine CAAT boxes were found in the promoter region of ESG gene. The protein‐coding sequence is interrupted by five introns. The length of introns 1 to 5 is 52, 252, 87, 53 and 518 bp, respectively. The overall structural relationships of the ESG‐related proteins among human, mouse, rat, Xenopus, Drosophila and nematode were also analyzed.
Binding characteristics of sle anti‐DNA autoantibodies to modified DNA analogue
Authors: Arjumand Sadia , Ali Asif 
Published: July 2007

AbstractNative calf thymus DNA was modified with furoconmarins and reactive oxygen species (ROS). The modifications were probed by UV & Fluorescence spectroscopy, thermal denaturation and hydroxyapatite chromatography. In CD spectroscopy changes in the ellipticity of the DNA molecule were observed as a result of modification. The binding specificity of naturally occurring anti‐DNA antibodies with modified DNA molecules was assessed by ELISA and quantitative precipitin titration. The affinity ofanti‐DNA antibodies for modified conformers was found to be quite high.
Expression of chloroplastic genes during autumnal senescence in a deciduous tree Populus deltiodes
Authors: Reddy M.S. Srinivasa , Trivedi Prabodh K. , Tuli Rakesh , Sane Prafullachandra V. 
Published: July 2007

AbstractIn Populus deltoides, a deciduous tree, the development on new leaves starts in the month of March, the leaves reach maturity by October and fall by December. Changes in the composition and function of the photosynthetic apparatus were analysed during autumnal senescence. With the progress of senescence, there was an initial increase followed by a decrease in the steady state levels of psbA, psbD/C and psaA/B gene transcripts. Decrease in the steady state level of D1 protein was faster than that of Cytochrome f. The decline in LHCP level was seen only during late senescence. Although the leaves continue to look green and healthy till late November, the electron transport driven by individual photosystems started declining by October end suggesting the onset of senescence.
Cloning and functional analysis of rat cyclin D2 promoter: multiple prolactin‐responsive elements
Authors: Yang Miao , Hosokawa Yoshitaka , Hu Ying , Kaneko Satoshi , Kaneko Hiroshi , Tanaka Minoru , Nakashima Kunio 
Published: July 2007

AbstractStructure and function of the promoter region of rat cyclin D2 gene were investigated by cloning, sequence analysis and DNA mobility shift assay using nuclear extracts of prolactin‐stimulated Nb2 cells. The results show that the expression of rat cyclin D2 gene is not dependent on the TATA‐box but is stimulated by multiple transcription factors induced by prolactin at an immediate‐early stage of the cell cycle.
Stimulation of cytosolic phospholipase A2‐catalyzed arachidonic acid liberation by low dose tert‐butyl hydroperoxide without an influence on the enzyme activity in rabbit platelets
Authors: Akiba Satoshi , Nagatomo Ryo , Hayama Misako , Sato Takashi 
Published: July 2007

AbstractThe effect of lipid peroxide on the hydrolytic action of cytosolic phospholipase A2 (cPLA2) in rabbit platelets was investigated. Ionomycin‐stimulated arachidonic acid liberation and lysophosphatidylcholine formation were significantly potentiated when platelets were pretreated with tert‐butyl hydroperoxide (BHP) and FeSO4, and then washed. Under the conditions, oxidizing reagents did not enhance the increase in cPLA2 activity by ionomycin or the basal activity in unslimulated cells. Furthermore, the treatment of a platelet lysate with BHP and FeSO4 did not affect Ca2+‐induced translocation of cPLA2 to the membranes. However, with a membrane fraction, arachidonic acid liberation catalyzed by the partially purified cPLA2 was synergistically enhanced by BHP and FeSO4. These results suggest that oxidative stress may potentiate the hydrolytic action of cPLA2 on membrane phospholipids without an influence on the processes leading to the enzyme activation.
Cloning and characterization of a rhamnogalacturonan hydrolase gene from Botrytis cinerea
Authors: Chen Huey‐Jen , Smith David L. , Starrett David A. , Zhou Dingbo , Tucker Mark L. , Solomos Theophanes , Gross Kenneth C. 
Published: July 2007

AbstractRhamnogalacturonan hydrolase (Rgase A) cleaves α1→2 linkages between rhamnosyl and galacturonosyl residues in pectin. A 1.9 kb RGase A cDNA clone (BCRHGA) was isolated from a B. cinerea cDNA library using a PCR‐amplified Aspergillus aculeatus RGase A probe. It's 1.7 kb open reading frame had 62% identity at the amino acid level with A. aculeatus RGase A. Northern blots of B. cinerea total RNA probed with BCRHGA revealed a 2 kb band, suggesting the cDNA clone is full or nearly‐full length. To determine mRNA expression of the gene, B. cinerea was grown in media containing 0.5% apple pectin, 0.5% rhamnogalacturonan‐I and 1% glucose carbon sources. Northern analysis revealed the BCRHGA gene was expressed on all carbon sources, but with different patterns of expression. B. cinerea RGase A appeared to be coded for by a single or low copy number gene based on Southern analysis.