bluuebunny/crossref_metadata_2025 huggingface dataset:
Monoclonal antibodies against vascular endothelial growth factor 165 (VEGF165): Neutralization of biological activity and recognition of the epitope
Authors: Chen Jun‐Hui , Wang Xin‐Chang , Sato J. Denry
Published: July 2007

Five antibodies against vascular endothelial growth factor 165 (VEGF165) were obtained. These antibodies, Ab‐2, Ab‐20, Ab‐153, Ab‐309, Ab‐342, were able to recognize not only native VEGF165, but also reducing VEGF165. Three of the antibodies, Ab‐153, Ab‐309, Ab‐342, were identified as VEGF165 neutralizing antibody on the basis of its ability to inhibit the proliferation of human umbilical vein endothelial cells (HUVEC) induced by VEGF165 and the binding of [125I]‐VEGF165 to receptors on HUVEC. The fragments of E1 (VEGF120‐135) and E2 (VEGF46‐60) recognized by neutralizing antibodies may be related the receptor binding domain of VEGF.
Cloning of a tellurite resistance determinant from Bacillus stearothermophilus V in Escherichia coli
Authors: Vásquez Claudio , Saavedra Claudia , Loyola Claudia , Moscoso Hugo , Pichuantes Sergio
Published: July 2007

A potassium tellurite‐resistance determinant was isolated from Bacillus stearothermophilus V and cloned in Escherichia coli. Transformed cells formed black colonies when grown on solid media containing permissive tellurite concentrations. The resistance determinant was contained in a B. stearothermophilus V chromosomal DNA fragment of 7 kb.
Cloning, sequence analysis and expression in E. coli of the cDNA of the thrombin‐like enzyme (pallabin) from the venom of Agkistrodon halys pallas
Authors: Fan Chun‐Yang , Qian You‐Cun , Yang Sheng‐Li , Gong Yi
Published: July 2007

The cDNA of the thrombin‐like enzyme (pallabin) from the venom of Agkistrodon halys pallas was cloned and sequenced. The length of the cDNA is 923bp which includes 120bp of noncoding region and 780bp of coding region. Pallabin was synthesized as a prozymogen with 260 amino acids, which includes a signal peptide of 18 amino acids, a proposed propeptide of 6 amino acids and a matured peptide of 236 amino acids. Pallabin exhibits a strong amino acid similarity to the serine proteases isolated from other snake venoms. It contains 12 cysteins which form 6 disulfide bridges. Like other serine proteases, it also has three conserved catalytically active sites: His41, Asp86 and Ser182. To our knowledge, this study is the first report concerning the cDNA of a thrombin‐like enzyme from Agkistrodon halys pallas. The cDNA was cloned into the expression plasmid pT7ZZa and expressed in E.coli. The recombinant pallabin immunologically reacted with its specific antibody.
Effect of dietary fish oil on tumor‐induced hyperlipidemia and tumor growth in rats implanted with a methylcholanthrene‐induced sarcoma
Authors: Radcliffe J. , Narins D. Czajka , Imrhan V. , Edwards M.
Published: July 2007

Male Fischer 344 rats implanted with a methylcholanthrene‐induced sarcoma (MCS), along with normal (or control) animals, were fed diets containing either 10% corn oil (CO) or 2% CO + 8% fish oil (FO), designated as diets CO and FO, respectively, in a study designed to determine the effect of dietary FO on serum lipids (in the presence or absence of a tumor) and the growth and fatty acid composition of the MCS. For both diets, MCS‐bearing rats had significantly (p<0.05) higher serum levels of triglycerides, cholesterol, phospholipids, and total lipids than controls. For both controls and tumor‐bearers, serum levels of all these lipids were, with the exception of cholesterol for the tumor‐bearers, significantly lower in rats receiving the FO diet than for the corresponding groups receiving the CO diet. Relative to rats fed the CO diet, those fed the FO diet had significantly higher serum levels of some fatty acids (e.g., 20:5n‐3) but significantly lower levels of others (e.g.,18:2n‐6), regardless of tumor status. For the tumor‐bearers, differences in the levels of fatty acids in MCS tissue reflected differences in the fatty acid composition of total serum lipids. Sarcoma growth was unaffected by diet. Thus, feeding dietary FO resulted in changes in the lipid status of both control and tumor‐bearing rats. Since sarcoma growth was unaffected by diet, the reduction in the severity of MCS‐induced hyperlipidemia by FO appears to be due to an effect of the oil per se.
Sedimentation analysis of muscle glycogen phosphorylase b entrapped in hydrated reversed micelles of aerosol OT in octane
Authors: Kurganov B. I. , Chebotareva N. A.
Published: July 2007

The oligomeric state and formation of supramolecular structures of glycogen phosphorylase b from rabbit skeletal muscles have been studied in the system of hydrated reversed micelles of aerosol OT (AOT) in octane. Sedimentation studies show that the oligomeric state of the enzyme is controlled by the degree of hydration of micelles. Monomeric, dimeric, trimeric, tetrameric, hexameric, or octameric forms of the enzyme were observed depending on the degree of micelle hydration.
Purification and characterization of a cell‐surface lectin (lectin II) from Agrobacterium radiobacter NCIM 2443
Authors: Syed F. B. F. , Joshi B. N. , SivaRaman H. , Khire J. M. , Khan M. I.
Published: July 2007

A lectin was isolated from Agrobacterium radiobacter cell surface and purified. It is a monomer of 40 kDa as shown by SDS‐PAGE. The lectin has a pI of 9.15 and amino acid composition of the lectin shows that 44% of the amino acids are hydrophobic. The lectin agglutinates rabbit erythrocytes but does not agglutinate human erythrocytes. It does not show specificity for monosaccharides except for D‐glucosamine. Fetuin and its N‐linked glycopeptide also inhibit the activity of the lectin but greater inhibition is shown by locust bean gum and Nicotiana tobaccum (tobacco) tissue extracts.
Concanavalin A binding to oligosaccharide chain leads to alterations in properties of Band 3
Authors: Zhang Zhen‐Gang , Lu Jing‐Ze , Chen Jian‐Wen
Published: July 2007

Anion transport activity and thermotropic behavior of Band 3 are found to be altered after binding of concanavalin (Con A) to human erythrocyte ghosts and isolated Band 3. At lower Con A concentration, the rate coefficients of anion transport enhance with increasing Con A concentration, while noticeable changes of the largest calorimetric endotherm of human erythrocyte membranes termed the C transition (Band 3) can not be observed. With 50μg/ml of Con A, the rate coefficient of Con A‐modified ghosts increases 34.4% in comparison with that of normal ghosts. Binding of Con A in lower concentration to ghosts bring about increase of fluidity of lipid which maybe contribute to increase anion transport via Band 3. At higher Con A concentration, the C transition tend to lower temperature with increase in Con A concentration. the C transition is shifted from 69.25°C to 66.25°C with 2.5mg/ml Con A. It is suggested that the Con A‐modified Band 3 possess a looser structure than normal one.
Modulation by nitric oxide (NO) of capsaicin‐induced calcium uptake into rat dorsal root ganglion neurons
Authors: Park Young‐Ho , Surh Young‐Joon , Lee Sang‐Sup
Published: July 2007

The regulatory role of nitric oxide in capsaicin‐induced 45Ca2+ accumulation in dorsal root ganglion neuronal cultures was investigated. Capsaicin‐activated calcium entry was subject to complicated tuning by NO‐releasing agents sodium nitroprusside, spermine/NO complex and NO synthase inhibitor NG‐nitro‐L‐arginine methyl ester in concentration and stimulation protocol‐dependent manner. In contrast, these agents failed to change depolarization‐induced calcium influx. In experiments using dithiothreitol and 5,5‐dithio‐bis‐2‐nitrobenzoic acid this modulation was independent of the oxidizing action of NO. It is suggested that NO exerts a novel feedback modulatory effects on capsaicin‐induced calcium entry into rat DRG neurons.
Cloning and characterization of the non‐catalytic heavy chain of mouse complement factor I gene: Structure comparison with the human homologue
Authors: Yun Young‐Sup , Goldberger Gabriel , Minta Joe O.
Published: July 2007

The gene sequence encoding the non‐catalytic heavy chain of mouse complement factor I (mCFI) was cloned and its exon‐intron organization and domain structure characterized. The genomic organization of mCFI differs in several aspects from its human homologue (hCFI). The intron sizes are remarkably different. Exons 2 and 4 in mCFI are larger than their counterparts in hCFI by 9 bp and 6 bp respectively. Whereas the diversity (D) region of hCFI is encoded by two exons (exon 7 or hD2 and exon 8 or hD4), this region in mCFI is encoded by three exons; exon 6A or mD1 (located at the 3′‐end of the LDLr A2 domain), exon 7 or mD2 and exon 8, an extended exon (56 bp) composed of mD3, fused upstream of mD4. In contrast, hCFI lacks D1 and D3 subregions and exon 8 in hCFI consists of only hD4, 36 bp in length. Thus the heavy chain of mCFI is organized into 10 exons compared to 9 exons in hCFI.
Introduction of NAD decreases fMLP‐induced actin polymerization in chicken polymorphonuclear leukocytes ‐ The role of intracellular ADP‐ribosylation of actin for cytoskeletal organization
Authors: Terashima Masaharu , Shimoyama Makoto , Tsuchiya Mikako
Published: July 2007

We reported previously that the arginine‐specific. ADP‐ribosyltransferase in chicken polymorphonuclear leukocytes specifically modified actin, thereby inhibiting actin polymerization in vitro. In the present study, we investigated the effect of ADP‐ribosylation on actin polymerization in situ. In the leukocytes, the introduction of NAD inhibited the increase in filamentous actin contents induced by a chemotactic peptide formyl‐methionyl‐leucyl‐phenylalanine, while introduction of NAD together with novobiocin, a specific inhibitor for ADP‐ribosyltransferase, did not. These results suggest that ADP‐ribosylation regulates the formation of filamentous actin by the covalent modification of the protein in vivo.
Polymerization of actin induced by a molar excess of ATP in a low salt buffer
Authors: Ikkai Takamitsu , Kondo Hiroshi
Published: July 2007

The polymerization of actin induced by dilution has previously been reported, where a 1000‐fold molar excess of ATP over actin resulted when actin was diluted to 4.0 μg/ml in low salt buffer A (0.1 mM ATP, 0.1 mM CaCl2, 2 mM Tris‐HCl, pH 8.0, 5 mM 2‐mercaptoethanol, 1 mM NaN3). Filaments formed by the addition of ATP to a 1000‐fold molar excess over actin in buffer B (0.1 mM CaCl2, 2 mM Tris‐HCl, pH 8.0, 1 mM NaN3) were then separated by gel‐filtration. When ATP was removed from these filaments using Dowex‐1, depolymerization occurred. Thus, the reversible polymerization induced by the dilution of actin or by addition of ATP can be ascribed to the binding of ATP at the low affinity site of actin.
C‐terminal region of hTPO is important for secretion and expression in insect cells
Authors: Man Hyea Kyung , Chung Joo Young , Park Sang Kyu , Joo Semg Myung , Park Seung Kook , Koh Yeo Wook
Published: July 2007

Human thrombopoietin (hTPO) variant cDNAs truncated in the C‐terminal regions of wild‐type hTPO (332 amino acids) were constructed by PCR and expressed in Trichoplusia ni (Tn5) insect cells using a baculovirus expression system. Each variant, hTPO163 (amino acids 1‐163), hTPO198 (1‐198) and hTPO245 (1‐245), was produced in insect cells with very low efficiency in comparison with wild‐type hTPO. Immunoblot analysis showed that the predicted 20, 25 and 34 kDa molecular sizes corresponding to hTPO163, hTPO198 and hTPO245, respectively, were barely detected in culture medium and most of the proteins remained within the cell. These results suggest that C‐terminal regions containing potential N‐glycosylation sites of hTPO are required for the secretion of hTPO into culture medium as well as expression in insect cells.
Phenotypic selection: A successful strategy to fix major genes of hypertension
Authors: Klöting Ingrid , Kovács Peter , Brandt Jens van den
Published: July 2007

Hypertension is dominantly inherited in cross hybrids between hypertensive SHR/Mol and normotensive BB/OK rats. We used these cross hybrids for repeated backcrossing of selected hypertensive animals onto normotensive BB/OK rats to fix high blood pressure and to generate a hypertensive and diabetic BB/OK rat subline. After 8 backcrosses, the backcross parents were genetically analysed with the aid of 259 microsatellite markers to identify SHR genes causing blood pressure of 177±10 mmHg in this BB/OK rat subline. Loci on chromosomes 1, 14 and 18 showed longest heterozygosity. These loci might contain major genes of the SHR rat causing hypertension in this BB/OK rat subline. This classical strategy seems to be most suitable to fix major genes of hypertension in particular and complex traits in general and therefore to generate new animal models.
Nucleoid proteins of pea chloroplasts: detection of a protein homologous to ribosomal protein
Authors: Oleskina Yu.P. , Yurina N. P. , Odintsova T. I. , Egorov Ts. A. , Otto A. , Wittmeann‐Liebold B. , Odintsova M. S.
Published: July 2007

Basic proteins were isolated from purified pea chloroplast nucleoids by acid extraction. Using RP‐HPLC, the component composition of the basic proteins was studied. SDS‐PAGE of major HPLC‐fractions showed that the basic nucleoid proteins are heterogeneous with mol. masses of components from 17 to 30 kDa. One polypeptide with mol. mass of 28 kDa (P28) was obtained by RP‐HPLC. The sequencing of three tryptic peptides of P28 (T6, T17, and T19) showed that they are homologous to the ribosomal protein L19 of Saccharomyces cerevisiae. The possible functional role of ribosomal proteins in chloroplast nucleoids is discussed.
Mitochondrial changes during the apoptotic process of HeLa cells exposed to cisplatin
Authors: Melendez‐Zajgla Jorge , Cruz Elizabeth , Maldonado Vihma , Espinoza Ana Maria
Published: July 2007

HeLa cells undergo apoptosis after exposure to cisplatin. Since mitochondria have recently been proposed as a probable effector of this type of cell death, we performed an analysis using the fluorescent cation rhodamine 123, which is transported actively by this organelle. Cisplatin induces a decrease in the mitochondrial staining, as assessed by cytofluorometric analysis. Microscopic analysis demonstrated that this effect was accompanied by damage of the mitochondria. These features were not exclusive of cisplatin, as other antineoplasic agents (taxol, etoposide) elicited similar effects. These results point toward the notion of a general effect of antineoplasic drugs over the mitochondria during induction of apoptotic cell death.
Induction of apoptosis by selenite and selenodiglutathione in HL‐60 cells: Correlation with cytotoxicity
Authors: Cho Dae‐Yeon , Jung Uhee , Chung An‐Sik
Published: July 2007

Effects of selenite and selenodiglutathione, an initial metabolite of selenite, on the induction of apoptosis and cytotoxicity were investigated in human promyelocytic leukemia HL‐60 cells. Treatment of selenite or selenodigtutathione resulted in concentration‐dependent cytotoxicity, measured by lactate dehydrogenase leakage assay, and by tetrazolium salt reduction assay. Selenodiglutathione has been shown to exert more cytotoxic effect than selenite in both assay systems. Time‐course study of cellular selenium uptake suggests that the higher cytotoxicity of selenodiglutathione be largely due to faster and greater selenium uptake rate. Treatment with selenite or selenodiglutathione also induced apoptosis in a dose‐dependent manner, as detected by enzyme‐linked immunosorbent assay and by DNA fragmentation assay. The dose‐response data of apoptosis induced by selenite or selenodiglutathione were similar to those of cytotoxicity, implicating a relationship between the induction of apoptosis and cytotoxicity. Zn, which is a well‐known inhibitor of apoptosis, dose‐dependently blocked not only the induction of apoptosis, but also the membrane damage induced by selenium, corroborating this hypothesis. It was noted that the inhibition of apoptosis by Zn exerted little protective effect on cytotoxicity at higher concentrations of selenium, compared with a perfect protective effect at low concentration of selenium. These results suggest that cytotoxicity induced by selenium may be partially correlated with apoptosis.
Chemical modification studies on the glucose/mannose specific lectins from field and lablab beans
Authors: Nadimpalli Siva Kumar
Published: July 2007

Seeds of Dolichos lablab var. lignosus (field beans) and variety typicus (lablab beans) contain glucose/mannose specific lectins that have been affinity purified and well characterised (Siva Kumar N., and Rajagopal Rao, D., J. Biosci., 1986, 10, 95‐109, (1) Rajasekhar etal., (Biochem.Archives. 1997, 13, 233‐240) (2). Purified lectins are glyeoproteins with a native molecular mass of 60 kDa and are made of two types of subunits (Gowda etal., 1994, J.Biol.Chem. 269, 18789‐18793) (3). Chemical modifications of various groups in purified lectins (using group specific reagents) such as lysine (citraeonie anhydride), carboxyl groups (water soluble earbodi‐imide) tyrosine (N‐acetyl imidazole) and tryptophan (2‐hydroxy 5‐nitro benzylbromide) revealed that 14 out of 21 residues of lysines 7 out of 92 residues of carboxyl groups, 16 out of 24 tyrosine residues and 2 out of 10 tryptophan residues were modified. Lysine and carboxyl group modification led to 95% loss in haemaglutinating activity compared to control while tyrosine and tryptophan modifications led to complete loss of lectin activity. Arginine and histidine modifications led to only 50% loss in activity. The extent of modification and loss in activity was same when the lysine and carboxyl groups were modified in the presence and absence of the inhibitory sugar methyl α‐D‐glucopyranoside at 0.1M concentration. However protection of modification and lectin activity was observed when the tyrosine and tryptophan residues were modified in the presence of the inhibitory sugar. Earlier CD studies carried out (1) and extensive chemical modification studies reported here substantiate the involvement of tyrosine and tryptophan residues in the sugar binding site of these lectins.
Changes in membrane‐bound leucine aminopeptidase activity during maturation and ageing of brain
Authors: Arechaga Garbiñe , Martínez José M. , Prieto Isabel , Ramírez María J. , Alba Fraucisco , Ramírez Manuel
Published: July 2007

Aminopeptidases are believed to be enzymes that regulate the activity of various neuropeptides. However, their physiological role, as well as their mechanisms of regulation, are not well understood. To analyze a part of the regulatory mechanisms that control the activity of these enzymes, the subcellular distribution of membrane‐bound leucyl aminopeptidase activity was studied in rat brain during development and ageing. Except in fetuses, the enzymic activity was greatest in the microsomal fraction in all ages tested. Except in microsomal and myelin fractions, compared with fetuses, leucyl aminopeptidase activity showed a decrease in 1‐week‐old rats and a subsequent increase to adult levels in 1‐month‐old rats. This profile differed in the microsomal fraction, where the activity increased steadily up to 1‐month‐old rats. After this age, the activity decreased progressively in 5‐month and 24‐month‐old rats. These results may reflect changes in the functional status of the susceptible substrates during development and ageing.
Regulation of an 8‐kDa peptide involved in testosterone production by luteinizing hormone in rat leydig cells
Authors: Ramaraj P. , Rao A. Jagannadha
Published: July 2007

Results of Western blot analysis carried out with an interstitial cell extract from male guinea pig and ovarian extract from immature female rats administered equine chorionic gonadotropin (eCG) provide supportive evidence to our earlier suggestion that an 8‐kDa peptide is involved in acquisition of steroidogenic capacity by the rat Leydig cells. It was found that though the signal was observed in other tissues such as liver, kidney and lung which do not produce gonadal hormones, the peptide was modulated only by lutenizing hormone (LH) in the rat Leydig cells.
The involvement of hexokinases in trehalose synthesis
Authors: Peixoto Daniele N. , Panek Anita D.
Published: July 2007

Yeast cells harboring a MAL2‐8c gene accumulate trehalose during the transition phase of growth on glucose due to the presence of the ADPG‐dependent trehalose 6‐phosphate synthase. Under these conditions, glueokinase appeared not to provide G‐6‐P for trehalose synthesis and the two hexokinases seemed to act synergistically. After incubation in d‐xylose, trehalose levels in these cells dropped almost in 90%, confirming the involvement of both hexokinases in the accumulation of this carbohydrate. Nevertheless, G‐6‐P levels appeared to be similar in all strains. Some explanations for this paradox are discussed. In stationary phase, neither of the three isoenzymes were involved in trehalose synthesis. Possibly, gluconeogenesis provides the substrate for trehalose synthesis at that stage.
The lysine and methionine rich basic subunit of buckwheat grain legumin: Some results of a structural study
Authors: Rout M. K. , Chrungoo N. K.
Published: July 2007

The 26 kD basic subunit of 280 kD buckwheat grain legumin has been partially characterized by measurement of its fluorescence and CD spectra. The protein has 22% α‐helix, 36% β‐sheet, 12% β‐turn and 30% random coil secondary structure. In comparison with the basic subunits of other legumin‐type proteins, the buckwheat legumin subunit has a high content of lysine and methionine. The protein also has higher ratios of lysine to arginine and methionine to arginine.
Porphyria‐induced hepatic porphyrinogen carboxy‐lyase inhibitor and its interaction with the active site(s) of the enzyme
Authors: de Catabbi S. Billi , de Cahmanovici R. Wainstok , Minutolo C. , Aldonatti C. , de Viale L.C. San Martin
Published: July 2007

Porphyrinogen carboxy‐lyase is an enzyme that sequentially decarboxylates uroporphyrinogen III (8‐COOH) to yield coproporphyrinogen III (4‐COOH). In mammals this enzyme activity is impaired by hexachlorobenzene treatment, through generation of an enzyme inhibitor. The interaction of porphyrinogen carboxy‐lyase inhibitor, extracted from the liver of hexachlorobenzene‐treated rats, with substrate decarboxylation sites on the enzyme, was studied using four different carboxylated substrates belonging to the isomeric III series of naturally‐formed porphyrinogens containing 8‐,7‐,6‐ and 5‐COOH.

Similar inhibitor effects were elicited against all the substrates assayed, with the exception of pentacarboxyporphyrinogen III in which decarboxylation was not inhibited to same extent.

Enzyme protection assays in the presence of the different substrates, indicated that each porphyrinogen protects its own decarboxylation from inhibitor action.

Preincubation of the inhibitor with normal enzyme increased its inhibitory effect. On the other hand, preincubation of both enzyme and inhibitor with superoxide dismutase or mannitol, did not alter inhibitory activity. Preincubation of the inhibitor with a number of aminoacids showed that only arginine and its derivative Nα‐Benzoyl‐L‐Arginine ethyl ester interact with the inhibitor, noticeably reducing its ability to inhibit porphyrinogen carboxy‐lyase. Albumin, histidine, serine, cysteine and imidazol, were unable to quench inhibitor activity.

The present results indicate that the inhibitor acts at the binding site of each porphyrinogen. Taking into account that arginine is related to enzyme activity, and that histidine is found at the binding site of the substrates, the results suggest that the inhibitor could bind to arginine residues, blocking the access of substrates to histidine and altering the adequate orientation for decarboxylation by masking the positively charged active site necessary for porphyrinogen binding to the enzyme. In addition an indirect effect of the inhibitor mediated through free radicals could be discarded.
cDNA Cloning of the Two Subunits of Phospholipase A2 Inhibitor PLIgamma from Blood Plasma of the Chinese Mamushi, Agkistrodon blomhoffii siniticus
Authors: Okumura Kohji , Inoue Seiji , Ohkura Naoki , Ikeda Kiyoshi , Hayashi Kyozo
Published: July 2007

Three phospholipase A2 (PLA2) inhibitors (PLI) have been purified from the blood plasma of the Chinese mamushi, Agkistrodon blomhoffii siniticus; 1 of these, PLI gamma, contains 2 homologous subunits, PLI gamma‐A and PLI gamma‐B. The cDNAs encoding these 2 subunits of PLI gamma were isolated from a liver cDNA library by using fragments from polymerase chain reaction amplifications as probes and sequenced. The respective nucleotide sequences encoded 19‐residue signal sequences, followed by 181‐residue proteins. The calculated molecular masses were 20123 and 20150 Da for the PLI gamma‐A and PLI gamma‐B subunits, respectively; and PLI gamma‐A included a N‐linked carbohydrate site at Asn‐157. The sequences of these subunits contained 2 internal repeats of disulfide‐bonding pattern characteristic to those of urokinase‐type plasminogen activator receptor and members of the Ly‐6 superfamily. A phylogenetic analysis comparing the amino acid sequences of PLI gamma‐A and PLI gamma‐B with those for other snakes revealed that the gene duplication leading to these 2 subunits occurred before the divergence of Viperidae and Elapidae.
Identification of a Novel Cytosolic Tocopherol‐Binding Protein: Structure, Specificity, and Tissue Distribution
Authors: Stocker Achim , Zimmer Sabine , Spycher Stefan E. , Azzi Angelo
Published: July 2007

alpha‐Tocopherol plays an important role as a lipid‐soluble antioxidant. It is present in all major mammalian cell types and shows tissue‐specific distribution. This suggests the presence of specific proteins involved in intracellular distribution or metabolism of alpha‐tocopherol. A diminution of tocopherol plasma concentrations contributes to the development of diseases such as vitamin E deficiency (AVED), atherosclerosis, and prostate cancer. Further evidence has been obtained for the existence of sites in cellular metabolism and signal transduction where alpha‐tocopherol potentially plays a regulatory role. A signal transduction modulation specific for alpha‐tocopherol has been described in several model systems. Using radioactively labeled alpha‐tocopherol as tracer, we have isolated a new alpha‐tocopherol‐associated protein (TAP) from bovine liver. This protein has a molecular mass of 46 kDa and an isoelectric point of 8.1. From its partial amino acid sequence, a human gene has been identified with high homology to the newly described protein. Sequence analysis has established that the new TAP has structural motifs suggesting its belonging to a family of hydrophobic ligand‐binding proteins (RALBP, CRALBP,alpha‐TTP, SEC 14, PTN 9, RSEC 45). Human TAP has been cloned into Escherichia coli, and its tissue‐specific expression has been assessed by Northern blot analysis.
Rate of 14CO2 Production from Variously Labeled Forms of [14C]Glucose in Human Breast Invasive Ductal Carcinoma Tissues
Authors: Yan Shou He , Wallon Joelle , Mayaudon Jacques
Published: July 2007

To establish possible cancerous aggressiveness between the metabolism of variously labeled [14C]glucose in the human breast invasive ductal carcinoma (IDC) tissues, we measured the rates of 14CO2 production from those tissues by using radiorespirometry, expressing the results as initial velocity (V) in nanomoles of 14CO2 min‐1 g‐1 of fresh tissues. The Vc data were compared with results of the SBR system, which grades up from I to III. Vc,1 values measured with [1‐14C]glucose increased from 1.99‐2.82 for SBR I to 3.90‐4.09 for SBR II, finally reaching 4.83‐7.04 for SBR III, thus matching clearly the increase of IDC cancerous aggressiveness. Conversely, data obtained from [3,4‐14C]glucose and [6‐14C]glucose decreased with increasing cancer stage: i.e., with [3,4‐14C]glucose, Vc,3,4 values were 5.79‐9.34 for SBR I, 4.45‐4.84 for SBR II, and 2.35‐1.90 for SBR III; with [6‐14C]glucose, the corresponding Vc,6 values were 1.34‐1.90, 1.33‐1.41, and 0.72‐0.79. The Vn,1/Vn,6 ratios were close to unity for normal tissues and for noncancerous tissues surrounding SBRI tumors. For cancerous tissues, however, the Vc,1/Vc,6 ratios were 1.5, 2.9, and 6.1‐9.8 in IDC tissues graded as SBR I, II, and III, respectively. The results suggest the possible use of radiorespirometry as a tool to assess IDC aggressiveness.
Organizational Complexity of a Rice Transgene Locus Susceptible to Methylation‐Based Silencing
Authors: Kumpatla Siva P. , Hall Timothy C.
Published: July 2007

Molecular analyses of a rice (Oryza sativa L.) transgene locus introduced using biolistic techniques revealed the presence of multiple copies of rearranged fragments, as well as an intact copy of the supplied constructs. Both the gene of interest (35S‐Btt cryIIIA) and the selectable marker used (Ubi1‐bar) were methylated and silenced. Additionally, vector sequences were present in great abundance and were also highly methylated, indicating that the entire transgene insert was marked for methylation. The rearrangement of input DNA resulted in interspersion of plasmid backbone regions with the gene of interest. Permutation of segments encoding the gene of interest and the selectable marker was also detected, perhaps explaining why sequences introduced on separate plasmids are frequently found to be inserted at the same locus. The 35S promoter contained several hotspots for fragmentation. These observations strongly support the concept that intrusive DNA is recognized by host surveillance systems and that transgene loci with anomalous structural organization are subjected to inactivation by processes such as methylation.
Origin of the Suppression of Chloride Ion Sensitivity in Human Embryonic Hemoglobin Gower II
Authors: Zheng Tao , Zhu Qing , Brittain Thomas
Published: July 2007

The molecular origin of the observed reduction of chloride ion sensitivity in the oxygen binding characteristics of the human embryonic hemoglobin Gower II has been probed by using site‐directed mutant forms of the protein. Separate mutation of the three amino acids previously identified as candidates for the suppression of chloride sensitivity in the epsilon chain allowed us to unambiguously identify the His beta77 epsilon Asn as the origin of the lower sensitivity towards chloride ions in hemoglobin Gower II, which allows oxygen exchange from the mother to the late embryo under physiological conditions.
Molecular Cloning, Sequencing, and Expression of Two Late Proteins of Bacteriophage MB78
Authors: Kolla Venkatadri , Datta Pinaki , Chakravorty Maharani
Published: July 2007

Bacteriophage MB78, a virulent phage of Salmonella typhimurium, does not allow other phages, such as P22 and 9NA, to grow in its presence. A detailed physical map of this phage has been constructed in our laboratory. In an ongoing effort to understand the genetics of this interesting phage, various genes were characterized. Here, we report cloning, sequencing, and expression of two late proteins, coded in a Sal I‐ Hind III fragment (SH9), by using the minicell expression system. Further, we performed a kinetic study of phage proteins by infecting the host LT2 cells and compared the proteins produced, with proteins obtained by the minicell expression system. Both sets of proteins run exactly parallel and migrated as 14‐ and 15‐kDa proteins on a polyacrylamide gel. The synthesis of these two proteins started 15 min after infection with MB78 and was prominent after 45 min. One of the proteins exhibited 57% homology to the structural protein of mycobacteriophage L5.
Thiolation of Low‐Mr Phosphotyrosine Protein Phosphatase by Thiol‐Disulfides
Authors: Degl'Innocenti Donatella , Caselli Anna , Rosati Fabiana , Marzocchini Riccardo , Manao Giampaolo , Camici Guido , Ramponi Giampietro
Published: July 2007

Thiol‐disulfides cause a time‐ and a concentration‐dependent inactivation of the low‐Mr phosphotyrosine protein phosphatase (PTP). We demonstrated that six of eight enzyme cysteines have similar reactivity against 5,5‐dithiobis(nitrobenzoic acid): Their thiolation is accompanied by enzyme inactivation. The inactivation of the enzyme by glutathione disulfide also is accompanied by the thiolation of six cysteine residues. Inorganic phosphate, a competitive enzyme inhibitor, protects the enzyme from inactivation, indicating that the inactivation results from thiolation of the essential active‐site cysteine of the enzyme. The inactivation is reversed by dithiothreitol. Although all PTPs have three‐dimensional active‐site structures very similar to each other and also have identical reaction mechanisms, the thiol group contained in the active site of low‐Mr PTP seems to have lower reactivity than that of other PTPs in the protein thiolation reaction.
Construction of Murine Phage Antibody Library and Selection of Ricin‐Specific Single‐Chain Antibodies
Authors: Gao Xinsheng , Huang Yixiu , Zhu Shenggeng
Published: July 2007

A murine phage antibody library containing 1.7 108 independent clones was constructed and then screened by affinity panning of anti‐ricin antibody fragments. First, mRNA was purified from the total RNA extracted from fresh spleens of nonimmunized mice of Kunming, and then the total antibody variable region cDNA was amplified via reverse transcription‐polymerase chain reaction. Gene fragments encoding VH and VL were amplified and assembled into a single gene by using a DNA linker encoding a pentadecapeptide (Gly4 Ser)3 through primary PCR. Finally the recombinant DNA fragments were cloned into the phagemid pCANTAB 5E vector and electroporated into Escherichia coli TG1. The recombinant phage particles displaying functional single‐chain fragment variable regions (scFvs) were rescued by reinfection of helper phage M13KO7, thus constructing a murine antibody library. Two ricinspecific scFvs strains were selected from the phage antibody library by using affinity panning. The target antigen, ricin, was biotinylated with photobiotin and the biotin‐labeled ricin interacted with the phage antibody library. Subsequent addition of streptavidin‐coated paramagnetic beads isolated the biotinylated ricin‐binding phage particle complex. Washing, elution, and reinfection of the isolated complex then proceeded sequentially. After six rounds of panning, 2 strains of the 34 clones were verified to show greater specific affinity to ricin.
5′ to 3′ Single Strand DNA Exonuclease Activity in a Preparation of Human Ku Protein
Authors: Morozov Viktor E. , Fuller Brian G.
Published: July 2007

We describe a novel 5′ to 3′ single‐strand exonuclease activity exhibited by a Ku preparation purified from a human cell line. The enzyme removes 5′ single‐strand extensions from duplex DNA molecules. The exonuclease and helicase activities respond reciprocally to changes in ATP concentrations: Nuclease activity is inhibited at the ATP concentrations that are optimal for the helicase. The exonuclease activity does not require divalent cations. The potential implications of the exonuclease activity findings for repair of double‐strand breaks and recombination processes are discussed.
A Short Region Containing an AP‐1 Binding Site Is Essential for Transforming Growth Factor‐beta‐Induced c‐jun Gene Expression in Osteoblastic Cells
Authors: Hata Satoshi , Shimizu Toshikatsu , Fujimoto Masafumi
Published: July 2007

Transforming growth factor‐beta (TGF‐beta) is a multifunctional regulatory peptide that elicits different responses in different cell types. Much remains unknown about the pathway of intracellular TGF‐beta signal transduction, but TGF‐beta is known to induce expression of several genes by way of the transcription factor AP‐1. We studied the mechanism that mediates TGF‐beta?induced gene expression of c‐jun, a component of AP‐1, in MC3T3‐E1 osteoblastic cells. To map in detail the corresponding responsive elements in the rat c‐jun promoter, we generated a series of 5′ deletion promoter/luciferase reporter gene constructs. Transient cell transfection assays identified the region located between positions ‐79 and ‐59 as being critical for the TGF‐beta response and for the basal activity of the promoter. Gel mobility shift assays indicated specific binding of nuclear proteins to this 21‐bp region of the c‐jun promoter containing an AP‐1 binding site. These results show that the AP1?dependent mechanism is involved in TGF‐beta?induced increase of c‐jun induction, suggesting positive autoregulation of AP‐1.
A Replicator Was Not Involved in the Origin of Life
Authors: Shapiro Robert
Published: July 2007

Many scientific theories of the origin of life suggest that life began with the spontaneous formation of a replicator (a self‐copying organic polymer) within an unorganized chemical mixture, or ?soup.? A profound difficulty exists, however, with the idea of RNA, or any other replicator, at the start of life. Existing replicators can serve as templates for the synthesis of additional copies of themselves, but this device cannot be used for the preparation of the very first such molecule, which must arise spontaneously from an unorganized mixture. The formation of an information‐bearing homopolymer through undirected chemical synthesis appears very improbable. The difficulties involved in such a synthesis are illustrated by considering the prospects for the assembly of a polypeptide of L‐alpha‐amino acids, based on the contents of the Murchison meteorite as an example of a mixture of abiotic origin. In that mixture, potential replicator components would be accompanied by a host of interfering substances, which include chain terminators (simple carboxylic acids and amines), branch‐formers, D‐amino acids, and many classes of substances for which incorporation would disrupt the necessary structural regularity of the replicator. Laboratory experiments dealing with the nonenzymatic synthesis of biopolymers have not addressed the specificity problem. The possibility that formation of the first replicator took place through a very improbable event cannot be excluded, but greater attention should be given to metabolism‐first theories, which avoid this difficulty.
Model of Detergent‐Induced Spectral Changes of the B800‐850 Complex from Chromatium minutissimum
Authors: Razjivin Andrei P. , Moskalenko Andrei A. , Novoderezhkin Vladimir I.
Published: July 2007

The absorption and circular dichroism spectra of the B800‐850 complex from Chromatium minutissimum
Innate immunity modulates autoimmunity: type 1 interferon‐β treatment in multiple sclerosis promotes growth and function of regulatory invariant natural killer T cells through dendritic cell maturation
Authors: Gigli Gianluigi , Caielli Simone , Cutuli Daniela , Falcone Marika
Published: July 2007

Type 1 interferon‐β (T1IFN‐β) is an innate cytokine and the first‐choice therapy for multiple sclerosis (MS). It is still unclear how T1IFN‐β, whose main function is to promote innate immunity during infections, plays a beneficial role in autoimmune disease. Here we show that T1IFN‐β promoted the expansion and function of invariant natural killer (iNKT) cells, an innate T‐cell subset with strong immune regulatory properties that is able to prevent autoimmune disease in pre‐clinical models of MS and type 1 diabetes. Specifically, we observed that T1IFN‐β treatment significantly increased the percentages of Vα24+ NKT cells in peripheral blood mononuclear cells of MS patients. Furthermore, iNKT cells of T1IFN‐β‐treated individuals showed a dramatically improved secretion of cytokines (interleukins 4 and 5 and interferon‐γ) in response to antigenic stimulation compared to iNKT cells isolated from the same patients before T1IFN‐β treatment. The effect of T1IFN‐β on iNKT cells was mediated through the modulation of myeloid dendritic cells (DCs). In fact, DCs modulated in vivo or in vitro by T1IFN‐β were more efficient antigen‐presenting cells for iNKT cells. Such a modulatory effect of T1IFN‐β was associated with up‐regulation on DCs of key costimulatory molecules for iNKT (i.e. CD80, CD40 and CD1d). Our data identified the iNKT cell/DC pathway as a new target for the immune regulatory effect of T1IFNs in autoimmune diseases and provide a possible mechanism to explain the clinical efficacy of T1IFN‐β in MS.
How Indirect Can Indirect Utilitarianism Be?*
Authors: WILAND ERIC
Published: July 2007

Most act‐utilitarians now reject the direct utilitarianism of Bentham. They do so because they are convinced of what I call the paradox of utilitarianism—the thought that one cannot maximize happiness if one is trying to maximize happiness. Instead, they adopt some form of indirect utilitarianism (IU), arguing that the optimal decision procedure may differ markedly from the criterion of rightness for actions. Here I distinguish between six different versions of indirect utilitarianism, arguing that the weaker versions of IU also fall prey to the paradox of utilitarianism, while the stronger versions of IU violate an overwhelmingly plausible moral principle, the principle that one ought to V only if one can V intentionally.
Political forecasting: The missing link
Authors: Ascher William
Published: July 2007

Political forecasting provides the contextuality needed for decision‐making and for forecasting ‘non‐political’ trends. To gear political forecasting to these needs, rather than mimicking approaches in other areas, requires recognition of the distinctive nature of political trends, and realism regarding forecast uses, which generally do not benefit from ‘precise’ probabilities, predictions of only major events, or ‘sophisticated’ methodology that sacrifices comprehensiveness for explicitness. Approaches borrowed from other forecasting disciplines have been counterproductive, although contextual approaches, including cross‐impact analyses and developmental constructs that integrate political and non‐political trends, are promising. Explorations of the consistency of scenario dynamics, taking into account policy responses and non‐formalizable complexity, are also useful. Thus the separation of political forecasting from political analysis should be minimized, calling for a redirection of effort away from developing methodology uniquely geared to forecasting, and towards organizing more comprehensive and systematic analytical efforts.
Aid co‐ordination
Authors: Ross David J.
Published: July 2007

Aid co‐ordination entails the exchange of information, an attempt to agree on aid objectives and priorities, and agreement on projects to be undertaken concurrently or jointly. It should be a mechanism for the more effective delivery of development assistance. If it works it produces several benefits. It prevents waste. It can reduce duplication of effort. It can free officials for more productive activities than unnecessary meetings with donors. Harmonizing aid procedures, reporting requirements and monitoring should reduce the administrative burden on recipient governments. For aid co‐ordination to work the administrative capacities of recipient governments need to be strengthened. Co‐ordination can be enhanced by improving the leadership role of multilateral agencies and recipient governments. Adequate resources need to be devoted to improving co‐ordination by the aid donors. Co‐ordination must be structured and organized. Monitoring capacities also need to be improved.
Problems of co‐ordination in upazila administration in Bangladesh
Authors: Minhaj Ahmad A. J. Uddin
Published: July 2007

The present government of Bangladesh has extensively reformed the administrative system and upgraded the previous thana to upazila (sub‐district) to serve as the focal point of development administration. A new tier of local government called Upazila Parishad (UZP) has been created and a large number of functions have been devolved to this institution along with the placement of central government employees at its disposal. The reformers argue that the present system will remove the inadequacies of the previous administrative system and will bring about an improvement in the co‐ordination of field services at the upazila level. However, the actual situation represents a totally different picture. The desired integration— both vertical and horizontal—and co‐ordination in local level planning is non‐existent due to the absence of any clear direction from the central government. Proper co‐ordination among the field services in the upazila administration does not exist due to the anomalous relation between the upazila and the national government, including excessive central control; the involvement of multiple agencies of the central government in guiding and supervising; ambiguity in government thinking; the absence of clarification from the ministries concerned and agencies on the timing of different development programmes; and conflict over the allocation of resources. There is also conflict between the different tiers of local government in respect of authority and resources. Relations between public representatives and officials at the upazila level are strained because a clear allocation of authority to political executives is lacking; areas for co‐operation among different agencies have not been clarified; and public representatives and government officials lack mutual respect.
Linking action training to bureaucratic reorientation and institutional reform
Authors: Schmidt Gregory D.
Published: July 2007

This article explores the use of action‐training methods in a highly centralized and control‐oriented public sector. It focuses on the Technical Assistance and Training Programme for Departmental Development Corporations (PATC‐CORDES), an innovative programme in Peru initially supported by the United States Agency for International Development (USAID). Successive sections review a particularly influential alternative to conventional classroom training proposed by Honadle and Hannah (1982); provide relevant background; describe PATC‐CORDES organization and objectives; examine its training activities; assess the resulting institutional impacts; and posit important insights from the case study. The PATC‐CORDES experience demonstrates that action‐training methods can produce positive results in agencies that are part of a rigid public sector and, more importantly, that such methods can be linked to bureaucratic reorientation and institutional reform. Perhaps the most interesting and controversial conclusion is that recruitment of central administrators to lead training exercises can be a useful tactic for bureaucratic reorientation. The article is based on review of documents, reports, and evaluations prepared by persons working for USAID/Peru, PATC‐CORDES, and departmental development corporations (CORDES); extensive interviews with political leaders, managers, and professionals in these same organizations; analysis of Peruvian legislation; and other references cited in the text.
Large scale public projects—The personal connection
Authors: Goldenberg Tzvika Y. , Mevorach Baruch
Published: July 2007

The paper expands on an interesting phenomenon accompanying large‐scale projects, namely, the parallel inflation in costs and deflation in benefits from initiation to implementation and final evaluation or elimination altogether. Historically, three explanations have been provided for the above phenomenon: the deterministic effects of uncertainty; self‐interest evaluators; and the ‘vision’ factor. We claim that on face value one cannot accept as scientifically sound evidence the above three explanations of the constant decline in actual benefits. The ‘acceptance‐avoidance’ model which we bring to bear predicts a constant deterioration in actual benefits as the project develops. This psychological factor results in the use of the three above‐mentioned explanations as a ‘cover‐up’ for real underlying human doubts regarding the chances of success of crucial decisions taken by individuals.