bluuebunny/crossref_metadata_2025_split huggingface dataset:
Sustaining Protein Synthesis in the Absence of Rapid Cell Division: An Investigation of Plasmid‐Encoded Protein Expression inEscherichia coliduring Very Slow Growth
Authors: Flickinger Michael C. , Rouse Michael P. 
Published: March 2005

AbstractThe minimum growth rate capable of supporting plasmid‐encoded gene expression is determined using continuous cultures ofEscherichia coliMZ9387 at dilution rates (D) as low as 5% of the maximum specific growth rate. Expression from a low copy number plasmid, pMPR166, encoding cyanase under the control of Placis investigated in order to study plasmid‐encoded gene expression under conditions approaching starvation. Plasmid copy number was stabilized by selection in the presence of 500 μg/raL chloramphenicol by constitutive expression of chloramphenicol acetyl transferase (CAT). Plasmid retention was determined by dot‐blot hybridization and chloramphenicol resistance. The contribution of plasmid maintenance and cyanase expression to the maximum cell yield (Y′x/s) and the maintenance coefficient (ms) was determined for MZ9387 and MZ9387: pMPR166 under uninduced and IPTG‐induced conditions. The values ofYx/sand msfor non‐plasmid‐bearing cultures were 0.56 g of cell dry mass (DCM)/g of glucose and 0.26 g of glucose/g of DCM·h, respectively. The cell yield for plasmid‐bearing cultures under uninduced conditions (Y′x/s) was 0.28 g of DCM/g of glucose, with m°s= 0.08 g of glucose/g of DCM·h. These values decreased following induction of cyanase expression. Glucose consumption in the presence of IPTG was linearly related to the growth rate atD< 0.28 h−1but nonlinear at dilution rates greater than 50% of the maximum specific growth rate, indicating that cyanase expression alters metabolism and glucose consumption. The fraction of plasmid‐free cells decreased with decreasing Damköhler number (Da). These data confirm the usefulness ofDafor predicting the relationship between plasmid‐free and plasmid‐bearing cells where plasmids are stabilized by concentrations of antibiotic greater than the minimum plasmid‐free host cell growth inhibitory concentration. Specific cyanase expression increased as the dilution rate decreased toD= 0.15 h−1. BetweenD= 0.15 h−1andD= 0.14 h−1, expression decreased 7‐fold. At very low dilution rates (D≤ 0.06 h−1), nonseptated filamentous cells appeared. The appearance of filamentous cells could be reversed by increasing the dilution rate. These data are evidence that when plasmid copy number is stabilized by chloramphenicol resistance, a minimum dilution rate exists below which stringent regulation of protein synthesis dramatically reduces gene expression.
Effect of Water Content on the Glass Transition and Caking of Fish Protein Hydrolyzates
Authors: Aguilera Jose M. , Levi Guy , Karel Marcus 
Published: March 2005

AbstractThe glass transition temperature (Tg) and structure collapse (volumetric shrinkage) of a freeze‐dried fish protein hydrolyzate (HFP) were studied. An increase in the water activity from 0 to 0.64 reduced the Tg of HFP from 79.1 to −42.8 °C. The Gordon—Taylor equation was a good predictor for the plasticizing effect of water on Tg. At room temperature (19 °C), collapse was initiated at aw = 0.44 corresponding to a T — Tg value of 35.8 °C, and browning was evident above aw = 0.55. Above these critical values several structural changes occurred: shrinkage, collapse, browning, and setting into a sticky, high‐viscosity brown liquid. The viscosity of the matrix at the onset of collapse was 105–107 Pa·s, as estimated using the Williams—Landel—Ferry equation.
Effects of Abrupt and Gradual Osmotic Stress on Antibody Production and Content in Hybridoma Cells That Differ in Production Kinetics
Authors: Reddy Sridhar , Miller William M. 
Published: March 2005

AbstractAB2‐143.2 cells exhibit non‐growth‐associated antibody production (constant specific antibody production rate (qAb) throughout batch culture), while IND1 cells exhibit growth‐associated production (decrease in qAb during stationary phase). AB2‐143.2 cells increase qAb following abrupt batch osmotic stress. Enhanced antibody production in IND1 cells after batch osmotic shock is evidenced as maintenance of the higher exponential phase QAb into the stationary phase. AB2‐143.2 cells also increase qAb in response to gradual osmotic stress in continuous culture. In contrast, IND1 cells decrease 9Ab during gradual osmotic stress. Although the two cell lines differ in antibody production, they have very similar intracellular antibody content profiles. Both show (1) constant antibody content during the exponential phase in control culture with a decrease as cells enter stationary phase; (2) maintenance of exponential‐phase antibody content into the stationary phase after batch osmotic shock; and (3) no change in antibody content in response to gradual osmotic stress.
Characterization and Polyelectrolyte Precipitation of β‐Galactosidase Containing Genetic Fusions of Charged Polypeptides
Authors: Niederauer Mark Q. , Suominen Ilari , Rougvie Malcolm A. , Ford Clark F. , Glatz Charles E. 
Published: March 2005

AbstractGenetically engineered versions of β‐galactosidase were constructed through the addition of charged polypeptide fusion tails for the purpose of enhancing polyelectrolyte precipitation. Negatively charged aspartic acid tails and positively charged poly(arginine) tails were added to β‐galactosidase from Escherichia coli. These fusion proteins were all shown to possess specific activity equal to that of the native enzyme. Gel permeation and ion‐exchange chromatography provided evidence concerning the integrity of the tails as well as their altered charge characteristics. All enzymes containing charged tails displayed enhanced polyelectrolyte precipitation over the native enzyme. An optimal number of charged residues, beyond which no further enhancement of precipitation was observed, was found to be approximately 10 residues for each type of tail. No interference from nucleic acids was observed in the precipitation of positively tailed β‐galactosidase.
Effect of Sodium Hydroxide on the Solubilities of L‐Isoleucine, L‐Leucine, and L‐Valine
Authors: Brown Marena Gatewood , Rousseau Ronald W. 
Published: March 2005

AbstractThe solubilities of L‐isoleucine, L‐leucine, and L‐valine in aqueous solutions containing sodium hydroxide were determined. In all cases, the solubility increased as the solution pH was increased above the isoelectric point, and then it decreased sharply with further additions of base, after 1 mol of NaOH/mol of amino acid was added to the solution.
Effects of Carbon Dioxide, Air Flow Rate, and Inoculation Density on the Batch Growth of Catharanthus roseus Cell Suspensions in Stirred Fermentors
Authors: Van Gulik Walter M. , Nuutila Anna Maria , Vinke Ko L. , Ten Hoopen Hens J. G. , Heijnen Joseph J. 
Published: March 2005

AbstractA high reproducibility of batch growth of Catharanthus roseus could only be achieved by using identical inoculum material. It was found that the initial growth rate of the cells increased with increasing inoculation density, whereas the maximum growth rate was not affected. Withdrawal of the conditioned medium from the inoculum resulted in a strong decline in the initial growth rate, indicating that conditioning factors affected the initial growth rate. Neither increasing the air flow rate from 0.5 to 1.5 volume of air per volume of broth‐min−1 (vvm) nor CO2 enrichment of the aeration gas had any effect on the growth of the cells.
In Pursuit of the Optimal Fed‐Batch Process for Monoclonal Antibody Production
Authors: Bibila Theodora A. , Robinson David K. 
Published: March 2005

AbstractFed‐batch culture currently represents the most attractive choice for large scale production of monoclonal antibodies (MAbs), due to its operational simplicity, reliability, and flexibility for implementation in multipurpose facilities. Development of highly productive cell lines, maximization of cell culture longevity, and maintenance of high specific antibody secretion rates through genetic engineering techniques, nutrient supplementation, waste product minimization, and control of environmental conditions are important for the design of high‐yield fed‐batch processes. Initially simple supplementation protocols have evolved into sophisticated serum‐free multi‐nutrient feeds that result in MAb titers on the order of 1–2 g/L. Limited research has been published to date on the effects of various culture parameters on potentially important quality issues, such as MAb glycosylation and stability. Although most fed‐batch protocols to date have relied on relatively simple control schemes, increasingly sophisiticated algorithms must be applied in order to take full advantage of the potentially additive effects of manipulating nutrient and environmental parameters to maximize fed‐batch process productivity.
Protective Effects of Polymer Additives on Animal Cells Exposed to Rapidly Falling Liquid Films
Authors: Wu Jianyong , Daugulis Andrew J. , Faulkner Peter , Goosen Mattheus F. A. 
Published: March 2005

AbstractThe protective effects of polymer additives on insect cells against fluid mechanical damage was investigated in a falling film‐flow device. The falling liquid film creates rapidly moving air&&beta;;liquid interfaces and high fluid shear stress, mimicking the characteristics of a bursting bubble in aerated cell culture. The additives tested included a group of surface‐active polymers, (i. e., Pluronic F68, poly(ethylene glycol)s, and Tween 80) and a group of viscosity‐enhancing polymers (i. e., dextrans, methyl‐cellulose, and (carboxymethyl)cellulose). We found that methylcellulose, which was previously considered a viscosity‐enhancing polymer, actually had significant surface‐active properties. All of the surface‐active polymers exhibited significant protective effects, with Pluronic F68 and the higher molecular weight poly(ethylene glycol), PEG 20M, providing the best protection. In contrast, the viscosity‐enhancing polymers, with the exception of methylcellulose, showed little or no protection for insect cells in the film flow. All of the protective polymers had surface‐active properties, even though some of them did not change the surface tension in the actual insect cell medium. There was no correlation between the protective effect and the changes in liquid viscosity and surface tension due to the polymer additives. The level of protection was shown to be dependent upon the type of polymer, its concentration in the culture medium, and the polymer molecular weight. We concluded that the mechanism of protection of these surface‐active polymers was through interaction of the polymer molecules with the cell plasma membranes: a fast‐acting biological mechanism.
Improved Homogenization of Recombinant Escherichia coli following Pretreatment with Guanidine Hydrochloride
Authors: Bailey Stuart M. , Blum Paul H. , Meagher Michael M. 
Published: March 2005

AbstractPretreatment of recombinant Escherichia coli, expressing human growth hormone inclusion bodies, with guanidine hydrochloride and Triton X‐100 prior to high‐pressure homogenization has been investigated. Homogenates were analyzed for protein release, viscosity, and particle size. We were able to reduce the number of passes required for cell disruption and the number of downstream processing steps required for the recovery of protein from inclusion bodies by pretreating cells with guanidine HCl and Triton X‐100. Pretreatment of exponential growth phase cells with 1.5 M guanidine HCl and 1.5% Triton X‐100 gave adequate disruption after one pass at 41 MPa with a particle size distribution similar to that for untreated cells disrupted after one pass at 62 MPa. This combination of guanidine HCl and Triton X‐100 was also selected so as to wash the inclusion bodies without solubilization of the human growth hormone. Pretreatment of cells with 4 M guanidine HCl produced cell debris that was substantially smaller than the debris from untreated cells and partially solubilized the inclusion bodies. Cells harvested in the stationary growth phase were more resistant to high‐pressure homogenization and pretreatment.
Evaluation of UNIFAC for the Prediction of Partitioning in Two‐Phase Aqueous‐Organic Systems Containing Amino Acids and Oligomers
Authors: Roberts Jason D. , Daugulis Andrew J. 
Published: March 2005

AbstractTernary liquid‐liquid equilibria were predicted by the UNIFAC method for organic/aqueous biphasic systems containing amino acids and amino acid oligomers composed of glycine, alanine, phenylalanine, and tyrosine. UNIFAC was used to screen solvents for use as potential extractants, and UNIFAC‐calculated distribution coefficients were compared to data obtained experimentally. UNIFAC identified some novel extractants from a larger database of solvents. Distribution into the organic phase generally increased with the degree of polymerization of the oligomers. The presence of nonpolar residues in oligomers was found to decrease extraction over hydrophobic species alone. It was concluded that the polarity of the amide backbone and the zwitterion effects were significant in reducing the hydrophobicity of the side chains.
Myricetin, quercetin and catechin-gallate inhibit glucose uptake in isolated rat adipocytes
Authors: STROBEL Pablo , ALLARD Claudio , PEREZ-ACLE Tomás , CALDERON Rosario , ALDUNATE Rebeca , LEIGHTON Federico 
Published: March 2005

The facilitative glucose transporter, GLUT4, mediates insulin-stimulated glucose uptake in adipocytes and muscles, and the participation of GLUT4 in the pathogenesis of various clinical conditions associated with obesity, visceral fat accumulation and insulin resistance has been proposed. Glucose uptake by some members of the GLUT family, mainly GLUT1, is inhibited by flavonoids, the natural polyphenols present in fruits, vegetables and wine. Therefore it is of interest to establish if these polyphenolic compounds present in the diet, known to be effective antioxidants but also endowed with several other biological activities such as protein-tyrosine kinase inhibition, interfere with GLUT4 function. In the present study, we show that three flavonoids, quercetin, myricetin and catechin-gallate, inhibit the uptake of methylglucose by adipocytes over the concentration range of 10–100 μM. These three flavonoids show a competitive pattern of inhibition, with Ki=16, 33.5 and 90 μM respectively. In contrast, neither catechin nor gallic acid inhibit methylglucose uptake. To obtain a better understanding of the interaction among GLUT4 and flavonoids, we have derived a GLUT4 three-dimensional molecular comparative model, using structural co-ordinates from a GLUT3 comparative model and a mechanosensitive ion channel [PDB (Protein Data Bank) code 1MSL] solved by X-ray diffraction. On the whole, the experimental evidence and computer simulation data favour a transport inhibition mechanism in which flavonoids and GLUT4 interact directly, rather than by a mechanism related to protein-tyrosine kinase and insulin signalling inhibition. Furthermore, the results suggest that GLUT transporters are involved in flavonoid incorporation into cells.
Beyond Bush: The future of popular movements & US Africa policy
Authors: Martin William G. 
Published: March 2005

This article reviews US policy towards Africa, arguing that continuity over the past four administrations far outweighs differences between presidential candidate Senator Kerry and President George Bush. If Kerry were to win the Presidential elections in November, this would not lead to any radical change in US-Africa relations. What is new over the longer term, and is posed so starkly by Bush's unilateralist and militarised actions, is the relentless development of a post-liberal world order and policy agenda. Opposition to this agenda by progressive movements and organisations focused upon such issues as debt cancellation, privatisation, and public health has already born fruit in Africa and elsewhere. The successes, failures, and contradictions of these new campaigns and organisations reflect the post-liberal conditions they work under, and are thus significantly different from the solidarity struggles of the past.
The assistance of Jim Cason was invaluable in constructing this essay, for which the author is very grateful.
Recombinant human granulocyte‐macrophage colony‐stimulating factor plus erythropoietin for the treatment of cytopenias in patients with myelodysplastic syndromes
Authors: Stasi Roberto , Pagano Adalberto , Terzoli Edmondo , Amadori Sergio 
Published: March 2005

In vitro studies have indicated that granulocyte‐macrophage colony‐stimulating factor (GM‐CSF) synergizes with erythropoietin (EPO) for the production of erythroid precursors in patients with myelodysplastic syndrome (MDS). We performed a clinical trial to evaluate whether the combination of these growth factors was effective in relieving the cytopenias associated with MDS. 31 anaemic patients with low and intermediate‐risk primary MDS were enrolled in a 12‐week study. Therapy was initiated with GM‐CSF at 1 μg/kg/d.s.c., and then adjusted to either normalize or double the absolute neutrophil count. EPO was given subcutaneously on alternate days starting from day 2. The EPO dose was initiated at 150 U/kg and increased to 300 U/kg if after 6 weeks there was no or suboptimal erythroid response. 26 patients completed the study treatment. All evaluable cases had a neutrophil response. Clinically significant erythroid responses with increases of haemoglobin levels of at least 1 g/dl and/or reduction of transfusion needs were seen in 9/26 (34.6%), five patients improving their response after dose escalation of EPO. Treatment had no apparent effect on mean platelet counts, a single case displaying a trilineage response. An elevated bone marrow erythroid infiltration and low concentrations of circulating tumour necrosis factor‐α were the only predictors of haemoglobin response both in univariate and in multivariate analysis.We conclude that the combination GM‐CSF + EPO can abrogate neutropenia and substantially relieve transfusion requirements in a large proportion of patients with low and intermediate risk MDS. However, in vivo synergy between these growth factors for the production of erythroid precursors is not supported by our data.
The prevalence of the cysteine1584 variant of von Willebrand factor is increased in type 1 von Willebrand disease: co‐segregation with increased susceptibility to ADAMTS13 proteolysis but not clinical phenotype
Authors: Bowen Derrick J. , Collins Peter W. , Lester Will , Cumming Anthony M. , Keeney Steven , Grundy Pamela , Enayat Saad M. , Bolton‐Maggs Paula H. B. , Keeling David M. , Khair Kate , Campbell Tait R. , Wilde Jonathon T. , John Pasi K. , Hill Frank G. H. ,   on behalf of UK Haemophilia Centre Doctors’ Organization
Published: March 2005

SummaryThe molecular pathogenesis of type 1 von Willebrand disease (VWD) is uncertain in most patients. We examined 30 type 1 VWD families in the UK Haemophilia Centre Doctors’ Organization study. Heterozygosity for Y/C1584 was present in eight of 30 (27%) families and 19 of 76 (25%) individuals with type 1 VWD recruited into the study. Eighteen (95%) of these 19 individuals were blood group O. C1584 did not co‐segregate with VWD in four families, and co‐segregated in one family; the results were equivocal in three families. In all families increased susceptibility of von Willebrand factor (VWF) to a disintegrin and metalloprotease with thrombospondin motifs (ADAMTS) 13 proteolysis co‐segregated with C1584 in affected and unaffected individuals. These data show that C1584, associated with blood group O, is prevalent among patients with type 1 VWD but not necessarily causative of disease and should not be used in isolation to diagnose VWD. Increased susceptibility of C1584 VWF to ADAMTS13 proteolysis may be physiologically significant and increase an individual's risk of bleeding and presenting with VWD.
Oestrogen Requires the Insulin‐like Growth Factor‐I Receptor for Stimulation of Prolactin Synthesis via Mitogen‐Activated Protein Kinase
Authors: Arroba A. I. , Frago L. M. , Argente J. , Chowen J. A. 
Published: March 2005

AbstractSex steroids and growth factors interact at the intracellular level in a variety of tissues to control numerous physiological functions. Oestrogen is known to stimulate prolactin synthesis and secretion, but the effect of insulin‐like growth factor (IGF)‐I is less clear. We used GH3 cells, a somatolactotroph cell line, to study the interaction of 17β‐oestradiol (E2) and IGF‐I on prolactin protein levels and the intracellular mechanisms involved. Cell cultures were treated with E2 (10 nM) and/or IGF‐I (10 ng/ml) for 8 h. The real‐time reverse transcriptase‐polymerase chain reaction, Western blot and enzyme‐immunoassay were used to determine changes in prolactin mRNA and protein levels. At this time‐point, there were no significant changes in cell number, prolactin mRNA expression, or the amount of secreted prolactin. However, E2 increased intracellular prolactin concentrations. IGF‐I alone had no effect, but blocked the stimulatory effect of E2. MAPK (ERK1/2) activation, as determined by Western blot analysis, increased with both E2 and IGF‐I, but not with the combination of these factors. The MAPK inhibitor PD98059 blocked the ability of E2 to increase intracellular prolactin concentrations. Similarly, the IGF‐I receptor antagonist, JB1, blocked the effect of E2 on prolactin synthesis and MAPK activation, as did the oestrogen receptor antagonist ICI182 780. These results suggest that, to stimulate prolactin synthesis, E2 activates the MAPK cascade and that this requires the presence of both oestrogen and IGF‐I receptors.
Mycosis fungoides – a retrospective study of 40 cases in Hong Kong
Authors: Lap‐shing Ku , Kuen‐kong Lo 
Published: March 2005

AbstractBackground  Mycosis fungoides is rare in Hong Kong and oriental data on the disease are lacking.Methods  This is a multiclinic, 13‐year, retrospective study to determine the clinicopathologic characteristics, treatment, and disease outcomes of 40 patients with mycosis fungoides/Sézary syndrome seen in the Social Hygiene Service, Hong Kong.Results  There were 27 males and 13 females with a mean age at diagnosis of 56.4 years. Based on figures in the Social Hygiene Service alone, the incidence in Hong Kong was estimated to be 0.044 per 100,000. Eighty‐five per cent of patients presented with skin‐limited disease, and pruritus was absent in 40% of patients. A mean of 1.48 biopsies was needed to establish the diagnosis, and only 58% of the mycosis fungoides skin biopsies were reported to be histologically diagnostic of the disease. Atypical lymphocytes, epidermotropism, interface changes, and Pautrier's microabscesses were the four most frequently encountered features that attained statistical significance. Sixty‐nine per cent of patients were treated with psoralen–UVA as their initial therapy, and the complete response and relapse rates were 78.3% and 66.6%, respectively. Disease progression to more advanced stages was only seen in 15% of patients. The 5‐year survival rates for the whole group and for stage IA and IB patients were 88.8% and 100%, respectively.Conclusion  Mycosis fungoides is rare amongst Hong Kong Chinese and the majority present with skin‐limited nonprogressive disease.
Fitting Regional Income Distributions in the European Union*
Authors: Pittau Maria Grazia 
Published: March 2005

AbstractCross‐sectional distribution of per capita (log) GDP across the European Union regions from 1977 to 1996 is analysed. Kernel density estimates reveal a multimodal structure of the distribution during the 1970s and early 1980s, and a tendency towards unimodality since the mid‐1980s. The distribution is further analysed by a mixture of normal densities. A two well‐separated component mixture fits the distributions in the 1970s and early 1980s. These two clusters tend to converge, supporting the idea of a process of catching up. In the mid‐1990s, a small group of very rich regions is generated by a separated component.