bluuebunny/crossref_metadata_2025_split huggingface dataset:
Plasmid instability kinetics of the yeast S288C pUCKm8 [cir+] in non‐selective and selective media
Authors: Schwartz L. S. , Jansen N. B. , Ho N. W. Y. , Tsao G. T. 
Published: December 2004

AbstractPlasmid loss kinetics for Saccharomyces cerevisiae transformed with the 2‐μm DNA‐based‐plasmid pUCKm8 were measured in nonselective and selective media. The plasmid pUCKm8 gives the organism two new phenotypes: resistance to the wide spectrum antibiotic G418 sulfate, and the ability to produce the enzyme, β‐lactamase. Plasmid stability was determined using the production of β‐lactamase as a marker. The effect of G418 on the growth rates of all organisms present in the culture and on plasmid stability was also determined. Mathematical models describing plasmid loss kinetics during exponential growth for both nonselective and selective conditions are used to simulate the experimental data. In nonselective medium, over 80% of the cells still exhibited the desired phenotype after 50 doublings. In medium containing G418, improvements in plasmid stability were only marginal due to the appearance of antibiotic‐resistant cells.
Variation in cellulase‐constituting components from Trichoderma reesei with agitation intensity
Authors: Mukataka Sukekuni , Kobayashi Nobuyuki , Sato Seigo , Takahashi Joji 
Published: December 2004

AbstractCellulase producing activity of Trichoderma reesei QM9414 was examined under various agitation intensities and at the dissolved oxygen concentration above 3 ppm. The producing activity greatly depended upon the agitation intensity, and the dependence on the agitation was different for each cellulase‐constituting component. The maximum producing activities of FPA, CM Case, and β‐glucosidase were obtained under different agitation conditions, 1.0, 0.7, and 1.4 m/s in tip velocity, respectively. Intensive agitation brought about remarkable reduction in all cellulase components. The mycelial transformation through agitation intensity was also observed. Comparatively mild agitation of 0.3–1.0 m/s caused pellet formation as the culture progressed, although the pelletization was delayed with increasing agitation intensity. The behavior of the pelletization did not occur at 1.3 and 1.7 m/s throughout the course of cultivation, and under the latter agitation condition hyphae were broken up into short fragments. The cellulase producing activity is discussed in relation to such morphological changes.
Substrate and energy costs of the production of exocellular enzymes by Bacillus licheniformis
Authors: Frankena J. , Van Verseveld H. W. , Stouthamer A. H. 
Published: December 2004

AbstractSubstrate and energy costs of the production of exocellular enzymes from glucose and citrate by B. Iicheniformis S1684 as well as molar growth yields corrected for these costs of product formation were calculated using data from chemostat experiments. The calculations showed that 1.46–1.73 mol glucose and 2.31–2.77 mol citrate are needed for formation and excretion of 1 mol protein. Consequently, the values of the maximal product yield from substrate (Ypsm' g/mol) are 80 < Ypsm < 95 when product is formed from glucose and 50 < Ypsm < 60 when product is formed from citrate. The higher substrate costs for product formation from citrate are due to a higher level of CO2 production during protein formation and a higher substrate requirement for the energy supply of product formation and excretion than when product is formed from glucose. The theoretical ATP requirement for protein synthesis could be determined reasonably well, but the energy costs of protein excretion could not be determined exactly. The energy costs of protein formation are higher than those of biomass formation or protein excretion. Molar growth yields corrected for the substrate costs of product formation were high, indicating a high efficiency of growth.Growth and production parameters were determined as well from experimental data of recycling fermentor experiments using a parameter optimization procedure based on a mathematical model describing biomass growth as a linear function of the substrate consumption rate and the rate of product formation as a linear function of biomass growth rate. The fitting procedure yielded two growth and production domains during glucose limitation. In the first domain the values for the maximal growth yield and maintenance coefficient were in agreement with those found in chemostat experiments at corresponding values of Yspm. Domain 2 could be described best with linear growth and product formation. In domain 2 the rate of product formation decreased and more substrate became available for biomass formation. As a consequence the specific growth rate increased in the shift from domain 1 to 2. Domain 2 behavior most probably is caused by the rel‐status of B. Iicheniformis S1684.
Continuous fermentation of feed streams containing D‐glucose and D‐xylose in a two‐stage process utilizing immobilized Saccharomyces cerevisiae and Pachysolen tannophilus
Authors: Slininger P. J. , Bothast R. J. 
Published: December 2004

AbstractIn the U.S., forest and crop residues contain enough glucose and xylose to supply 10 times the country's usage of ethanol and ethylene, but an efficient fermentation scheme is lacking,1,2,3 To develop a strategy for process design, specific ethanol productivities and yields of Pachysolen tannophilus NRRL Y‐2460 and Saccharomyces cerevisiae NRRL Y‐2235 were compared. Batch cultures and continuous stirred reactors (CSTR) loaded with immobilized cells were fed glucose and xylose. As expected from previous reports, Y‐2235 fermented glucose but not xylose. Y‐2460 consumed both sugars but fermented glucose inefficiently relative to Y‐2235, and it suffered a diauxic lag lasting 10‐20 h when given a sugar mixture. Immobilized Y‐2235 exhibited increasing productivity but constant yield with in creasing glucose concentration. In contrast, Y‐2460 exhibited an optimum productivity at 30–40 g/L xylose and a declining yield with increasing xylose concentration. Immobilized Y‐2235 tolerated more than 100 g/L ethanol while the productivity and yield of Y‐2460 fell by 80 and 58%, respectively, as ethanol reached 50 g/L. A 38.8‐g/L ethanol stream could be produced as 103 g/L xylose was continuously fed to Y‐2460. If it was blended with a 274 g/L glucose stream to give a composite of 23.7 g/L ethanol and 107 g/L glucose, Y‐2235 could en rich the ethanol to 75 g/L. Taken together these results suggest use of a two‐stage continuous reactor for pro cessing xylose and glucose from lignocellulose. An immobilized Y‐2460 CSTR (or cascade) would convert the hemicellulose hydrolyzate. Then downstream, an immobilized Y‐2235 plug flow reactor would enrich the hemicellulose‐derived ethanol to more than 70 g/L upon addition of cellulose hydrolyzate.
Estimation of intracellular sugar phosphate concentrations in Saccharomyces cerevisiae using 31P nuclear magnetic resonance spectroscopy
Authors: Shanks Jacqueline Vanni , Bailey James E. 
Published: December 2004

AbstractA systematic procedure has been formulated for estimating the relative intracellular concentrations of sugar phosphates in Saccharomyces cerevisiae based upon 31P nuclear magnetic resonance (NMR) measurements. The sugar phosphate region of the 31P NMR spectrum is first decomposed by computer analysis, and the decomposition consistency and identification of individual sugar phosphate resonances are established based on in vitro chemical shift calibrations determined in separate experiments. Numerous evaluations of intracellular S. cerevisiae compositions for different strains and different cell environments provide the basis for in vivocorrelations of inorganic phosphate chemical shift with the chemical shifts of 3‐phosphoglycerate, beta;‐fructose 1,6‐diphosphate, fructose 6‐phosphate, and glucose 6 phosphate. Relative intracellular sugar phosphate concentrations are obtained by correcting peak areas for partial saturation during transient in vivo experiments. In vivo concentrations estimated by this method agree well with estimates for similar systems based on other techniques. This approach does not require costly la belled compounds, and has the advantage that other important metabolic state variables such‐as internal and external pH and intracellular levels of phosphate, ATP, ADP, NAD(H), and polyphosphate may be determined from the same 31P spectrum. Extension of this strategy to other cellular systems should be straightforward.
Experimental multivariable adaptive optimization of the steady‐state cellular productivity of a continuous baker's yeast culture
Authors: Semomes Gary B. , Lim Henry C. 
Published: December 2004

AbstractA multivariable adaptive optimization algorithm that uses transient data to improve the optimization speed was successfully implemented on‐line to maximize the steady‐state cellular productivity of a continuous culture of baker's yeast. The algorithm was shown to be stable even during periods of oscillatory growth and was able to reoptimize the culture when planned disturbances were introduced. Although adaptive tuning of the forgetting factor improved the performance, further refinements in the adaptive forgetting factor algorithm are necessary for completely satisfactory results.
Protein purification by affinity binding to unilamellar vesicles
Authors: Powers Johnny D. , Kilpatrick Peter K. , Carbonell Ruben G. 
Published: December 2004

AbstractA novel purification technique is proposed which employs affinity‐ligand‐modified liposomes to specifically purify bioactive macromolecules from solution. This process is demonstrated with avidin as the model biomolecule and biotin as the affinity ligand. Biotin is covalently bound to the surface of small unilamellar vesicles composed of dimyristoyl phosphatidylcholine (DMPC) and dimyristoyl phosphatidylethanolamine (DMPE). The number of accessible binding sites on the liposomes is determined by titration with avidin, and the kinetics of binding are evaluated by monitoring the concentration of free avidin in solution after the addition of biotinylated liposomes. The specificity of the process is determined by following the affinity binding of avidin to biotinylated liposomes in the presence of model impurities (i.e., lysozyme and cytochrome C). Liposome‐bound avidin is separated from the impurities by ultrafiltration through a membrane which retains the liposomes.
A heuristic approach for the economic optimization of a series of CSTR's performing michaelis–menten reactions
Authors: Malcata F. Xavier 
Published: December 2004

AbstractThe estimation of the size of each reactor of a series of CSTR's performing a Michaelis–Menten reaction in the liquid phase can be obtained to advantage via an optimization technique leading to the minimum overall capital cost. The cost scaleup is assumed to be described by a power rule on the equipment capacity. Various contributions are lumped into the exponent, thus leading to values above unity. The analytical development leading to the optimal intermediate concentrations of substrate according to the foregoing criterion is presented. A short‐cut method based on an empirical expression that approximates the numerical solution is reported. This correlation is found to be exact at the asymptotic behaviors, and to give accurate results within an acceptable error level for the range with physical interest. Therefore, it is particularly useful during the predesign steps of equipment for the biochemical industry.
New sensor allowing continuous water activity measurement of submerged or solid‐substrate fermentations
Authors: Gervais P. 
Published: December 2004

AbstractWater activity of the substrate is an important and acute factor for growth as well as for metabolic production of microorganisms or for biocatalyst systems. A sensor has been designed in order to control this parameter on‐line during submerged and solid‐substrate fermentations. This sterilizable sensor allows the measurement of the relative humidity of the atmosphere in a small chamber by means of a capacitive element separated from the medium by a thin ethylenepolytetrafluoride membrane. For high aw values (>0.90) a sequential circulation of a dried gas prevents the sensor saturation. Measurements are rapid and accurate, and control of the water activity of a fermentation medium has been carried out for 5 days using this sensor.
A bio‐mimetic cadmium adsorbent: Design, synthesis, and characterization
Authors: Yin John , Blanch Harvey W. 
Published: December 2004

AbstractA cadmium‐binding adsorbent has been designed based on insights gained from the study of cadmium‐binding molecules isolated from living organisms. Cadmium‐chelating thiolate groups—common in proteins that bind cadmium—have been covalently attached to a commercially available ion‐exchange resin. The new adsorbent exhibits a favorable affinity (2 × 10−10M), selectivity (25‐fold greater affinity for Cd2+ than Zn2+), and capacity (1.4 mmol Cd2+/g dry resin) for cadmium ion. Adsorbed Cd2+ may be released with 20mM pyro‐phosphate at pH 2. The adsorbent also recovers Cd2+ in the presence of NH4CI and KCN.The apparent adsorption rate at pH 7.0 (2M−1 min−1) increases 30‐fold as pH is increased to 11. The rate dependence on pH may be due to the inhibition of adsorbent‐metal association by intramolecular hydrogen bonding at neutral pH.
Kinetic aspects of the bioconversion of L‐tyrosine into L‐DOPA by cells of Mucuna pruriensL. Entrapped in different matrices
Authors: Pras Niesko , Hesselink Paul G. M. , ten Tusscher Jan , Malingré Theo M. 
Published: December 2004

AbstractPlant cells of Mucuna pruriens L. entrapped In calcium alginate, calcium pectinate, agarose, or gelatine were able to convert L‐tyrosine to L‐DOPA, which was released Into the medium. Michaelis–Menten kinetics could be applied on the entrapped cells, based on the measurement of initial rates of L‐DOPA production. The calculated apparent affinity constants were comparable with the affinity constants obtained with enzyme preparations. Comparison of the apparent maximum rate of bioconversion of the entrapped cells and the maximum rate of bioconversion of a derived cell homogenate indicated that the systems were not operating optimally. Measurement of the effective diffusion coefficients of L‐tyrosine pointed out that this substrate could diffuse freely into the matrices. From the initial rates of bioconversion and the effective diffusion coefficients, the observable modulus was calculated for each system. The obtained values confirmed that the diffusional supply rate of L‐tyrosine was not the limiting factor. For oxygen, which was utilized for byconversion as well as for cell respiration, the calculated observable moduli was directed toward strong oxygen transfer limitations. The values found for the oxygen consumption indicated that the entrapped cells remained partly or totally viable in the four matrices tested. Based on the highest viability and the highest rates of bioconversion, it was concluded that alginate‐entrapped cells of M. pruriens formed the most suitable biocatalytic system for the production of L‐DOPA from L‐tyrosinre.
Cell mass synthesis and degradation by immobilized Escherichia coli
Authors: Karel Steven F. , Robertson Channing R. 
Published: December 2004

AbstractEscherichia coli K‐12 cells were grown in a confined volume using microporous hollow fiber membranes. The local cell concentrations in the reactors were above 400 g dry mass/L, in excess of the predicted limit based on the specific volume of free cells determined by tracer exclusion. Cell mass synthesis and degradation rates in these reactors were measured using radioisotope labeling with 35S. Net accumulation of cell material persisted at these high cell densities. The rates of substrate uptake and cell growth were predicted from the theory of reaction and diffusion assuming that kinetics of cell metabolism are identical for free‐living and immobilized cells. This theory was tested by comparison of overall rates and by the size of the region in which cell growth occurred, measured by autoradiography. A yield coefficient of 4 ± 1 mol sulfur/mol glucose was measured, in agreement with the value determined for free‐living cells in similar conditions. Cell growth occurs in a thin layer (10–30 μm), at a rate similar to the growth rate for free cells. Volume expansion by the cells as a consequence of proliferation induces convection of cell mass out of the growth region into a region of the reactor filled with starving cells, which then accumulate in the reactor. The combination of mass‐balance and spatial distribution measurements made possible by the use of radioisotope labeling enables a direct test for mass transfer limitations, the determination of the intrinsic cell kinetics, and noninvasive measurements of cell growth in immobilized cell reactors.
Compartment model concept used in the design of fermentation with recombinant microorganisms
Authors: Nielsen J. , Emborg C. , Halberg K. , Villadsen J. 
Published: December 2004

AbstractSimple models are of great value in order to simulate cell dynamics, and a class of potentially useful models are the so‐called compartment models, which are a simple extension of the unstructured approach. Compartment models have earlier been used with great success to describe cell dynamics. In this article the authors present a 4‐compartment model that is able to describe a majority of the observations that have been made through fermentations with recombinant microorganisms. The compartment model is especially suited for description of dynamic changes in plasmid copy number, e.g., runaway replication. Simulation of batch fermentation with runaway conditions corresponds very well with experimental results. Finally the model is used in the design of an optimal fermentation strategy in a CSTR.
Graphical determination of mean activation energy and standard deviation in a microheterogeneity model of enzyme deactivation
Authors: Henley James P. , Sadana Ajit 
Published: December 2004

AbstractTraditionally, enzyme populations have been treated as if they were either homogenous, or heterogeneous with distinct and separable subpopulations. The microheterogeneity model, however, assumes that there is a continuous distribution of properties in the population. In the area of enzyme deactivation kinetics, this model describes the heterogeneous population as having a continuous distribution of activation energy of deactivation. This distribution is characterized by mean activation energy, and a standard deviation of activation energy. The microheterogeneity model contains two parameters, ϵ0 and σ. Parameter ϵ0 is the mean value of ϵ for a heterogeneous enzyme population; ϵ is the activation energy divided by absolute temperature and the ideal gas constant. Parameter σ is the standard deviation of the Gaussian distribution of ϵ values in the population. If the population is homogeneous, then ϵ = ϵ0 for all enzyme molecules and σ = 0. There are certain ratios which are independent of ϵ0 and dependent upon σ. Two important ratios are t1/4/t1/2 and t1/2/t, where t represents t1/2 for a homogeneous enzyme population with the same mean ϵ (ϵ0), as the heterogeneous population. If there is experimental deactivation data for the heterogeneous population which is well behaved, the first ratio, t1/4/t1/2, can be determined by estimating the time in minutes at which the enzyme has lost 25% of its activity (t1/4), and the time in minutes at which the enzyme has lost 50% of its activity (t1/2), and then taking the ratio t1/4/t1/2. The corresponding value of σ can be estimated from a graph. The ratio t1/2/t can be found directly as a function of t1/4/t1/2, and can be estimated from another graph. The value of ϵ0 can then be calculated from the formulasgiven in the article.
Determination of protein expression and plasmid copy number from cloned genes in Escherichia coli by flow injection analysis using an enzyme indicator vector
Authors: Schendel F. J. , Baude E. J. , Flickinger M. C. 
Published: December 2004

AbstractOn‐line determination of expression rates from cloned genes in Escherichia coli and of plasmid copy number would be useful for monitoring accumulation of non‐secreted proteins. As an initial model for monitoring gene expression in intact cells, a non‐gene‐fusion enzyme‐based indicator plasmid has been constructed containing the phoA gene coding for alkaline phosphatase (AP) in pUCIS and pACYC184. The activity of AP can be rapidly determined in permeabilized cells. A flow injection analysis (FIA) assay has been developed which allows the direct real‐time measurement of the AP activity during cell growth. A model target gene coding for E. coli cyanase (cynS) has been inserted in order to determine the ratio between the expression of the target and indicator, AP. A linear relationship has been found between plasmid copy number and AP activity for the high‐copy pUC vector. To minimize indicator expression, transcription terminators have been inserted between the cynS and phoA genes, altering the target‐to‐indicator ratio by 10‐ to 40‐fold. These vectors may be useful for the rapid continuous determination of plasmid copy number and target gene expression for nonsecreted proteins and would overcome the limitations of in situ probe biosensors for real‐time determination of the accumulation of proteins from cloned genes in E. coli.
How suspension cultures of Catharanthus roseus respond to oxygen limitation: Small‐scale tests with applications to large‐scale cultures
Authors: Snape J. B. , Thomas N. H. , Callow J. A. 
Published: December 2004

AbstractThe effect of oxygen supply on the growth of suspension cultures of Catharanthus roseus in Erlenmeyer flasks was investigated. Below a critical oxygen supply rate the culture could not survive. By increasing the oxygen supply, a point is reached where the culture survives but no growth is possible. At higher oxygen supply rates there is a regime where both growth rate and the maximum biomass concentration increase with oxygen supply. Eventually there comes a point where no further increase in biomass is achieved, probably due to the depletion of the sugars; however, the growth rate continues to increase with oxygen supply until a maximum growth rate is obtained. The ratio of fresh to dry weight at maximum fresh weight increased with shaker table speed of rotation accompanied by a greater rate of sugar depletion.
Cell adsorption and local accumulation of extracellular polysaccharide in an immobilized Acinetobacter calcoaceticus system
Authors: Wang Sy‐Dar , Wang Daniel I. C. 
Published: December 2004

AbstractAcinetobacter calcoaceticus can be immobilized on Celite by adsorption. The salt concentrations suitable for immobilized cell fermentation are between 10 and 50 mM phosphate concentration. Low salt concentrations cause desorption of immobilized cells while high salt concentrations inhibit the adsorption of cells on Celite. It is also found that cell adsorption is better at lower pH than at higher pH. An airlift fermentation using immobilized cells at 300 g/L Celite loading shows that about 70% of the total polymer produced is accumulated in Celite pores at a concentration (15.4 g/L) almost threefold higher than that in the bulk liquid (5.7 g/L).