bluuebunny/crossref_metadata_2025_split huggingface dataset:
Reductive biotransformations of organic compounds by saccharomyces cerevisiae: Study of oxidoreductases involved using electrophoretic zymograms
Authors: Young C. S. , Ward O. P. 
Published: December 2004

AbstractThe role of oxidoreductases in reduction of carbonyl compounds was investigated by application of zymogram techniques. Eight bands were observed using ethanol with nicotinamide adenine dinucleotide (NAD) as coenzyme. Bands observed with lactic acid and (R)‐(‐)‐phenyl‐1,2‐ethanediol with nicotinamide adenine dinucleotide phosphate (NADP) had similar Rm values. 2‐Hydroxyvalerate and malate manifested bands having similar Rm values and were active with both NAD and NADP. Based on their structural similarity and identical Rm values, oxidation of 1,4‐cyclooctanediol (band #2) and cis‐1,5‐cyclooctanediol may be due to a common enzyme. The PAGE‐zymogram technique may be used on a preparative scale to facilitate purification and full characterization on the observed stained bands.
Continuous photoautotrophic cultures of the eukaryotic alga chlorella vulgaris can exhibit stable oscillatory dynamics
Authors: Javanmardian Minoo , Palsson Bernhard O. 
Published: December 2004

AbstractSustained oscillations in cell concentration, average per cell DNA content, and average cell size were found in continuous photoautotrophic cultures of Chlorella vulgaris at low dilution rates (0.1/day). The period of oscillation was approximately 10 days. DNA histograms determined by flow cytometry exhibited reproducible pattern through consecutive oscillations. At the maximum cell concentration during an oscillation, the DNA histograms showed that the majority of the cells were not replicating their chromosomes, and most of the culture was comprised of single cells in G0/G1 phase. The cells then initiated DNA replication; however, because of the long generation time, the cell concentration decreased to a minimum, and at the same time the average per cell DNA content reached its maximum value. At this point the cells began to divide, and the cell concentration increased until it reached its maximum value at the beginning of the next oscillation. Calculations based on the supplied nutrients and comparison to biomass generation showed that the oscillatory behavior in continuous photoautotrophic cultures of C. vulgaris was not due to nutrient limitation, but most likely was due to the secretion of compounds that alter cell cycle kinetics. The oscillatory behavior disappeared when the dilution rate was increased to 0.3/day and the culture reached a stable steady state.
Application of empirical design methodologies to the study of the influence of reaction conditions and N‐α‐protecting group structure on the enzymatic X‐Phe‐Leu‐NH2 dipeptide synthesis in buffer/dimethylformamide solvents systems
Authors: Calvet S. , Clapés P. , Vigo J. P. , Xaus N. , Jorba X. , Mas R. M. , Torres J. L. , Valencia G. , Serralheiro M. L. , Cabral J. M. S. , Empis J. M. A. 
Published: December 2004

AbstractThe influence of five different N‐terminal protecting groups (For, Ac, Boc, Z, and Fmoc) and reaction conditions (temperature and dimethylformamide content) on the α‐chymotrypsin‐catalyzed synthesis of the dipeptide derivative X‐Phe‐Leu‐NH2 was studied. Groups such as For, Ac, Boc, and Z always rendered good peptide yields (82% to 85%) at low reaction temperatures and DMF concentrations, which depended on the N‐α protection choice. Boc and Z were the most reactive N‐α groups and, in addition, the most suitable for peptide synthesis. On the other hand, the use of empirical design methodologies allowed, with minimal experimentation and by multiple regression, to deduce an equation, which correlates the logarithm of the first order kinetic constant (log k') with reaction temperature, DMF concentration, and hydrophobicity (log P values) of the different protecting groups. The predictive value of the equation was tested by comparing the performance of another protective group, such as Aloc, with the performance predicted by said equation. Experimental and calculated k' values were found to be in good agreement.
Quantification of cell culture factors affecting recombinant protein yields in baculovirus‐infected insect cells
Authors: Lindsay David A. , Betenbaugh Michael J. 
Published: December 2004

AbstractAn experimental study was undertaken to quantify the effects of infection cell density, medium condition, and surface aeration on recombinant protein yields in insect cells. In the absence of surface aeration and fresh medium, insect cells generated higher product yields (on a per cell basis) when infected with recombinant baculovirus at low cell densities, LCD (3 × 105−4 × 105 cells/mL), than at high cell densities, HCD (>0.9 × 106 cells/mL), for two distinct baculovirus types. Surface aeration of a HCD culture infected in spent medium improved β‐glactosidase yields 5‐fold over the nonaerated case. Surface aeration and medium replenishment improved β‐galactosidase yields of a HCD culture by 20‐fold (compared to a 1.6‐fold improvement for a LCD culture), resulting in cultures with productivties that were independent of the cell density at infection.
Ultrafiltration membranes for chemical bonding of urease
Authors: Schmidt‐Steffen Alfred , Staude Eberhard 
Published: December 2004

AbstractAsymmetric ultrafiltration membranes suitable for covalent bonding of urease can be prepared from membranes based upon polyamide or polysulfone. Because the original membrane polymers are not chemically reactive, they have to be converted in such a way that known reactions for enzyme fixation can be used such as the diazo, the acyl‐acid, the carbodiimide, and the methylbromide reaction. The enzyme was fixed within the porous substructure of the membrane. The amount of enzyme immobilized at the membrane dense skin was found to be negligible. The kinetics can be described according to the Michaelis–Menten model. Compared to the native urease, the activity of the membrane‐bonded enzyme was very low. The reasons can be sought in the transmembrane transport and in the fixation procedure as well.
Development of a multienzyme reactor for dopamine synthesis: I. Enzymology and kinetics
Authors: Anderson William A. , Moo‐Young Murray , Legge Raymond L. 
Published: December 2004

AbstractThe enzymology and kinetics of tyrosine phenol lyase (TPL) from Erwinia herbicola, and tyrosine decarboxylase (TDC) from Streptococcus faecalis have been investigated for potential use in a coimmobilized multienzyme biocatalytic system for the production of dopamine. In this multienzyme biotransformation using whole cells optimized for each of the respective enzymes, TPL catalyzes the production of 3,4‐dihydroxyphenyl‐L‐alanine (L‐dopa) from catechol, pyruvate, and ammonium, and this is subsequently decarboxylated by TDC to produce dopamine. Performing the reactions simultaneously, thereby removing L‐dopa, is one option for overcoming the TPL equilibrium constraints. The enzymes have different optimal pH values, so the reaction kinetics at a compromise pH of 7.1, where both enzymes could be operated simultaneously, were investigated. For the concentration range investigated, TPL followed pseudo‐first‐order kinetics with respect to catechol, pyruvate, and ammonium. TDC exhibited significant product inhibition as well as inhibition by combinations of catechol and pyruvate.
Protein extraction and activity in reverse micelles of a nonionic detergent
Authors: Ayala Guadalupe A. , Kamat Sanjay , Beckman Eric J. , Russell Alan J. 
Published: December 2004

AbstractWe describe, for the first time, the ability of a polyoxyethylene sorbitan trioleate‐isopropanol microemulsion in hexane to solubilize pure proteins. The dependences of cytochrome c extraction and buffer solubilization by the reverse micellar system on ionic strength of the aqueous phase, detergent concentration, and cosurfactant concentration result in increased extraction. In addition, subtilisin (a serine protease) is shown to be active in this microemulsion. Further the activity of the enzyme can be regulated by the water content of the micelles, enabling control of enzyme activity by “solvent engineering.”
Analysis of mammalian viable cell biomass based on cellular ATP
Authors: Sonderhoff Stefan A. , Kilburn Douglas G. , Piret James M. 
Published: December 2004

AbstractAnalysis of cellular ATP as a means of measuring viable biomass loading was investigated in hybridoma cell culture. ATP analysis by the luciferin–luciferase assay was compared with trypan blue‐stained hemocytometer counts. The cell‐specific ATP content varied between 2 and 6 fmol per viable cell over a batch culture. ATP levels were highest during exponential growth, and decreased during the stationary and decline phases. Electronic counting and volume measurements were performed to assay the viable cell biomass. Cell sorting, using fluorescein diacetate, was used to separate viable and nonviable cells in cultures with between 35% and 90% viable cells. Viable cells contained over 2 orders of magnitude greater cell‐specific ATP than nonviable cells. Cell‐specific ATP correlated directly with the viable cell volume rather than viable cell numbers. Over the range of batch culture conditions, ATP analysis should provide a more accurate measurement of hybridoma viable biomass than hemocytometer counts.
Oxygen transfer modeling and simulations for an industrial continuous airlift fermentor
Authors: Pigache S. , Dhoms P. , Trystram G. 
Published: December 2004

AbstractThis article deals with the modeling of the oxygen transfer in an industrial airlift fermentor used for lactic yeast production on whey substrates. The purpose of this study was to improve the understanding of the interactions among the various parameters that govern the oxygen transfer phenomena in this type of fermentor. The reliability of the proposed model is demonstrated. The results of the investigations have been put into practice on the industrial scale and have contributed to monitor better the fermentation process. The model was also used to develop new ways of industrial fermentor design.
Production of a discrete, heterogeneous population of β‐galactosidase polypeptides using baculovirus expression vectors
Authors: Licari P. , Bailey J. E. 
Published: December 2004

AbstractGel electrophoresis analysis of immunoprecipitated β‐galactosidase and polyhedrin‐β‐galactosidase expressed in Spodoptera frugiperda cells infected with recombinant Autograph californica nuclear polyhedrosis virus revealed the existence of a population of discrete β‐galactosidase polypeptides. Several of the polypeptides observed in the fusion protein expression experiments exhibit a consistent pattern of slightly greater molecular weight when compared to the nonfusion β‐galactosidase that is compatible with the hypothesis that these fusion protein fragments retain the N‐terminal polyhedrin residues. Pulse‐chase experiments showed that overall β‐galactosidase degradation occurred at a negligible rate compared to the synthesis rate at 96 h postinfection, yet the fragments are observed for short pulse times. Degradation of several different β‐galactosidase polypeptides was observed 24 h postinfection. Ribonucleic acid hybridization analysis of lacZ transcripts shows significant heterogeneity that may result from premature transcription termination. Although a proteolytic origin cannot be excluded, the data assembled suggest that premature termination of transcription or translation is the likely cause for the heterogeneous population of immunoreactive peptides observed. Many discrete forms of β‐galactosidase polypeptides were also observed in studies with Escherichia coli, indicating that production of these heterogeneous forms is not a consequence of heterologous expression of the enzyme.
Classification and measurement of fungal pellets by automated image analysis
Authors: Cox P. W. , Thomas C. R. 
Published: December 2004

AbstractAn automated image analysis method for classifying and measuring pellets of filamentous fungi growing in submerged fermentations has been developed. The method discriminates between pelleted mycelial growth and loose aggregates of dispersed hyphae. Pellets are classified into smooth and hairy types. In both cases, the core of the pellet is identified and its shape and size characterized. For hairy pellets the annular region is also characterized. The method was tested on pellets of Aspergillus niger ATCC 11414 grown in a defined medium in shake flasks. This rapid method makes practical extensive studies on the morphology of pellets in submerged fermentations and the influence of fermentation conditions on that morphology.
Hydrolysis of butteroil by immobilized lipase using a hollow‐fiber reactor: Part II. Uniresponse kinetic studies
Authors: Malcata F. Xavier , Hill Charles G. , Amundson Clyde H. 
Published: December 2004

AbstractA lipase from Aspergillus niger immobilized by adsorption on microporous, polypropylene hollow fibers was used to effect the hydrolysis of the glycerides of melted butterfat at 40°C and pH 7.0. Mcllvane buffer was pumped through the lumen and melted butterfat was pumped courrently through the shell side of a shell‐and‐tube reactor. Nonlinear regression methods were employed to determine the kinetic parameters of three nested rate expressions derived from a Ping Pong Bi Bi enzymatic mechanism coupled with three nested rate expressions for the thermal deactivation of the enzyme. For the reaction conditions used in this research, a four‐parameter rate expression (which includes a two‐parameter deactivation rate expression and a two‐parameter hydrolysis rate expression) is sufficient to model the overall release of free fatty acids from the triglycerides of butterfat as a function of space time and time elapsed after immobilization. At a space time of 3.7 h immediately after immobilization of lipase, 50% of the fatty acid residues esterified in the sn‐1,3 positions of the triglycerides can be released in the hollow‐fiber reactor.
The use of Fab‐masking antigens to enhance the activity of immobilized antibodies
Authors: Velander William H. , Subramanian Anuradha , Madurawe Rapti D. , Orthner Carolyn L. 
Published: December 2004

AbstractThis study demonstrates that masking the Feb regions of a monoclonal antibody (Mab) with synthetic antigens prior to covalent immobilization efficiency. Water‐soluble adducts of poly(2‐methyloxazoline) polymers and a syntheticpeptide epitope for the Mab were constructed. These synthetic antigens are referred to as Fab‐masking antigents (FMAs). The antibody used in this study is a Ca2+‐dependent murine monoclonal lgG directed against the plasma protein, human protein C (hPC). The FMAs were pre‐equilibrated with Mab in the presence of calcium prior to immobilization and were then removed by EDTA, which destabilized the FMA‐Mab complexes. The antigen binding efficiency and accessibility of the Fab domain of the immobilized antibody was significantly increased for Mab immobilized in the presence of FMA relative to those Mab immobilized without FMA. The increase in binding efficiency was most pronounced for the largest FMA employed. No appreciable differences were detected in the avidity of hPC‐Mab complexes formed by immunosorbents produced by either masked or unmaked antibody. These results provide evidence that orientgation may play an important role in the binding activity of immobilized antibodies.
Characterization of initial events in bacterial surface colonization by two Pseudomonas species using image analysis
Authors: Mueller Robert F. , Characklis William G. , Jones Warren L. , Sears John T. 
Published: December 2004

AbstractThe processes leading to bacterial colonization on solidwater interfaces are adsorption, desorption, growth, and erosion. These processes have been measured individually in situ in a flowing system in real time using image analysis. Four different substrata (copper, silicon, 316 stainless‐steel and glass) and 2 different bacterial species (Pseudomonas aeruginosa and Pseudomonas fluorescens) were used in the experiments. The flow was laminar (Re = 1.4) and the shear stress was kept constant during all experiments at 0.75 N m−2. The surface roughness varied among the substrata from 0.002 μm (for silicon) to 0.015 μm (for copper). Surface free energies varied from 25.1 dynes cm−1 for silicon to 31.2 dynes cm−1 for copper. Cell curface hydrophobicity, reported as hydrocarbon partitioning values, ranged from 0.67 for Ps. fluorescens to 0.97 for Ps. aeruginosa.The adsorption rate coefficient varried by as much as a factor of 10 among the combinations of bacterial strain and substratum material, and was positively correlated with surface free energy, the surface roughness of the substratum, and the hydrophobicity of the cells. The probability of desorption decreased with increasing surface free energy and surface roughness of the substratum. Cell growth was inhibited on copper, but replication of cells overlying an initial cell layer was observed with increased exposure time to the cell‐containing bulk water. A mathematical model describing cell accumulation on a substratum is presented.
Enzyme activity and shikonin production in Lithospermum erythrorhizon cell cultures
Authors: Srinivasan Venkatesh , Ryu D. D. Y. 
Published: December 2004

AbstractThe activities of the biosynthetic enzmes phenylalanine ammonia lyase (PAL) and 3‐hydroxy‐3‐methylglutaryl‐CoA‐reductase (HMGR) were measured in cells transferred from growth to production medium in a two‐stage batch culture. It was found that both these enzymes showed transient increases, PAL (three‐ to fourfold) and HMGR (two‐ to four‐fold), at or near the point of exhaustion of nitrogen source (NO3). Production of shikonin derivatives also started at this time. The addition of excess nitrate to the medium shortly before nitrate exhaustion (days 6–8) markedly reduced the final product yield (by 70–80%) while addition of excess nitrate in the later stationary growth phase (days 14–16) had no significant effect. When the production rate of shikonin derivatives was correlated with PAL activity, it was observed that production rate is very low (less than 1 mg/L · day) at low levels of PAL activity (below 0.1 unit/mg protein). Once a threshold level of PAL activity (about 0.15 unit/mg protein) is reached, the biosynthetic rate of shikonin derivatives increases. Such a relationship could not be deduced for HMGR activity. It was concluded that the production of shikonin derivatives may be limited at the phenylalanine deaminating step at low levels of PAL activity.
Formation of polyol–fatty acid esters by lipases in reverse micellar media
Authors: Hayes Douglas G. , Gulari Erdogan 
Published: December 2004

AbstractThe synthesis of polyol–fatty acid esters has strong implications in such industries as foods, cosmetics, and polymers. We have investigated these esterification reactions employing the polyols ethylene glycol, 2‐monoglyceride, and sugars and their dervatives with the biocatalyst lipase in water/AOT/isooctane reverse micellar media. For the first reaction, 50–60% conversion was achieved and product selectivity toward the monoester over the diester shown possible by employing lipase from Rhizopus delemar. A simple kinetic model based on the formation of acyl‐enzyme intermediate accurately predicted the effect of polyol concentration but not the effect of fatty acid or water concentration probably due to the model exclusion of paritioning effects. The success of this reaction in reverse micellar media is due greatly to its capacity to solubilize large quantities of glycol despite the media's overall hydrophobicity. The second reaction, investigated for its potential for production of “mixed” glycerides, also achieved about 50% conversion but had only a small portion of triglyceride in its product distribution. Also, isomerization of the 2‐monoglyceride to 1‐monoglyceride, followed by hydrolysis of the latter, unfortunately occurred to a significant extent. Attempts at esterification with hexoses and their derivatives such as glucose and mannitol produced no convesion.
Anomeric specificity of glucose uptake systems in Lactococcus cremoris, Escherichia coli, and Saccharomyces cerevisiae: Mechanism, kinetics, and lmplications
Authors: Benthin Stig , Nielsen Jens , Villadsen John 
Published: December 2004

AbstractThe mechanism and kinetics of the glucose uptake systems of three representative microorganisms are studied during cultivation in a chemostat. The three microorganisms are Lactococcus cremoris, Escherichia coli, and Saccharomyces cervisiae. Two models describing respectively competitive and independent uptake of the two glucose anomers are tested on experimental data where α‐ and β‐glucose are determined by flow injection analysis after pulse addition of the pure anomers to a chemostat. The very accurate experimental results are used to give a convincingly clear model discrimination for all three microorganisms. The uptake of glucose by S. cervisiae occurs by a competitive mechanism with preference for α‐glucose (Kα = 32 mg/L and Kβ = 48 mg/L). Surprisingly, the glucose uptake by the two bacteria is shown to be mediated by anomer specific transport systems with no competitive inhibition from the other glucose anomer. This novel finding has not been described in the literature on the phosphotransferase system. In L. cremoris the relative uptake rates of the glucose anomers match the equilibrium composition exactly (36% α‐glucose). In E. coli the relative uptake rate of α‐glucose at glucose unlimited growth is 26%, which means preference for β‐glucose. However, the saturation constants of the two sites in E. coli are Kα = 2 mg/L and Kα = 15 mg/L, and a preference for α‐glucose is exhibited at very low glucose concentrations. The results are of considerable improtance in relation to enzyme based on‐line measurements during fermentations as well as to the modeling of glucose limited growth and product formation.
Incapability of Gluconobacter oxydans to produce tartaric acid
Authors: Klasen Ralf , Bringer‐Meyer Staphanie , Sahm Hermann 
Published: December 2004

AbstractThe dependence of tartaric acid production by Gluconobacter oxydans ssp. oxydans ATCC 19357 and G. oxydans ssp. suboxydans ATCC 621 on vanadate was investigated. It was found with both organisms that trataric acid could only be produced in a medium containing vanadate (NH4VO3). A proposed intermediate of the tartaric acid metabolism in G. oxydans, 5‐ketogluconic acid, was tested on its reactivity in the presence of the oxidizing catalyst vanadate. It could be shown that 5‐ketogluconic acid and the catalyst vanadate, but not the activity of G. oxydans, were responsible for the formation of tartaric acid. G. oxydans was not able to produce tartaric acid by itself. The stereochemical identity of the formed tartaric acid could be identified as the L‐(+)‐type. Oxalic acid was formed from 5‐ketogluconic acid with vanadate in the absence and in the presence of G. oxydans. The ratio of oxalic acid to tartaric acid was 1:1.