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  • Life and Medical Sciences  (1,345)
  • Biochemistry and Biotechnology  (589)
  • 1985-1989  (1,934)
  • 1950-1954
  • 1988  (1,934)
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  • 1985-1989  (1,934)
  • 1950-1954
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  • 1
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 31 (1988), S. 160-167 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Six endoglucanases (Endo I, II, III, IV, V, and VI), three exoglucanases (Exo I, II, and III), and a β-glucosidase (β-gluc I) isolated from a commercial cellulase preparation of Trichoderma viride origin were examined as to their activities on xylan ex oat spelts. Endo I, II, and III as well as Exo II and III showed no activity toward xylan and were classified as specific glucanases. Less specificity was found for the endoglucanases Endo IV, V, and VI, Exo I, and β-gluc I, whose enzymes were able to hydrolyze xylan. With respect to product formation these xylanolytic cellulases fit the classification of xylanases generally accepted in the literature. Kinetic experiment with xylan, CM-cellulose, and p-nitrophenyl-β-D-glucoside revealed that Endo IV, V, an VI and Exo I prefer to hydrolyze β-1, 4-D-glucosidic linkages. β-Gluc I showed no clear substrate preference.
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  • 2
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    Biotechnology and Bioengineering 31 (1988), S. 173-178 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
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  • 3
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    Biotechnology and Bioengineering 31 (1988), S. 179-182 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
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  • 4
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    Biotechnology and Bioengineering 31 (1988), S. 895-904 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Native starch granules from wheat have been subjected to enzymatic depolymerization with an α-amylase from Bacillus subtilis. Crystallites made from short-chain amylose and residues from mild acid hydrolysis have been also tested. Electron microscopy, particle size analysis, DSC, and x-ray diffractometry reveal that enzymatic degradation occurs granule by granule. Gel permeation chromatography shows off the macromolecular nature of the remaining material. In contrast, acid erodes simultaneously all the granules, leading to a splitting into small particles. Crystalline fractions are completely degraded by α-amylase. These results support evidence for an active disentanglement of chains, carried out by the different subsites of α-amylase molecules. A simple mathematical treatment is proposed to explain the results of the kinetics.
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  • 5
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    Biotechnology and Bioengineering 31 (1988), S. 958-968 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: The influence of operational, parameters, such as hydraulic retention time, organic loading rate, influent substrate concentration, pH, and temperature, on the performance of the first phase of anaerobic digestion has been investigated. A complex substrate based on beef extract was used, and six series of experimental runs were conducted, each one showing the effect of one operational variable. The predominant fermentation products were always acetic and propionic acid, independent of the values of the operational parameters. For initial COD concentrations and hydraulic retention times above the critical values identified as 3 g/L and 6 h, respectively, the degree of acidification achieved was between 30 and 60%. The degree of acidification was found to increase with the hydraulic retention time and decrease with the influent substrate concentration and organic loading rate, while the opposite held true for the rate of product formation. Furthermore, it has been demonstrated that acidification is primarily determined by the hydraulic retention time and the rate of product formation by the influent substrate concentration. The concentration of the acetic acid produced was found to depend on the operational parameters. However, the concentration of propionic acid produced depended only on the substrate availability with a consistent proportion of 8% initial COD converted to it. The optimum pH and temperature were 7 and 40°C, respectively. The percentage of acetic acid as a proportion of the total volatile fatty acids produced was found to increase with increasing pH and temperature, while the percentage of propionic acid seemed to decrease accordingly. Finally the effect of the temperature on the rate of acidification followed an Arrhenius type equation with an activation energy equal to 4739 cal/mol.
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  • 6
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    Biotechnology and Bioengineering 31 (1988), S. 979-983 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: A new method for determination of the binodial of an aqueous two-phase system, using spectrophotometric measurements of the turbidity, is described in this article. The method is especially designed to characterize phase systems composed of polydisperse phase components. It gives information about the area representing the transition from homogeneous solution to a two-phase system. The two-phase systems studied were first a conventional Dextran T40-polyethylene glycol 20M (PEG 20M) system, then a less-well-defined phase system based on PEG 20M and partially hydrolyzed starch. The PEG 20M-starch system could be changed with respect to the volume ratio between the phases with time using hydrolytic enzymes, and the possibility of using the glucose released from the starch polymer is pointed out. Then the system is transformed to an extractive biconversion where the bottom phase polymer also served as the substrate.
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  • 7
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    Biotechnology and Bioengineering 31 (1988), S. 984-994 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: The use of microporous membranes has been examined for the recovery of precipitated protein suspensions and related soluble protein. Membrane flux rates and soluble protein transmissions are reported for unstirred batch-cell studies and cross-flow experiments. The unstirred batch-cell gave soluble protein transmissions in the range 80-100% for feeds containing either soluble protein or a mix of soluble and isoelectrically precipitated protein. In all cases a sharp decline in flux was observed which was, for example, considerably greater for soluble protein at its isoelectric point, pH 4.6, than at pH 8.8. The presence of precipitated protein led to a further decrease in flux rate. In cross-flow studies, flux decline was eventually accompanied by a significant decline in soluble protein transmission. The flux protein-transmission characteristics of microporous membranes are discussed in terms of the interaction of the soluble and precipitated protein with the membrane.
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  • 8
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    Biotechnology and Bioengineering 31 (1988), S. 995-1005 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: A membrane-covered oxygen electrode was used to measure oxygen diffusion coefficients and solubilities in aqueous glucose solutions and various fermentation media following a newly developed methodology. The fermentation media studied were tryptic soy broth and those for fermentations of Penicillium chrysogenum, Saccharomyces cerevisiae, and Micrococcus glutamicus. The experimental results of oxygen diffusion coefficients and solubilities in glucose solutions were in good accord with the literature data. As for the fermentation media, both oxygen diffusion coefficients and solubilities were found to decrease with an increased fractional composition of these media, and log-additive behaviors of the oxygen diffusion coefficients and solubilities in fermentation media were observed.
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  • 9
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    Biotechnology and Bioengineering 31 (1988), S. 1006-1009 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
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  • 10
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    Biotechnology and Bioengineering 31 (1988), S. 1010-1011 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
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  • 11
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    Biotechnology and Bioengineering 32 (1988) 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
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  • 12
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    Biotechnology and Bioengineering 32 (1988), S. iii 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
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  • 13
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    Biotechnology and Bioengineering 31 (1988), S. 495-501 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
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  • 14
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    Biotechnology and Bioengineering 31 (1988), S. 487-494 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Gas holdup and oxygen transfer studies in non-Newtonian suspensions of cellulose fibres conducted in two large (0.098 m3 each) reactors are described. Both reactors - a bubble column and a similar internal loop airlift - were unusual in that they had rectangular cross-sections. In all cases gas holdups and kLaL declined with increasing solid concentration and, under identical conditions, the bubble column performed better than the airlift. The fluid systems used were carefully selected to represent mould fermentation broths.The behavior of true mass transfer coeffcient kL with changes in bubble size is discussed for these systems.
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  • 15
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    Biotechnology and Bioengineering 31 (1988), S. 502-506 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
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  • 16
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    Biotechnology and Bioengineering 31 (1988), S. 287-294 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: A simple adaptive control algorithm, for which theoretical stability and convergence properties had been previously demonstrated, has been successfully implemented on a biomethanation pilot reactor. The methane digester, operated in the CSTR mode was submitted to a shock load, and successfully computer controlled during the subsequent transitory state.
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  • 17
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    Biotechnology and Bioengineering 31 (1988), S. 295-299 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: The influence of temperature, pH, and substrate and product concentrations on the oxidation rate of ferrous iron by biofilm of Thiobacillus ferrooxidans was determined. The experiments were performed in an inverse fluidized-bed biofilm reactor in which the biofilm thickness was kept constant at 80 μm. Oxygen concentration and diffusion through the biofilm did not limit the oxidation rate. The oxidation rate was almost unaffected by temperature between 13 and 38°C, pH between 1.3 and 2.2, ferric iron concentration up to 14 g/L, or ferrous iron concentration from 4 to 13 g/L. The kinetics of the process was described by the Monod equation with respect to the mass of the biofilm and with ferrous ions as the limiting substrate.
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  • 18
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    Biotechnology and Bioengineering 31 (1988), S. 304-310 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: A segregated model of multicopy plasmid propagation has been formulated which incorporates plasmid replication and partition functions, as well as the effect of plasmid presence on host growth rate. Growth of plasmid-free cells in selective medium is explicitly analyzed. The model parameters can be determined from experimentally measurable quantities. Propagation of a recombinant multicopy plasmid in the yeast Saccharomyces cerevisiae is analyzed using this model.
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  • 19
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    Biotechnology and Bioengineering 31 (1988), S. 300-303 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: A total of 58 yeast strains from 12 genera were assayed for their ability to grow and ferment carbohydrates in standard Durham tube test at 40, 43, and 46°C. Based on the kinetic parameters for glucose fermentation in shaken flask cultures, the strain Fabospora fragilis CCY51-1-1 was chosen for further studies. It reached about 56.0 and 35.0 g ethanol/L from ∼140 g glucose/L at 43 and 46°C in less than 48 h, respectively. Trichoderma reesei cellulase preparation (400 FPU/L) had not distinct effect on the ethanol yield and biomass production by the selected strain in the first 12 h fermentation at 46°C. Later a negligible decrease in both yields was observed. It was found that Fabospora fragilis did not grow or produce ethanol at 46°C as tho initial ethanol concentration overcame 40 g/L.
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  • 20
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    Biotechnology and Bioengineering 31 (1988), S. 311-320 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: The objective of this work was to relate macroscopically measurable on-line fermentation parameters such as dissolved oxygen, off-gas oxygen and carbon dioxide, and cell mass, to the controlled production of key intracellular enzymes under carbon limited conditions. Both batch and perturbed batch aerobic fermentations were performed using two different strains of Escherichia coli, with glucose and lactose as the sole carbon sources. The two strains differed from each other only in the lac operon region of their genome. The parent strain, E. coli 3000, was inducible for the enzyme β-galactosidase. The other strain, E. coli 3300, was a constitutive mutant in the production of β-galactosidase. In all experiments, off-line assays of sugars and β-galactosidase activity were performed. It was observed that there is a clear relationship between the macroscopic on-line measurements, dissolved oxygen tension, carbon dioxide evolution rate and oxygen uptake rate, and the microscopic control phenomena of catabolite repression, catabolite inhibition, and inducer repression.
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  • 21
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    Biotechnology and Bioengineering 32 (1988), S. 64-67 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Photoacoustic spectroscopy was used to monitor photo synthetic electron transfer in native and immobilized thylakoid membranes. The photoacoustic parameter φr′ (the percentage of absorbed energy that is stored in photo chemical intermediates) and i50 (the half-saturation modulated light intensity) were directly correlated to electron transfer rates. As previously shown, thylakoids immobilized in an albumin-glutaraldehyde matrix were more resistant to aging. The inhibitory effects of the immobilization procedure and of aging at 4°C were detected as a decrease in i50 values. In analogy with enzyme kinetic analysis, the effect could be characterized as a competitive type of inhibition. Photoacoustic measurements are performed in conditions similar to a working bioreactor cell with regards to the sample preparation.
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  • 22
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    Biotechnology and Bioengineering 31 (1988), S. 559-566 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Naphthalene-2-sulfonate was degraded by submerse growing Pseudomonads in a chemostat culture. The kinetic parameters for the Monod equation, including Pirts maintenance energy, were calculated from these experiments regarding naphthalene-2-sulfonate as substrate and oxygene as cosubstrate. By immobilizing the bacteria on sand particles, the degradation of naphthalene-2-sulfonate was carried out in a specialy designed three-phase airlift-loop reactor in a completely fluidized state. From these experiments, the influence of biofilm diffusion limitation on reaction kinetics and criteria for stable biofilm formation on sand particles were obtained.
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  • 23
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    Biotechnology and Bioengineering 31 (1988), S. 553-558 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: A reactor, using the enzymatic electrocatalysis scheme, was developed on a laboratory preparative scale for the catalytic oxidation of glucose into gluconic acid. Glucose oxidase was directly immobilized on the surface of a carbon felt electrode and a solution of glucose and benzo-quinone passed through the electrode in order to regenerate the electron acceptor. The reactor was able to produce continuously 1.5 g gluconate/h with a catalytic current of 0.4 A. This gave a high productivity ca. 100 g/h/L reactor. A one-dimensional model was developed which demonstrated the efficiency of coupling between enzymatic and electrochemical reactions due to the proximity of the two reaction sites. For example the catalytic current was practically independent of mass transfer parameters. The operational stability of immobilized glucose oxidase was increased 50 times at least when electroregenerated benzoquinone was used as oxidant instead of oxygen.
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  • 24
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    Biotechnology and Bioengineering 31 (1988), S. 567-578 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: In production-scale bioreactors microorganisms are exposed to a continually changing environment. This may cause loss of viability, reduction of the yield of biomass or desired metabolites, and an increase in the formation of by-products. In fed-batch production of baker's yeast, profiles may occur in substrate and oxygen concentrations and in pH. This article deals with the influence of a periodically changing oxygen concentration on the growth of baker's yeast in a continuous culture. Also, influences on the production of ethanol, glycerol, acetic acid, and on the composition of the cells were investigated. It was found that relatively fast fluctuations between oxygen-unlimited and oxygen-limited conditions with a frequency of 1 or 2 min had a distinct influence on the biomass and metabolite production. However, RNA, protein, and carbohydrate contents measured in cells exposed to fluctuations differed little from those in cells from an oxygen-unlimited or an oxygen-limited culture. The respiration and fermentation capacities of cells exposed to fluctuations can be larger than the capacities of cells grown under oxygen-unlimited conditions.
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  • 25
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    Biotechnology and Bioengineering 32 (1988), S. 348-355 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Development of a novel two-layer anaerobic biofilm model is based on substrate utilization kinetics and mass transport. The model is applied to steady-state conditions for a fixed-film anaerobic reactor. The microbial film is considered to consist of two distinct biofilm layers, one adjacent to the second, with an acidogenic bacteria biofilm forming the outer layer and a methanogenic film the inner one. The model assumes that sugars are only metabolized by the first layer and converted into volatile fatty acids (VFA), while fatty acids are taken up only by the inner layer. The model is able to predict both substrate flux net uptake and methane production for steady-state conditions. The results of modeling agree with methane production experimental data published elsewhere. Further, the model shows why layered fixed-film reactors can withstand high and inhibitory concentrations of volatile fatty acids as well as severe overloading without failure.
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    Biotechnology and Bioengineering 32 (1988), S. 356-362 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: The fermentation of Streptomyces griseus for the production of cycloheximide is similar to other antibiotic fermentations in that product synthesis is subject to feedback regulation and the desired product is degraded in the fermentation broth. The productivity of this fermentation can thus be dramatically increased by removing the antibiotic from the whole broth as it is produced. One means for effecting this on-line product removal is to contact the whole fermentation broth with neutral polymeric resin immobilized in hydrogel beads. The antibiotic adsorbs to the immobilized resin via hydrophobic interactions. In this work, the adsorption of the antibiotic onto the immobilized resin was characterized. A biochemical model of the fermentation was then used to describe the time profiles of biomass, substrate, and antibiotic in a fermentation system in which whole broth is circulated from the fermentor through a continuously stirred extractor containing the adsorbent beads. Various operating conditions were examined to optimize the productivity of the fermentation.
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  • 27
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    Biotechnology and Bioengineering 32 (1988), S. 669-676 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: A continuous production process of maltotetraose was investigated by using immobilized maltotetraose (G4)- forming amylase (1,4-α-D-glucan maltotetraohydrolase, EC3.2.1.60) from Pseudomonas stutzeri adsorbed on a macroporous hydrophobic resin. The maximum reaction rate was obtained at 55°C and the activation energy of hydrolysis by immobilized G4-forming amylase was calculated to be 8.45 kcal/mol. The maltotetraose yield was greatly influenced by the flow rate of substrate solution, its concentration, and the immobilized enzyme activity. The newly defined factor “specific space velocity” was successfully introduced to normalize the operating parameters. Using this factor, the immobilized enzyme reactor then can be simulated and the operating dynamics can be determined.
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    Biotechnology and Bioengineering 32 (1988), S. 664-668 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Invertase was ionically immobilized on the poly(ethylene-co-vinyl alcohol) hollow fiber inside surface, which was aminoacetalized with 2-dimethylaminoacetaldehyde dimethyl acetal. Immobilization and enzyme reaction were carried out by letting the respective solutions pass or circulate through the inside of the hollow fiber, and the activity of invertase was determined by the amount of glucose produced enzymatically from sucrose. Immobilization conditions were examined with respect to the enzyme concentration and to the time, and consequently the preferable conditions at room temperature were found to be 5 μg/mL of enzyme concentration and 4 h of immobilization time. Under those conditions the immobilization yield and the ratio of the activity of the immobilized invertase to that of the native one were 89 and 80%, respectively. For both repeating and continuous usages, the activity fell to ca. 60% of the initial activity in the early stage and after that almost kept that value. The apparent Michaelis constant Km′ for the immobilized invertase decreased with increasing the flow rate of the substrate solution, to be close to the value for the native one. Furthermore, the possibility of the separation of the enzymatically formed glucose from the reaction mixture through the hollow fiber membrane was preliminarily examined.
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  • 29
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    Biotechnology and Bioengineering 32 (1988), S. 396-399 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
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    Biotechnology and Bioengineering 32 (1988), S. 174-183 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Batch foam separation has been employed to separate Saccharomyces carlsbergensis cells from their broth without the use of any external surface-active agent. A model has been developed to predict the foamate cell concentration as well as the variation of cell concentration in the bulk liquid in the foam column as a function of time. The model assumes a linear equilibrium relation between the cell concentrations at the interface and the bulk. The foam has interface as well as interstitial liquid. The interface is assumed to be in equilibrium with the interstitial liquid, which in turn is assumed to have the same concentration as the bulk. The interfacial area is calculated by assuming the foam bubbles to be pentagonal dodecahedral in shape. The model has been found to explain the results of foam separation of cells quite well, particularly with respect to the effect of bubble size and aeration rate.
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    Biotechnology and Bioengineering 32 (1988), S. 159-173 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Anaerobic degradation performance of a laboratory-scale packed-bed reactor (PBR) was compared with two fluidized-bed biofilm reactors (FBRs) on molasses and whey feeds. The reactors were operated under constant pH (7) and temperature (35°C) conditions and were well mixed with high recirculation rates. The measured variables were chemical oxygen demand (COD), individual organic acids, gas composition, and gas rates. As carrier, sand of 0.3-0.5 mm diameter was used in the FBR, and porous clay spheres of 6 mm diameter were used in the PBR. Startup of the PBR was achieved with 1-5 day residence times. Start-up of the FBR was only successful if liquid residence times were held low at 2-3 h. COD degradations of 86% with molasses (90% was biodegradable) were reached in both the FBR and PBR at 6 h residence time and loadings of 10 g COD/L day. At higher loadings the FBR gave the best performance; even at 40-45 g COD/L day, with 6 h residence times, 70% COD was degraded. The PBR could not be operated above 20 g COD/L day without clogging. A comparison of the reaction rates show that the PBR and FBR per formed similarly at low concentrations in the reactors up to 1 g COD/L, while above 3 g COD/L the rates were 17.4 g COD/L day for the PBR and 38.4 g COD/L day for the FBR. This difference is probably due to diffusion limitations and a less active biomass content of the PBR compared with the fluidized bed.The results of dynamic step change experiments, in which residence times and feed concentrations were changed hanged at constant loading, demonstrated the rapid response of the reactors. Thus, the response times for an increase in gas rate or an increase in organic acids due to an increase in feed concentration were less than 1 day and could be explained by substrate limitation. Other slower responses were observed in which the reactor culture adapted over periods of 5-10 days; these were apparently growth related. An increase in loading of over 100% always resulted in large increases inorganic acids, especially acetic and propionic, as well as large increases in the CO2 gas content. In general, the CO2 content of the gas was very low, due to the large amount of dissolved CO2 that exited with the liquid phase at low residence times. The performance of the FBR with whey was comparable to its performance with molasses, and switching of molasses to whey feed resulted in immediate good performance without adaptation.
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  • 32
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    Biotechnology and Bioengineering 32 (1988), S. 184-191 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: A new combined bioreactor-separator system was designed and its operational feasibility demonstrated in order to develop a bioprocess that enables us to handle simultaneous biotransformation and recovery of product by crystallization. Enzymatic conversion of L-aspartate to L-alanine by L-aspartate β-decarboxylase from Pseudomonas dacunhae (ATCC 21192) was used as a model system for this study to demonstrate the principles involved in the bioprocess design. Immobilized cells of P. dacunhae containing the enzyme were fluidized in a tapered column type of bioreactor and a filter-crystallizer combination was used as a separator unit in our experimental system.It was found that almost a theoretical yield was achieved, and the process control for both the bioreactor operation and separation was relatively easy. The Production systems, namely, the recirculating bioreactor separator combination system and the conventional batch reactor system, were analyzed and compared based on the results obtained form this study, and it was found that a significant cost reduction, by about 20%, can be achieved when the recirculating bioreactor-separator combination system was employed. Based on these findings, it is anticipated that the conceptual design of the bioreactor-separator combination system evaluated in this study has some potential for industrial application.
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  • 33
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    Biotechnology and Bioengineering 32 (1988), S. 192-204 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: The performance of differential contactors for use in extractive fermentation is complicated by the effects of product formation in the contactor. When product formation is significant, approximate analytical solutions are presented for the performance of the contactor for two limiting cases: high and low substrate concentrations. When products are formed at a constant rate, there is a minimum raffinate solute concentration that can be obtained, in contrast to the behavior of a column in the absence of product formation. General equations describing the behavior of the system for product formation with backmixing in both phases are presented. The case of a stripping factor not equal to unity is considered.
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  • 34
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    Biotechnology and Bioengineering 32 (1988), S. 966-974 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Nuclear magnetic resonance (NMR) spectroscopic analysis of whole cells is an important emerging technique for noninvasive and nondestructive monitoring of cell physiology. However, this technique requires extremely high cell densities. Attempts to maintain densities above the carrying capacity of a maintenance system result in the demise of the entire culture. To define conditions for maintaining mammalian cells at high densities for NMR studies, we have designed a bioreactor to operate under defined, oxygen-limited conditions within an NMR spectrometer. The bioreactor utilizes hollow fibers to deliver nutrients and remove wastes from an agitated cell suspension. The mass transfer properties of the fibers with respect to oxygen were determined. Ehrlich Ascites Tumor (EAT) cells were supplied with glutamine as the respiratory carbon source. The maximum viable cell density supported by a given oxygen concentration in the fluid flowing through the fiber lumen was predicted and then confirmed experimentally on the bench and in the spectrometer.
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    Biotechnology and Bioengineering 32 (1988), S. 975-982 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Results are presented which show how the microcarrier concentration affects the hydrodynamic environment in animal cell bioreactors. At low levels of agitation, no physical effects of microcarrier concentration were found. However, cell growth was strongly influenced by cell concentration. At high levels of agitation, a strong detrimental effect of microcarrier concentration was found. A new mechanism of hydrodynamic damage was identified which is second order in microcarrier concentration. The identification of this mechanism adds to the fundamental understanding of hydrodynamic phenomena in microcarrier bioreactors.
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  • 36
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    Biotechnology and Bioengineering 31 (1988), S. 675-681 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: A novel human protein exhibiting erythroid differentiation activity was discovered in the culture fluids of phorbol ester-stimulated human cells. The differentiation assay system involving Friend virus-derived mouse leukemia cells was used. THP-1 cells of myelomonocytic origin were typical producers. 4β-Phorbol 12-myristate 13-acetate (PMA) was essential for inducible excretion of the erythroid differentiation factor (EDF). The factor was stable toward heat and pH (acidic or alkaline) but lost its activity on pronase treatment, which suggested its proteinous nature. After an optimization of the condition, production of EDF was performed on a 200-L scale for purification of the protein.
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  • 37
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    Biotechnology and Bioengineering 31 (1988), S. 682-695 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: A mathematical model was developed to simulate the operation of an activated sludge system with biomass support particles. Based on the results of this simulation study, it would appear that a system with biomass particle supports offers several advantages not offered by conventional systems. These include the ability to resist failure from large hydraulic surges; stable nitrification under transient inhibitory conditions resulting from temperature changes, hydraulic surges, and/or toxic chemicals; and the ability to establish stable operating conditions with respect to both carbon oxidation and nitrification at short hydraulic retention times and low sludge ages.
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  • 38
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    Biotechnology and Bioengineering 32 (1988), S. 983-992 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: A high surface area hollow fiber reactor was developed for mammalian cell culture. The reactor employs an interfiber gel matrix of agar or collagen for cell support. A model was developed to predict cell density as a function of fiber spacing. Optimum spacings are calculated for two sizes of Celgard hollow fibers. Ehrlich Ascites Tumor (EAT) cells were grown to an estimated density of 1.1 × 108 viable cells/mL in the extracapillary space - corresponding to an overall reactor density of 7 × 107 cells/mL. On the basis of available kinetic and diffusivity data, the model predicts that lactate accumulation may limit cell growth in the early stage of medium utilization, while oxygen delivery becomes limiting at later stages.
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  • 39
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    Biotechnology and Bioengineering 32 (1988), S. 993-1000 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Monoclonal antibodies are being manufactured for clinical trials in suspension culture at the 1300-L scale. Suspension culture offers some advantages relative to high-density mammalian cell culture methods; in particular, the ability to closely monitor the behavior of cells in a homogeneous environment. Computer control and on-line mass spectrography of exit gases provide instantaneous information about the culture metabolic activity. Air sparging and agitation by marine impeller provide aeration sufficient to maintain a constant dissolved oxygen tension at cell concentrations up to 5.0 × 106 cells/mL without causing apparent cell damage.
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  • 40
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    Biotechnology and Bioengineering 32 (1988), S. 803-812 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Substrate 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.
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    Biotechnology and Bioengineering 32 (1988), S. 826-830 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
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    Biotechnology and Bioengineering 32 (1988), S. 831-834 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
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    Biotechnology and Bioengineering 32 (1988), S. 835-840 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
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  • 44
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    Biotechnology and Bioengineering 32 (1988), S. 263-270 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: A laboratory model constant-depth film fermentor was developed. Film grew on the surface of polytetrafluoroethylene (PTFE) plugs and was limited to a predetermined depth by mechanically removing excess film. Six film-forming organisms were isolated from river water and used to assess the operating characteristics of the fermentor. Film accumulation was logarithmic, and a steady state was maintained. Electron micrographs show early film development. The fermentor enables film to be grown on any substratum and allows discrete, reproducible, and representative samples to be taken.
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  • 45
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    Biotechnology and Bioengineering 32 (1988), S. 277-288 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: In this study, a cephalosporin C producing strain, Cephalosporium acremonium (ATCC 36225), was chosen to determine the optimal conditions that maximize antibiotic production in a mixed substrate of glucose and sucrose. A model for cell growth and cephalosporin C production at different pH and temperature was developed and the associated parameters were evaluated experimentally. Pontryagin's maximum principle, in conjunction with the model, was used to predict the optimal temperature and pH control profiles to maximize the production of antibiotic.
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  • 46
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    Biotechnology and Bioengineering 32 (1988), S. 271-276 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: A kinetic model of ethanol fermentation conducted under a variety of conditions in a continuous four-stage reactor is proposed. The expressions for specific growth and product formation rates are: \documentclass{article}\pagestyle{empty}\begin{document}$$ \mu = \mu _0 {\rm exp( - }k_1 P){\rm (1 - }X/X_1) \\ \nu _P = \nu _0 {\rm exp( - }k_2 P){\rm (1 - }X/X_2) \\ $$\end{document} Parameters were identified by nonlinear programming and shown to fit data correctly for steady states of seven different experiments. The product inhibition constants were of 27 and 84 g/L, respectively. Secondary inhibitions were represented by the linear biomass term. The proposed model gave a better description of phenomena than one which only took ethanol inhibition into account. The same model also fitted batch fermentation data, with only some parameters altering significantly. The use of this model for on-line purposes is discussed.
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  • 47
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    Biotechnology and Bioengineering 32 (1988), S. 289-294 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Poly(styrene) microcapsules, prepared by depositing the polymer around emulsified aqueous droplets, were coated with a synthesized polyelectrolyte; i.e., copolymer of maleic acid (MA) with methyl vinyl ether (MVE), co-poly(MA, MVE), or with styrene (St), copoly(Ma, St). The permeability of the capsule membrane was investigated under various pHs of the outer medium using n-propyl alcohol as a permeant. It became apparent that either copoly(MA, St)- or copoly(MA, MVE)-coated microcapsules function as a pH-sensitive capsule. In particular, the former showed a dramatic change of the permeability in response to small differences in pH (5-6). By reference to the viscometric and electrophoretic studies of both copolymers, these were interpreted as being due to a pH-induced alteration of the configuration of the copolymer coating on the surface of the capsule membrane. When sucrose was hydrolyzed in an aqueous suspension of the copoly(MA, St)-coated capsules into which invertase was loaded, the hydrolytic reaction was initiated at pH 5. 5 and stopped at pH 4. 5. Such initiation-cessation control was repeated reversibly without damaging the capsules.
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  • 48
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    Biotechnology and Bioengineering 31 (1988), S. 770-782 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Batch fermentations of glucose to ethanol by Z. Mobilis.(ATCC 10988) were examined in several semidefined nutrient media. The measurement of acid produced by the microorganism was used to study its transient fermentation characteristics. Limitation of nitrogen source in the semidefined medium of Rogers and coworkers2 was found to limit the growth of this microorganism in the late stages of batch fermentations, when the initial glucose concentration was 75 g/L and higher. The microorganism exhibits a preference for inorganic nitrogen over preformed organic nitrogen provided by yeast extract. The microbial growth occurs exponentially in the presence of ammonium sulfate and yeast extract. However, in the absence of ammonium sulfate, the growth occurs in a linear fashion. The “linear” growth phase is characterized by poor cell-mass yields, and during this phase, growth and ethanol production are decoupled. An improved semi-defined growth medium is established which supports better growth rate and cellular yield, without affecting the ethanol yield.
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    Biotechnology and Bioengineering 32 (1988), S. 910-915 
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    Keywords: Chemistry ; Biochemistry and Biotechnology
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    Biotechnology and Bioengineering 32 (1988), S. 916-919 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
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    Biotechnology and Bioengineering 32 (1988), S. 920-922 
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    Keywords: Chemistry ; Biochemistry and Biotechnology
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    Biotechnology and Bioengineering 32 (1988), S. 1128-1137 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: The interrelationships between the three parts of the air lift reactor, the riser, the downcomer, and gas-liquid separator, were examined with relation to the overall mass transfer in the reactor. This involved studying the mass transfer of oxygen from the gas phase to the liquid phase for 20 different reactor geometries. Both one- and two-sparger systems were studied. It was demonstrated that the gas-liquid separator plays a major role in reactor behavior and must be considered in reactor design. It was found that the overall reactor mass transfer coefficient KLA could be correlated to the pneumatic power of gas input per total dispersion volume (P/VD) and to the true riser superficial gas velocity JGR for all experimental conditions examined. The KLA is directly related to the P/VD with an exponent of approximately 1. “Two-sparger” systems, where an auxiliary gas sparger is placed near the downcomer entrance, have higher ab solute values for KLA than single-sparger systems.
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  • 53
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    Keywords: Chemistry ; Biochemistry and Biotechnology
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    Notes: An activated sludge conditioned to a low concentration of phenol was used for the biodegradation of phenol in the presence of Chlorophenols, measuring the inhibiting action of all the latter compounds and expressing it as p/50. As a structural property to be correlated with the measured values, the lipophilicity (log P) of the Chlorophenols was reevaluated with a new apparatus. Values of p/50p are better correlated with the number of chloro substituents in the inhibitor, provided a specific effect of substituents ortho to the phenolic function is considered. New results are compared with previous measurements effected on a different activated sludge.
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    Biotechnology and Bioengineering 31 (1988), S. 829-833 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: We have recently prepared some crosslinked derivatives of Escherichia coli β-galactosidase by treating the enzyme with bisimidoesters. In this article, we report the results obtained when the native and these crosslinked derivatives are entrapped in polyacrylamide gel lattice. It was found that use of combination of three protective agents, viz., bovine serum albumin, cysteine, and lactose, during immobilization gave an increased yield of 190% in the case of DMA crosslinked preparation. In the case of native enzyme, the Km, pH optimum, and temperature optimum were found to remain unchanged on immobilization. The DMA crosslinked preparation entrapped in polyacrylamide in the presence of BSA, lactose, and cysteine was found to be a significantly better catalyst and hydrolyzed 47% milk lactose as compared to 31% hydrolysis by entrapped native enzyme in 6 h.
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  • 55
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    Keywords: Chemistry ; Biochemistry and Biotechnology
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    Notes: Among various microbial cells examined under screening conditions, Nocardia opaca showed the highest activity for production of phenylalanine from phenylpyruvate. Here NH4Cl as well as amino acids were used as an amino donor for phenylalanine production. The phenylalanine production rate increased with increasing hydrogen pressure. The specific activity of phenylalanine dehydrogenase was increased by culturing N. opaca cells in nutrient broth containing 0.3% phenylalanine. As a result, the phenylalanine production rate increased from 0.69 to 4.4 μmol/min g dry cells. Immobilized cells were activated in nutrient broth containing ZnCl2 before phenylalanine production. Phenylalanine dehydrogenase activity and cell number in the gel increased with increasing incubation time, and the maximum phenylalanine dehydrogenase activity was obtained at 36 h incubation. Then, phenylalanine was produced from phenylpyruvate, NH4Cl, and 100 atm H2 with the activated immobilized cells. The rate of phenylalanine production was 0.24 μmol/min cm3 gel. The conversion of phenylpyruvate to phenylalanine was 82%. Immobilized cells retained 76% of the initial phenylalanine production rate after 10 h reactions were repeated 11 times with two intervening reactivations.
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    Biotechnology and Bioengineering 32 (1988), S. 460-466 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Experimental data from the enzymatic hydrolysis of phenol-pretreated Swedish wheat straw have been used to evaluate the cost fractions of capital and utility, enzyme, and raw material. Two different raw material prices and varying enzyme prices have been used. The evaluation is based on an empirical model for the enzymatic hydrolysis and a computer program where utility and equipment, enzyme, and raw material prices can be varied. The optimal residence time for the enzymatic hydrolysis is in the range of 70-110 h. A fed-batch procedure with substrate concentrations higher than 10% oven-dried material (ODM) and enzyme concentrations in the range (6-10) · 106 FPU/ton ODM should be used.
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    Biotechnology and Bioengineering 32 (1988), S. 467-474 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Antisera against charged (p-azobenzoate and p-azoben zenesulfonate) and uncharged (dinitrophenyl) haptenic groups were produced in rabbits, and the equilibrium characteristics of hapten-antibody were measured by use of immunoadsorbents. The antibody to the uncharged hapten formed a stable binding with the hapten to the changes in ionic strength and pH. On the other hand, the antibodies to the charged haptens showed affinities sensitive to the changes in pH and ionic strength. Therefore, the effect of the pKa of ionizable haptens on the pH dependence of the hapten-antibody binding was studied by comparing the interactions between a series of para-substituted benzoic acids and the anti-p-azobenzoate antibody. The pH dependence of the interactions was strongly affected by the pKa of ionizable groups in haptens. Furthermore, the equilibrium characteristics of anti-p-aminobenzoyl dipeptides were compared. The characteristics of interactions were affected by the features of amino acid residues.
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    Biotechnology and Bioengineering 32 (1988), S. 475-481 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: A comparison of different immobilized enzyme reactors has been made for the limited modification of soya storage proteins and the products compared with those from action of the soluble enzyme. Clarified total water extracts of soya protein were subjected to the action of subtilisin in a soluble and immobilized form. The sodium dodecyl sulfate (SDS) electrophoresis patterns of soya proteins modified by enzyme in the two forms differed for unbuffered soya protein at the same pH of 8.0. However, identical patterns could be obtained by a downward adjustment of the pH of soya protein treated with immobilized enzyme. The same SDS electrophoresis pattern could be obtained for a packed column of immobilized enzyme and a well-mixed vessel by buffering. Operation of the column reactor at higher superficial linear velocities (above 1.47 cm/min), higher protein concentrations (8.8% w/v), and prolonged periods (24 h) led to a bed compression attributed to the protein coating of the support.
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  • 59
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    Biotechnology and Bioengineering 32 (1988), S. 482-490 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Empirical equations used so for for computing kla in mechanically agitated aerated vessels agree well with the measured dependence of Kla on the power input of the stirrer and the superficial velocity of the gas. However, they do not fit the dependence of Kla on the physical properties of the liquid batch. From the data published so far it can be deduced that the equations suitable for the description of the dependence of kmola on the liquid physical properties must include some independent variable, which characterizes the rate of coalescence of the bubbles. About 400 experimental values of Kla were compared with those calculated from different relations. This comparison shows that the rate of coalescence for the aqueous electrolyte solutions is well described on the basis of the Gibbs theory of adsorption of the solute in the surface layer of the solution.
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  • 60
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    Biotechnology and Bioengineering 32 (1988), S. 491-506 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: The lack of quantitative descriptions of mammalian culture kinetics limits the ability to optimally design and control cell culture bioreactors. This limitation is ad dressed by developing mathematical equations relating the initial growth rate and the antibody productivity of the hybridoma cell line, CRL-1606, to its environmental state. This initial rate approach, in contrast with steady state analysis of chemostat cultures, approximates steady state behavior, since the rates were measured over brief time intervals and at low cell concentrations (〈 50,000 cells/mL). The advantage of this approach is that it is much faster than the chemostat approach.An equation for the growth rate was developed that superimpoed Monod equations in serum and glutamine with “noncompetitive” -type inhibition constants were inversely proportional to the lactate and ammonium concentrations. The Monod constant is critical for relating initial, low cell level culture states to other states.Lactate was found to be the only environmental parameter that significantly inhibited antifibronectin monoclonal antibody production by the CRL-1606 hybridimas. Volumetric productivity was strictly related to culture viability, which was observed to decline at growth rates below 0.02h-1. Lactate was also found to significantly inhibit ammonium production.
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  • 61
    ISSN: 0886-1544
    Keywords: actinogelin ; α-actinin ; reconstituted actin-gel ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: We studied the properties of actinogelin, a Ca2+-regulated actin cross-linking protein isolated from Ehrlich tumor cells or rat liver. Chicken gizzard α-actinin was used as a Ca2+-insensitive control. Actinogelin, which has very high gelation activity under low Ca2+ conditions, was found using electron microscopic or fluorescence studies to induce formation of a characteristic structure in which actin filaments and bundles radiate to (or converge from) all directions from spot-like core structures. A similar structure was induced with actinogelin, even in the presence of 0 7 saturation of tropomyosin. No such structure was detected with actinogelin under high Ca2+ conditions, and only a few were found with gizzard α-actinin. Because reconstituted structures are similar to those observed intracellularly, actinogelin may be important in the formation of similar microfilament organization in the cells. It seems also important that these structures are reconstituted with only two purified protein components, i.e., actinogelin and actin.Immunocompetition studies showed that actinogelin and gizzard α-actinin partially shared antigenicity, and their molecular shape and peptide maps were similar. Their amino acid compositions [Kuo et al., 1982], subunit and domain structures, and binding sites on actin [Mimura and Asano, 1987] are also very similar. Therefore, it is concluded that actinogelin belongs to α-actinin superfamily proteins. Furthermore, the presence of functionally different subfamilies concerned with Ca2+ sensitivy, gelation-efficiency, and others is discussed. Actinogelin, which induces networks of actin filaments, may be classified as high gelation type.
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  • 62
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    Cell Motility and the Cytoskeleton 10 (1988), S. 471-481 
    ISSN: 0886-1544
    Keywords: Dictyostelium ; limited proteolysis ; thick filaments ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Dictyostelium myosin was associated into dimers and small oligomers at very low ionic strength, filamentous at intermediate ionic strength, and monomeric in solution conditions of high ionic strength. These different associations were probed by fragmenting myosin with chymotrypsin, trypsin, or V-8 protease. All three proteases digested monomeric myosin giving rise to multiple fragments with a wide range of molecular weights. Filamentous myosin was not digested by the V-8 protease, was preferentially cleaved at a single site in the middle of the heavy chain by chymotrypsin, and was cleaved at several sites by trypsin. If the reaction was carried out in very low ionic strength, however, two of these proteases generated stable fragments of high molecular weight. Electron microscopic analysis of these stable fragments showed that tails were shorter than in intact myosin, indicating that the cleavage sites were in the rod portion of the molecule. Under the same conditions of enzymatic digestion, myosin that had been radio labeled in vivo with 32P was analyzed by SDS-PAGE and autoradiography. By comparing the state of phosphorylation and the size of the stable fragments, it was determined that the heavy chain phosphorylation site was located between 55 and 70 kD from the tip of the myosin tail, near a region where the tail displayed sharp bends.
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  • 63
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    Cell Motility and the Cytoskeleton 10 (1988), S. 482-495 
    ISSN: 0886-1544
    Keywords: organelle motility ; kinesin ; cytoplasmic dynein ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Directed movements of organelles have been observed in a variety of cultured cells. To study the regulation and molecular basis of intracellular organelle motility, we have prepared extracts from cultured chick embryo fibroblasts (CEF cells) which support the movement of membranous organelles along microtubules. The velocity, frequency and characteristics of organelle movements in vitro were similar to those within intact cells. Organelles and extract-coated anionic beads moved predominantly (80%) toward the minus ends of microtubules that had been regrown from centrosomes, corresponding to retrograde translocation. Similar microtubule-dependent organelle movements were observed in extracts prepared from other cultured cells (African green monkey kidney and 3T3 cells).Organelle motility was ATP and microtubule dependent. The frequency of organelle movement was inhibited by acidic (pH〈7) or alkaline (pH〉8) solutions, high ionic strength ([KCl] = 0.1 M), and the chelation of free magnesium ions. Treatment of the extracts with adenylyl imidodiphosphate (AMP-PNP, 7 mM), sodium orthovanadate (vanadate; Na3VO4, 20 μM), or N-ethylmaleimide (NEM, 2 mM) blocked all organelle motility. The decoration of microtubules with organelles was observed in the presence of AMP-PNP or vanadate. Motility was not affected by cytochalasin D (2 μM) or cAMP (1 mM). Kinesin (Mr= 116,000), an anterograde microtubule-based motor, was partially purified from the CEF extract by microtubule affinity purification in the presence of AMP-PNP, and was able to drive the movement of microtubule on glass coverslips. A similar preparation made in the presence of vanadate contained a different subset of proteins and did not support motility. These results demonstrate that intracellular organelle motility can be reproduced in vitro and provide the basis for investigating the roles of individual molecular components involved in the organelle motor complex.
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  • 64
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    Cell Motility and the Cytoskeleton 10 (1988), S. 496-505 
    ISSN: 0886-1544
    Keywords: centrosome ; spindle matrix ; postembedding immunofluorescent labeling ; mitotic apparatus ; sea urchin eggs ; 51-kD protein ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The 51-kD protein, a protein component of the mitotic apparatus in sea urchin eggs, is involved in the aster-forming activity previously shown in vitro [Toriyama et al., 1988]. Postembedding immunofluorescent labelings of eggs from fertilization through first cleavage showed that the 51-kD protein is localized in sperm asters, centrosomal regions, spindles, basal regions of astral microtubules, and regions surrounding daughter nuclei at telophase in situ. Immunofluorescence and immunoblot analyses detected the 51-kD protein uniformly in unfertilized eggs, but not in spermatozoa. When unfertilized eggs were treated with taxol, the 51-kD protein was shown to be associated with taxol-induced cytasters. Immunoblot analysis revealed that similar protein species are present in the mitotic apparatus of other species of sea urchin. It was suggested that the 51-kD protein may be involved in microtubule nucleation and microtubule matrix in sea urchin eggs in vivo.
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  • 65
    ISSN: 0886-1544
    Keywords: axoneme ; flagellar movement ; helical wave ; planar wave ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: American horseshoe crab sperm flagella have the typical 9+2 structure whereas Asian horseshoe crab sperm flagella have a 9+0 axoneme lacking central pair and central sheaths. Beat patterns of the American and the Asian horseshoe crab sperm were recorded by means of a high-speed video system (200 fields/second) and were compared in order to study the role of the central pair of the axoneme in ciliary and flagellar movement.The American horseshoe crab sperm beat with relatively planar waves, whereas the Asian horseshoe crab sperm beat with right-handed helical waves. These results suggest that the central complex plays an important role in forming planar waves, whereas it is not essential for the conversion of microtubule sliding into axonemal bends.
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  • 66
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    Cell Motility and the Cytoskeleton 9 (1988), S. 73-84 
    ISSN: 0886-1544
    Keywords: cilia ; metachronal waves ; electron microscopy ; calcium ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Structural and behavioral features of intact and permeabilized Paramecium tetraurelia have been defined as a basis for study of Ca2+ control of ciliary reversal. Motion analysis of living paramecia shows that all the cells in a population swim forward with gently curving spirals at speeds averaging 369 ± 19 μm/second. Ciliary reversal occurs in 10% of the cell population per second. Living paramecia, quick-fixed for scanning electron microscopy (SEM), show metachronal waves and an effective stroke obliquely toward the posterior end of the cell. Upon treatment with Triton X-100, swimming ceases and both scanning and transmission electron microscopy reveal cilia that uniformly project perpendicularly from the cell surface. Thin sections of these cells indicate that the ciliary, cell, and outer alveolar membranes are greatly disrupted or entirely missing and that the cytoplasm is also disrupted. These permeabilized paramecia can be reactivated and are capable of motility and regulation of motility. Motion analysis of cells reactivated with Mg2+ and ATP in low Ca2+ buffer (pCa7) shows that 71% swim forward in straight or curved paths at speeds averaging 221 ± 20 μm/second. When these cells are quick-fixed for SEM the metachronal wave patterns of living, forward swimming cells reappear. Motion analysis of permeabilized cells reactivated in high Ca2+ buffers (pCa 5.5) shows that 94% swim backward in tight spirals at a velocity averaging 156 ± 7 μm/second. SEM reveals a metachronal wave pattern with an effective stroke toward the anterior region. Although the permeabilized cells do not reverse spontaneously, the pCa response is preserved and the Ca2+ switch remains intact. The ciliary axonemes are largely exposed to the external environment. Therefore, the behavioral responses of these permeabilized cells depend on interaction of Ca2+ with molecules that remain bound to the axonemes throughout the extraction and reactivation procedures.
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    Cell Motility and the Cytoskeleton 9 (1988), S. 85-96 
    ISSN: 0886-1544
    Keywords: fertilization ; echinoderm eggs ; egg cortex ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Changes in the distribution and organizational state of actin in the cortex of echinoderm eggs are believed to be important events following fertilization. To examine the initial distribution and form of actin in unfertilized eggs, we have adapted immunogold-labeling procedures for use with eggs of Strongylocentrotus purpuratus. Using these procedures, as well as fluorescence microscopy, we have revealed a discrete 1-μm-thick concentrated shell of actin in the unfertilized egg cortex. This actin is located in the short surface projections of unfertilized eggs and around the cortical granules in a manner that suggests it is associated with the cortical granule surface. The actin in the short surface projections appears to be organized into filaments. However, most if not all of the actin surrounding the cortical granules is organized in a form that does not bind phalloidin, even though it is accessible to actin antibody. The lack of phalloidin binding is consistent with either the presence of nonfilamentous actin associated with the cortical granules or the masking of actin-filament phalloidin-binding sites by some cellular actin-binding component. In addition to the concentrated shell of actin found in the cortex, actin was also found to be concentrated in the nuclei of unfertilized eggs.
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    Cell Motility and the Cytoskeleton 9 (1988) 
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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  • 69
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    Cell Motility and the Cytoskeleton 9 (1988), S. 283-298 
    ISSN: 0886-1544
    Keywords: colchicine ; microtubule ; mitosis ; rhodamine-phalloidin ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The effect of cytochalasins on F-actin was investigated in Allium root cells stained with rhodamine-phalloidin. With cytochalasin D (CD), the normal interphase network of actin fibers is replaced by dispersed rods and specks similar to those seen in animal cells. However, during division, the specks accumulate at the poles in the form of one to a few large aggregates. The effects intensify with increasing concentration (0.5-5 m̈g/ml) and exposure time (0.5-3 hr). Further, similar behavior is observed with cytochalasin B, but dihydroCB has little effect. Double localizations show that during preprophase, aggregates cluster in association with microtubule foci at the new poles located near the nuclear envelope. From metaphase through anaphase, the aggregates are often found near the tips of kinetochore fibers, while in telophase they are often appressed to the pole side of the daughter nuclei. No association is seen between actin and the other microtubule arrays. The reorganization of F-actin into small specks is unaffected by sodium azide, but aggregation at the poles is very sensitive to this agent. Polar clustering is also blocked by oryzalin, colchicine, and isopropyl n-(3-chlorophenyl) carbamate, but taxol has no effect. Experiments with scleroderma serum 5051 show that CD-induced aggregates are embedded in centrosomal material at the poles. The results reveal that the reorganization of actin in response to cytochalasins differs during the cell cycle. Furthermore, the aggregation of actin during division is probably governed by an energy-dependent interaction with microtubules.
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    Cell Motility and the Cytoskeleton 10 (1988) 
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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  • 71
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    Cell Motility and the Cytoskeleton 9 (1988), S. 361-374 
    ISSN: 0886-1544
    Keywords: microtubular cytoskeleton ; Dinoflagellates ; immunofluorescence ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The cytoskeletal microtubule system has been studied in six species of unarmoured Dinoflagellates using immunofluorescence and electron microscopy. Several structures have been detected and described: (1) a subpellicular layer of microtubules, constituting the microtubular cytoskeleton, running singly or in bundles from the anterior part of the cell to the posterior; (2) a feeding apparatus, containing a ribbon of microtubules, which corresponds to a small peduncle in some species and is simply represented by a cytostome in some other species; and (3) the longitudinal flagellum that runs in a long intracytoplasmic pocket before becoming free at the extremity of the sulcus. A thorough study of the organization of the microtubular structures in a wide spectrum of Dinoflagellates is a prerequisite for understanding the evolutionary history of the group.
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    Cell Motility and the Cytoskeleton 9 (1988), S. 117-128 
    ISSN: 0886-1544
    Keywords: centrosome ; aster-forming activity ; tubulin polymerization ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Mitotic apparatuses (MAs) isolated from sea urchin metaphase eggs were chilled on ice to depolymerize microtubules, homogenized, and incubated with tubulin. This caused formation of many small asters with microtubules focusing on granules which were probably fragments of the centrosome. The aster-forming protein components of the granules in the homogenized MAs were solubilized in 0.5 M KCl containing 50% glycerol. After dialysis against low-ionic-strength buffer solution, proteins congregated to form granular assembly capable of initiating aster formation. Phosphocellulose column chromatography enabled the separation of the aster-forming protein fraction which contained a 51,000 molecular weight protein (51-kd protein) as a major component. The protein fraction possessing the aster-forming activity was also prepared from methaphase whole egg homogenate, and the elution profile of the 51-kd protein on phosphocellulose column also coincided with that of the aster-forming activity. The granular assembly reconstituted from the phosphocellulose fraction formed asters whose microtubules show the same growth rate and length distribution as those of asters reconstructed from the granules in the homogenized MAs. Anti-51-kd protein antibody that was raised in rabbit and affinity-purified stained the center of asters which were reconstructed either from the granules in the homogenized MAs or from the granular assembly reconstituted from the phosphocellulose fraction. These results suggest that the 51-kd protein is a component in the aster-forming activity of the centrosomal component in vitro.
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    Cell Motility and the Cytoskeleton 9 (1988), S. 129-139 
    ISSN: 0886-1544
    Keywords: microtubules ; motility ; cilia ; surface lattice ; biotin ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Studies were conducted to determine if dynein could bind to unpolymerized tubulin. Tubulin alone normally fractionated in the included volume of a molecular sieve Bio-Gel A-1.5m column. Incubated together, tubulin and dynein coeluted in the void volumn, suggesting that a complex had formed between the two. In addition, immunoelectron microscopy revealed preassembled microtubules were labeled with biotin antibody only when incubated in both dynein and biotinylated tubulin, evidence that dynein with bound biotinylated tubulin had decorated the microtubules. A fraction of the tubulin could be dissociated from dynein by addition of ATP and vanadate, as assayed by molecular sieve chromatography followed by densitometry of gels, suggesting that some tubulin bound to the B end of the dynein arm. Additional tubulin dissociated from the dynein under conditions of high salt. These studies, together with those indicating that tubulin blocked the A end of the dynein arm from binding to microtubules and promoted the interaction of two arms at their A ends, provide evidence that the A end of the arm also can bind tubulin. Thus, the tubulin subunits, themselves, on a microtubule rather than a particular surface lattice structure formed by adjacent protofilaments may provide the binding sites for both ends of the dynein arm.
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  • 74
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    Cell Motility and the Cytoskeleton 9 (1988), S. 140-152 
    ISSN: 0886-1544
    Keywords: cell surface ; cytoskeleton ; receptor-mediated endocytosis ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Cell-surface IgM (antigen receptor) sediments with the membrane fraction following osmotic lysis and homogenization of cells of the human lymphoblastoid cell line WiL2. In nonreducing buffers, SDS PAGE analysis of membrane pellets demonstrates that “native” membrane IgM exists as a dimer. In contrast to osmotic lysis, lysis of cells with the nonionic detergent Triton X-100 releases approximately 90% of the membrane-bound IgM into the supernatant; approximately 10% of the IgM pellets with the cytoskeletal fraction on centrifugation. Ligand challenge with either m̈-chain-specific antibodies or concanavalin A induces a change in the state of membrane IgM making it refractory to detergent extraction, such that 43% of the IgM pellets during centrifugation. This ligand-induced retention of IgM is significantly diminished by the microfilament-disrupting agent cytochalasin D, whereas pretreatment of cells with sodium azide or colchicine results in no significant change in the percentage of membrane IgM retained by Triton X-100 residues. These results indicate that retention of IgM involves an association with the cortical actin-based cytoskeleton. Investigation of the structural basis for ligand-induced Triton X-100 retention of membrane IgM by using ferritin-conjugated antibodies, myosin subfragment S1, and stereo-imaging electron microscopy has revealed linkages between ligand-receptor (antigen-IgM) complexes and elements of the cortical actin-based cytoskeleton.
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  • 75
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    Cell Motility and the Cytoskeleton 9 (1988), S. 153-163 
    ISSN: 0886-1544
    Keywords: fertilization cone ; fluorescence redistribution after photobleaching ; fluorescent analog cytochemistry ; microinjection of actin ; microvilli ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Actin from sea urchin eggs was fluorescently labeled with fluorescein isothiocyanate (FITC), N-(7-dimethylamino-4-methylcoumarinyl)-maleimide (DACM), or 5-iodoacetamidofluorescein (IAF) and microinjected into sea urchin eggs and oocytes. It distributed evenly in the cytoplasm of unfertilized eggs. Upon fertilization, actin accumulated first around the sperm binding site and, soon afterwards, in the fertilization cone. The accumulation propagated all over the cortex after a latent period of 10-20 sec. In the case of Clypeaster japonicus eggs, propagation of the accumulation coincided with a shape change in the egg, suggesting that the accumulated actin in the cortex generates forces. FITC-actin was incorporated into microvilli and retained in the cortex after cleavage. On the other hand, DACM- or IAF-actin was not incorporated into microvilli and was dispersed from the cortex by cleavage. These differences may be attributable to differences in the properties of the actins labeled at different sites. After photobleaching by laser light irradiation, FITC- or IAF-actin redistributed in the cortex of fertilized egg as quickly as it did before fertilization. When an unfertilized egg was injected with both actin and a calcium buffer (intracellular free Ca2+ concentration 9 μM), the actin accumulation was similar to that during fertilization but without the latent period. This suggests that the accumulation depended on the increase in the intracellular free Ca2+ concentration. When the unfertilized egg was injected with 0.2 M EGTA after injection of labeled actin and then inseminated, it accumulated only in the protrusion of cytoplasm where the sperm had entered, and fertilization was not completed. In immature oocytes, the accumulation was observed in the cortical region, including the huge protrusion of the cytoplasm where the sperm had entered. These results suggest that actin accumulation in the sperm binding site plays an important role in the sperm reception mechanism of the egg.
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  • 76
    ISSN: 0886-1544
    Keywords: Ca2+-ATPase of sarcoplasmic reticulum ; immunofluorescence ; myofibers types I (slow) and II (fast) ; II D8 monoclonal antibody ; II H11 monoclonal antibody ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Ca2+ -ATPase of the sarcoplasmic reticulum was localized in cryostat sections from three different adult canine skeletal muscles (gracilis, extensor carpi radialis, and superficial digitalis flexor) by immunofluorescence labeling with monoclonal antibodies to the Ca2+ -ATPase Type I (slow) myofibers were strongly labeled for the Ca2+ -ATPase with a monoclonal antibody (II D8) to the CA2+ -ATPase of canine cardiac sarcoplasmic reticulum; the type II (fast) myofibers were labeled at the level of the background with monoclonal antibody II D8. By contrast, type II (fast) myofibers were strongly labeled for Ca2+ -ATPase of rabbit skeletal sarcoplasmic reticulum. The subcellular distribution of the immunolabeling in type I (slow) myofibers with monoclonal antibody II D8 corresponded to that of the sarcoplasmic reticulum as previously determined by electron microscopy. The structural similarity between the canine cardiac Ca2+ -ATPase present in the sarcoplasmic reticulum of the canine slow skeletal muscle fibers was demonstrated by immunoblotting. Monoclonal antibody (II D8) to the cardiac Ca2+ -ATPase binds to only one protein band present in the extract from either cardiac or type I (slow) skeletal muscle tissue. By contrast, monoclonal antibody (II H11) to the skeletal type II (fast) Ca2+ -ATPase binds only one protein band in the extract from type II (fast) skeletal muscle tissue. These immunopositive proteins coelectrophoresed with the Ca2+ -ATPase of the canine cardiac sarcoplasmic reticulum and showed an apparent Mr of 115,000. It is concluded that the Ca2+ -ATPase of cardiac and type I (slow) skeletal sarcoplasmic reticulum have at least one epitope in common, which is not present on the Ca2+ -ATPase of sarcoplasmic reticulum in type II (fast) skeletal myofibers. It is possible that this site is related to the assumed necessity of the Ca2+ -ATPase of the sarcoplasmic reticulum in cardiac and type I (slow) skeletal myofibers to interact with phosphorylated phospholamban and thereby enhance the accumulation of Ca2+ in the lumen of the sarcoplasmic reticulum following β-adrenergic stimulation.
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    Cell Motility and the Cytoskeleton 9 (1988), S. 175-183 
    ISSN: 0886-1544
    Keywords: cytoskeleton ; microtubules ; monoclonal antibodies ; cell morphogenesis ; tubulin ; Trypanosoma brucei ; subflagellar microtubule quartet ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Tubulin from Trypanosoma brucei was characterized by Western blotting using well defined monoclonal antibodies reacting with α- or β-tubulin and a new monoclonal antibody, 1B41, raised against a microtubule-enriched fraction of T. brucei, which specifically reacts with the β-subunit of tubulin from either T. brucei or rat brain. This antibody has been used to examine the subcellular distribution of the corresponding antigen in T. brucei by indirect immunofluorescence. The epitope recognized by 1B41 is restricted to a thin line extending from the basal body region to the anterior end of the cell body. To determine the relationship between the immunoreactive zone and the flagellum, double-label immunofluorescence was performed in both interphase and mitotic cells with 1B41 and a flagellar marker, the monoclonal antibody 5E9, specific for the paraflagellar rod polypeptides of trypanosomes. These experiments revealed that the immunoreactive tubulin was contained in a part of the subpellicular cytoskeleton that remained in a constant spatial correspondence with the flagellum throughout the cell division cycle. The β-tubulin recognized by 1B41 may be segregated into the microtubular structures associated with a cisterna of the endoplasmic reticulum forming the subflagellar microtubule quartet (SFMQ). These results suggest that the presence of an antigenically unique β-tubulin defines a subpopulation of microtubules possessing specfic dynamic properties that may be involved in the morphogenesis of daughter cells during the division of T. brucei.
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  • 78
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    Cell Motility and the Cytoskeleton 9 (1988) 
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
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  • 79
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    Cell Motility and the Cytoskeleton 9 (1988), S. 184-189 
    ISSN: 0886-1544
    Keywords: cell locomotion ; cell motility ; calcium ; polymorphonuclear leukocyte ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Chemotactic factors stimulate the rate of locomotion of polymorphonuclear leukocytes (PMNs). To investigate the importance of cytoplasmic calcium we have examined the ability of the chemotactic peptide N-formylnorleucyl eucylphenalanine (FNLLP) to stimulate the locomotion of PMNs whose cytoplasmic calcium levels were reduced by incubation in EGTA or in EGTA plus the calcium ionophores, ionomycin or A23187. Locomotion was assayed by migration through micropore filters and by time-lapse videomicroscopy. Cells in EGTA exhibited similar or slightly reduced rates of locomotion compared to cells in Hanks' balanced salt solution (HBSS). The peptide dose dependence for the stimulation of locomotion was similar in medium containing calcium or EGTA. The presence of 1 μM ionophore plus EGTA had no effect on the stimulation of locomotion by peptide. The presence of ionophores (1 μM) plus external calcium inhibited locomotion.
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  • 80
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    Cell Motility and the Cytoskeleton 10 (1988), S. 54-61 
    ISSN: 0886-1544
    Keywords: immunofluorescence ; optical sectioning ; cytoskeleton ; microscopy ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: A wide variety of specimens has been examined with our apparatus, a commercial version of which is being manufactured by Bio-Rad/Lasersharp. The advantages expected of a confocal system have been realised in practice, the most striking advantage being the exclusion of glare from out-of-focus structures. This has made it possible to image cytological details in unflattened cells and intact tissues that were previously inaccessible to the light microscope.
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  • 81
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    Cell Motility and the Cytoskeleton 10 (1988), S. 62-70 
    ISSN: 0886-1544
    Keywords: interference filters ; fluorescence spectroscopy ; fluorescence microscopy ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Recent advances in thin film optical coating technology significantly improve the filters available for fluorescence spectroscopy. Bandpass and long- and shortpass filters with very sharply defined edges can provide from 10-5 to 10-6 blocking within 10-15 nm of the transmission region and are ideal for use as excitation and emission filters. A variety of nonpolarizing dichroic beamsplitters for use in epi-illumination configurations or in multiple emission configurations provides optimum longpass, shortpass, band reflection, or bandpass spectral control. These dichroics, used with high-performance bandpass, longpass, or shortpass filters, form matched sets that optimize the signal-to-noise ratio and system efficiency for fluorescence spectroscopic systems in single or multiple dye applications. Specially designed dichroic beamsplitters are used to reduce excitation filter overheating. Other dichroic beamsplitters efficiently separate two planes of polarization in a narrow wavelength band. Rejection band filters can be used to measure the fluorescent dye Indo 1 with very low emission signals.
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  • 82
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    Cell Motility and the Cytoskeleton 10 (1988), S. 71-76 
    ISSN: 0886-1544
    Keywords: myosin ; actin ; filament structure ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: We used video-fluorescence microscopy to directly observe the sliding movement of single fluorescently labeled actin filaments along myosin fixed on a glass surface. Single actin filaments labeled with phalloidin-tetramethyl-rhodamine, which stabilizes the filament structure of actin, could be seen very clearly and continuously for at least 60 min in O2-free solution, and the sensitivity was high enough to see very short actin filaments less than 40 nm long that contained less than eight dye molecules. The actin filaments were observed to move along double-headed and, similarly, single-headed myosin filaments on which the density of the heads varied widely in the presence of ATP, showing that the cooperative interaction between the two heads of the myosin molecule is not essential to produce the sliding movement. The velocity of actin filament independent of filament length (〉1 μm) was almost unchanged until the density of myosin heads along the thick filament was decreased from six heads/14.3 nm to 1 head/34 nm. This result suggests that five to ten heads are sufficient to support the maximum sliding velocity of actin filaments (5 μm/s) under unloaded conditions. In order for five to ten myosin heads to achieve the observed maximum velocity, the sliding distance of actin filaments during one ATP cycle must be more than 60 nm.
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  • 83
    ISSN: 0886-1544
    Keywords: birefringence ; Physarum ; acellular slime mold ; cytoplasmic streaming ; contractility ; rhythm ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Birefringent fibrils (BRFs) with a positive sign composed of bundles of F-actin were found throughout the Physarum plasmodium with the mode of existence differing regionally. In the zone behind the leading edge of an advancing plasmodium, where cytoplasmic sol and gel were still not well differentiated, more BRFs came to the foreground when the endoplasm flowed backward (emptying phase), and a substantial portion disappeared when the endoplasm flowed forward (filling phase), except for nodes, from which BRFs were reorganized in the early emptying phase of each cycle. BRFs found in the wall of the streaming channel in the posterior network and the branched vein section ran in parallel to or helically around the channel. They were much more stable and maintained strong birefringence irrespective of the direction of the cytoplasmic flow. When the fan-like expanse ceased moving forward, the BRFs no longer appeared and disappeared cyclically but persisted in the area which had previously been the front. We concluded that the site of the active contraction-relaxation rhythm in an advancing plasmodium with antero-posterior polarity is restricted to its frontal zone and that the rest of the plasmodium is in a state of “tonus” which continuously imparts a certain level of hydrostatic pressure to the interior. The meaning of the tonus and the mechanics of tensile force production in the plasmodium are discussed in terms of a working hypothesis arrived at from the phase relationship between isometric and isotonic contraction waves.
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  • 84
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    Cell Motility and the Cytoskeleton 10 (1988), S. 126-136 
    ISSN: 0886-1544
    Keywords: microtubules ; Allogromia ; intracellular transport ; surface motility ; actin ; morphogenesis ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Microtubules are the major cytoskeletal component of foraminiferan reticulopodia. Video-enhanced differential interference contrast light microscopy has demonstrated that the microtubules serve as the intracellular tracks along which rapid bidirectional organelle transport and cell surface motility occurs. Microtubules appear to move, both axially and laterally within the pseudopodial cytoplasm, and these microtubule translocations appear to drive the various reticulopodial movements. F-actin is localized to discrete filament plaques form at sites of pseudopod-substrate adhesion. Correlative immunofluorescence and electron microscopy reveals a structural interaction between microtubules and the actin-containing filament plaques. Our recent data on reticulopodial motility are discussed in an historical context, and a model for foram motility, based on motile microtubules, is presented.
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  • 85
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    Cell Motility and the Cytoskeleton 10 (1988), S. 137-142 
    ISSN: 0886-1544
    Keywords: substrate-adhesion ; focal contact ; actin filaments ; talin ; vinculin ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Evidence for a structural precursor of the focal contact in cultured fibroblasts and continuing studies on the development of the precursor and contact are discussed. The structural precursor consists of an F-actin-rich, rib-like fiber within the motile lamellipodium. The focal contact forms beneath the fiber, part of which is retained at the contact as the initial adhesion plaque. Therefore, F-actin is present at the contact from the beginning. Vinculin accumulates at the plaque during a 90-second period after the contact forms. A novel feature of the distribution of talin has been found. The protein is present along the distal margin of the lamellipodium, where it is further concentrated as a series of nodes at the tips of each precursor and between precursors. This distribution of talin is independent of that which develops at the plaque after the contact forms. The structural development of the precursor has been followed with AVEC-DIC optics. The process begins with the development of fine oblique fibers from small structural nodes at the margin of the lamellipodium, and continues with the fusion of the nodes at the margin and inward coalescence of the fibers. It is suggested that talin may function as a cross-linking protein in the convergence of actin filaments at the membrane, while other actin-bundling proteins participate in the inward coalescence of the filaments to form fibers. The F-actin core of the precursor could provide a structural framework against which differences at the external surface of the membrane develop prior to contact formation.
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  • 86
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    Keywords: pigment organelle ; xanthophore ; microtubule ; F-actin ; intermediate filament ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: In goldfish xanthophores, the formation of pigment aggregate requires: (1) that a pigment organelle (carotenoid droplet) protein p57 be in the unphosphorylated state; (2) that self-association of pigment organelles occur in a microtubule-independent manner; and (3) that pigment organelles via p57 associate with microtubules. In the fully aggregated state, the pigment organelles are completely stationary. Pigment dispersion is initiated by activation of a cAMP-dependent protein kinase, which phosphorylates p57 and allows pigment dispersion via an active process dependent on F-actin and a cytosolic factor. This factor is not an ATPase, and its function is unknown. However, its abundance in different tissues parallels secretory activity of the tissues, suggesting a similarity between secretion and pigment dispersion in xanthophores. The identity of the motor for pigment dispersion is unclear. Experimental results show that pigment organelles isolated from cells with dispersed pigment have associated actin and ATPase activity comparable to myosin ATPase. This ATPase is probably an organelle protein of relative molecular mass ∼72,000, and unlikely to be an ion pump. Isolated pigment organelles without associated actin have 5× lower ATPase activity. Whether this organelle ATPase is the motor for pigment dispersion is under investigation. The process of pigment aggregation is poorly understood, with conflicting results for and against the involvement of intermediate filaments.
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  • 87
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    Cell Motility and the Cytoskeleton 10 (1988), S. 164-171 
    ISSN: 0886-1544
    Keywords: motility ; lamella ; cytoskeleton ; membrane ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
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  • 88
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    Cell Motility and the Cytoskeleton 10 (1988), S. 153-163 
    ISSN: 0886-1544
    Keywords: intracellular particle motions ; cytoplasmic streaming ; onion (Allium) epidermal cells ; video microscopy ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The endoplasmic reticulum (ER) and associated organelle and particle movements in onion (Allium cepa) bulb scale epidermal cells were observed, recorded, and analyzed using computer-assisted video (AVEC-DIC, AVEC-POL and fluorescence) microscopy. The ER is composed of two interconnected sets of filamentous membrane tubules with diameters ranging from 0.1 to 0.5 μm. The first form a more stable, stationary network of intersecting polygonal membrane tubules lying closely appressed to the plasma membrane and continuous with a second very dynamic set of longer membrane tubules that often are located parallel to each other, shifting rapidly around the cytoplasm and forming dynamic knots or organization centers. The ER, mitochondria, and spherosomes fluoresced upon chlortetracycline treatment and are therefore presumed to sequester calcium. ER and mitochrondria also stain with the fluorescent dye, rhodamine 123. Mitochrondria and spherosomes are seen to move in the cytoplasm only along paths parallel to the axis of the ER tubules. Smaller particles (0.5 μm) tend to follow these same paths but may occasionally move independently. Particles and organelles move in close, but not in direct, association with the ER tubules. In optically favored cells, actin filaments were occasionally recorded located in parallel with the ER tubules and directly associated with moving particles. Streaming ceased promptly and reversibly upon treatment with cytochalasin B, which did not visibly disrupt the ER. Short-term treatment with colchicine did not inhibit streaming or disrupt the ER network, whereas long-term (hours) colchicine treatments caused the disappearance of the stationary, cortical polygonal networks and an aggregation of still slowly moving organelles and particles onto now visible actin filaments. This suggests that microtubule breakdown disrupts the three-dimensional distribution of the ER and rearranges actin filaments in the cell's cytoplasm. Actin filaments must be directly involved in generation of movement of the particles and organelles. A three-dimensional model, based on optical sectioning of the epidermal cells, is proposed to illustrate the distribution of the endoplasmic reticulum in onion epidermal cell cytoplasm.
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  • 89
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    Cell Motility and the Cytoskeleton 10 (1988), S. 172-184 
    ISSN: 0886-1544
    Keywords: chromosome movement ; spindle elongation ; micromanipulation ; mechanical properties ; mitosis ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Mechanical properties of the mitotic spindle and the effects of various operations of the mitotic apparatus on the chromosome movement and spindle elongation were investigated in fertilized eggs and blastomeres of the sand dollar, Clypeaster japonicus. On the basis of results with mechanical stretching and compression of the spindle with a pair of microneedles and the behavior of an oil drop microinjected into the spindle, it was concluded that the equatorial region of the spindle is mechanically weaker than the half-spindle region. Anaphase chromosome movement occurred in the spindle from which an aster had been removed or separated with its polar end and in the spindle in which the interzonal region had been removed. This fact indicates that chromosomes move poleward in anaphase by forces generated near the kinetochores in the half-spindle. Because of the effects of separation or removal of an aster from the spindle on the spindle elongation in anaphase and the behavior of the aster, it was concluded that the spindle elongation in anaphase is caused by pulling forces generated by asters attached to the ends of the spindle.
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  • 90
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    Cell Motility and the Cytoskeleton 11 (1988), S. 303-317 
    ISSN: 0886-1544
    Keywords: spectrin ; actin ; membrane skeleton ; cytoskeleton ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: We have used a polyclonal affinity-purified antibody made against chicken brain fodrin (both 240 and 235 Kd subunits) as a probe to determine if a fodrinlike protein exists in amoebae of Dictyostelium discoideum. In Western blots of whole cells and the isolated cell cortex, polypeptides measuring 220 and 70 Kd are recognized by the fodrin antibodies. In situ localization by indirect immunofluorescence with antifodrin indicates that the immunoreactive polypeptides are cortical. The immunoreactive analogues copatch and cocap with concanavalin A. At the level of resolution of the electron microscope, immunocytochemistry with antifodrin and colloidal gold confirms that the immunoreactive analogues are cortical proteins associated with microfilaments on the cytoplasmic side of the plasma membrane. We have isolated and characterized the 220 Kd protein to determine if it is similar to fodrin and to investigate its relationship to the 70 Kd polypeptide. The 220 Kd protein can be extracted from the cortex in the absence of detergent and isolated by gel filtration and sucrose density gradient sedimentation. The 220 Kd is a rod-shaped protein 118 ± 17.8 nm (N = 37) in length. It has a sedimentation coefficient of 9.3 S and Stokes' radius of 13 nm and exists as a dimer of approximately 500,000 daltons (Mr). Isolated 220 Kd binds to actin filaments in vitro when assayed by rotary shadowing. Morphological criteria distinguish 220 Kd from Dictyostelium myosin II heavy chain (215 Kd) and the filaminlike protein at 240 Kd. The 70 Kd polypeptide appears to be a cleavage fragment of the 220 Kd, since it is found after prolonged storage when formerly only the 220 Kd was present. Furthermore, the 220 and 70 Kd polypeptides exhibit similar one-dimensional peptide maps when treated with TPCK trypsin. On the basis of its physical and immunoreactive characteristics, and location in the cell, the 220 Kd may be a fodrinlike protein.
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    Cell Motility and the Cytoskeleton 11 (1988), S. 326-326 
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
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  • 92
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    Cell Motility and the Cytoskeleton 11 (1988), S. 318-325 
    ISSN: 0886-1544
    Keywords: actin ; contractile proteins ; microvilli ; cytoskeleton ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The mammalian genome contains 20-30 genes encoding a family of actins. To date, however, only six proteins (four muscle and two nonmuscle isoforms) encoded by this multigene complex have been identified. We have isolated two actins from the brush border of rat intestinal epithelial cells that have isoelectric points and N-terminal peptides characteristic of the cytoplasmic β- and γ-actins. However, using a panel of actin-specific monoclonal antibodies, we show that these actins contain a set of epitopes that distinguishes them from any of the known cytoplasmic or muscle isoforms. These unique actins share features of both the nonmuscle and muscle isoforms, suggesting that they represent an intermediate in the evolution of the specialized muscle actins.
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  • 93
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    Cell Motility and the Cytoskeleton 9 (1988), S. 9-16 
    ISSN: 0886-1544
    Keywords: adaptation ; cAMP ; cell motility ; chemotaxis ; Dictyostelium discoideum ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: When developing amebae of Dictyostelium discoideum are treated with constant concentrations of cAMP above 10-8 M, the average rate of motility is depressed, with maximum inhibition at roughly 10-6 M. It is demonstrated that shifting the concentration of cAMP from 0 M to concentrations ranging from 10-8 to 10-6 M in a perfusion chamber results in the immediate inhibition of motility. After shifting from 0 M to 10-8 or 10-7 M, the rate of cell motility remains low, then rebounds to a higher level, exhibiting a standard adaptation response. No adaptation is exhibited after a shift from 0 M to 10-6 M, a concentration resulting in maximum inhibition. It is demonstrated that the level of inhibition and the extent of the adaptation period are dependent upon the concentration of cAMP after the shift, and that submaximal inhibition is additive. The characteristics of adaptation in this motility response are very similar to the characteristics of adaptation for the relay system and phosphorylation of the putative cAMP receptor.
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  • 94
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    Cell Motility and the Cytoskeleton 9 (1988), S. 231-242 
    ISSN: 0886-1544
    Keywords: tubulin ; microtubules ; photobleaching ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: We have compared the exchange kinetics of fluorescein-labeled calmodulin and tubulin in the spindles of living mitotic cells at metaphase. Cultured mammalian cells in early stages of mitosis were microinjected with labeled calmodulin or tubulin and returned to an incubator to allow equilibration of the fluorescent protein with the endogenous protein pools. Calmodulin becomes concentrated in the mitotic spindle, and treatments with inhibitors of tubulin assembly show that this concentration is dependent on the presence of microtubules. The steady-state exchange rates of both tubulin and calmodulin were measured by an analysis of fluorescence redistribution after photobleaching (FRAP), using cells preequilibrated to either 26 ± 2°C or 36 ± 2°C. A pulse of laser light focused to a 5-μm diameter column was used to destroy the fluorescence at one pole of a metaphase mitotic spindle. Ratios of fluorescence intensity from the two half-spindles and from the two polar regions were calculated for each image in a post-bleach time series to determine the rates and extents of FRAP. For tubulin, we confirm earlier observations concerning the temperature dependence of the extent of FRAP, but our data do not show a significant temperature dependence for the rate of FRAP. We hypothesize that the reduced extent of tubulin FRAP at the lower temperatures is a result of microtubules that are stable to depolymerization at 26°C and are thus less likely to exchange subunits. Calmodulin's FRAP, however, does not exhibit any of the temperature dependence observed with fluorescent tubulin. At 26 ± 2°C calmodulin exchanges rapidly with the relatively stable population of microtubules, suggesting that calmodulin is bound, either directly or indirectly, to microtubule walls.
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  • 95
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    Cell Motility and the Cytoskeleton 10 (1988), S. 506-517 
    ISSN: 0886-1544
    Keywords: axoneme ; spokehead ; dynein ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: A high-molecular-weight polypeptide, named B-band, was partially purified from sea urchin sperm flagella using selective extraction, hydroxylapatite chromatography, and sucrose density gradient centrifugation. The molecular weight of the B-band was 440,000 by continuous system of sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). Sedimentation coefficient of the B-band protein was 10.5 S, and its Stokes radius was 10 nm. When examined by low-angle rotary shadowing electron microscopy, this molecule appeared to be composed of four globular heads and two curved linkers (“double headphone shape”), which was quite different from the shape of 21 S dynein, the outer arm dynein. Flagellar axonemes were also subjected to several chemical dissections. The B-band was not extracted with treatments that remove both arm structures but was solubilized with treatments that extract other components such as radial spokes and nexin links. The B-band protein in the axoneme was also more susceptible to trypsin digestion than the arm structures. These results suggest that the B-band protein is a “double headphone-shaped” component of the axonemal structures and makes up the elastic structure that might regulate the active sliding between adjacent doublet microtubules.
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    Cell Motility and the Cytoskeleton 10 (1988), S. 518-527 
    ISSN: 0886-1544
    Keywords: 9 + 2 flagellar beating ; aprotinin ; axonemes ; protease inhibitor ; sperm motility ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The effects of protease substrates and inhibitor, which have been previously shown to inhibit mammalian sperm motility (de Lamirande, E., and Gagnon, C. [1986] J. Cell Biol. 102:1378-1383), were investigated using reactivated sea urchin and carp spermatozoa as models of “9 + 2” flagella. Aprotinin in the 2 to 20 μM range interfered with sperm motility by reducing both the beat frequency and the percentage of motile spermatozoa. These inhibitory effects of aprotinin were reversible either by dilution or by the addition of high concentrations of MgATP to the incubation medium. Protease substrates with a lys-ester bond, such as N-α-benzyloxycarbonyl-lys thiobenzyl ester (BLT), also affected motility, but in the 0.1 to 0.5 mM range. As with aprotinin, both the flagellar beat frequency and the percentage of motile spermatozoa were partially and completely decreased, respectively. Analysis of the beat frequencies as a function of MgATP concentration in the presence and absence of 6 μM aprotinin indicated that this protease inhibitor affects sperm motility by decreasing the maximal flagellar beat frequency rather than by altering the axoneme's apparent Km for MgATP. Furthermore, aprotinin concentrations that blocked flagellar reactivation completely inhibited the sliding of microtubules from trypsinized axonemes. Basic proteins or polypeptides of pI close to that of aprotinin (10.3) were also potent inhibitors of the reactivation of motility. However, the characteristics of their inhibition of flagellar beat frequencies and reversibility of their effects suggested that they might be acting on sites different from those sensitive to aprotinin. The inhibitory effects of protease inhibitor and substrates, as well as results of experiments showing the absolute requirement of an intact ester bond for the inhibitory action of protease substrates, suggest that the involvement of a protease in the reactivation of 9 + 2 flagellar beating might be considered as a possible mechanism to explain aprotinin and BLT actions.
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    Cell Motility and the Cytoskeleton 10 (1988), S. 528-536 
    ISSN: 0886-1544
    Keywords: Na/K-ATPase ; keratocytes ; Xenopus laevis ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The migration of keratocytes isolated from Xenopus tadpole epidermis has been investigated in vitro. In saline the cells move with a mean speed of 5-6 μm/min. Migration is slowed down in saline with diminished sodium content and ceases in media containing not more than 4 mM sodium. Inhibition of the Na+/K+-2Cl- cotransporter by piretanide reduces the speed of migrating cells to about one-third of the control level, the same accounts to inhibition of the Na+/H+ antiport with amiloride at pH 7.2. At pH 6.6, however, amiloride only slightly influences locomotion. Depolarization of the plasma membrane by increased extracellular K+ concentration or by inhibition of the Na+/K+ pump by ouabain is only of minor influence during more than 1 h. Hyperpolarization of the cells using the sodium ionophore monensin impedes locomotion; this inhibition depends on an active Na+/K+ pump. Ionophore-mediated breakdown of the K+ gradient strictly inhibits locomotion. The experiments have shown that a continuous flux of sodium ions is indispensable for the maintenance of cell locomotion. These ions may exert their action primarily by affecting cytosolic free calcium concentration and pH.
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    Cell Motility and the Cytoskeleton 10 (1988), S. 537-537 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 10 (1988) 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 10 (1988), S. 1-1 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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