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  • 1
    Electronic Resource
    Electronic Resource
    Comparison between isoelectric focusing methods for the detection of orosomucoid phenotypes (1988)
    Weinheim : Wiley-Blackwell
    Electrophoresis 9 (1988), S. 268-272 
    Details
    ISSN: 0173-0835
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Orosomucoid (ORM) polymorphism was investigated by different methods including isoelectric focusing in acid pH ranges followed by silver staining, print immunofixation of desialyzed ORM, fixation using a lectin from the sea-weed Codium tomentosum, isoelectric focusing followed by immunofixation in miniaturized gels and isoelectric focusing in immobilized pH gradients. Population genetics studies were carried out in Galicia (NW Spain) and two new ORM variants were found.
    Additional Material: 5 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/elps.1150090605
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  • 2
    Electronic Resource
    Electronic Resource
    Determination of the specific growth of molds on semi-solid cultures (1981)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 23 (1981), S. 321-333 
    Details
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: The determination of growth constant of Aspergillus niger were obtained for semisolid cultures on cassava flour, Manihot esculenta, as a sole carbon source. As a consequence, a technique was developed that consisted of the use of a packed-bed microfermentor with a working volume of 16 cm3. The bed consisted of gelatinized and granulated cassava flour containing mineral nutrients and mold spores. The carbon dioxide produced during the respiration was drawn off with a current of air and then absorbed in a solution of sodium hydroxide. The absorption of CO2, P, was correlated with the specific growth rate μ, by means of the equation P=Keμt, where t is time and K is a constant. Ammonium nitrogen was used as a limiting substrate and its concentration was varied from 0.039 to 2.5% in dry base. The maximum growth rate, μmax, and the saturation constant, Ks, were 0.31 hr-1 and 0.065 mmol (NH4)2SO4/g total dry solids. Substrate inhibition was presented and the constant ki gave a value of 1.5 mmol (NH4)2SO4/g total dry solids. The proposed method is highly recommended for the evaluation of the semisolid fermentation of molds and for strictly aerobic bacteria and yeasts. It can be used especially in the evaluation of the growth of microorganisms on peanut shells, coffee residues, sugar cane bagasse, and other agricultural wastes.
    Additional Material: 5 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/bit.260230207
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  • 3
    Electronic Resource
    Electronic Resource
    Mutant Escherichia coli penicillin acylase with enhanced stability at alkaline pH (1995)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 48 (1995), S. 141-148 
    Details
    ISSN: 0006-3592
    Keywords: protien stability ; protein surface charge ; 6- amino-penicillanic acid ; biocatalysis ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Increased stability at alkaline pH should be a valuable attribute for the utilization of penicillin acylase in bioreactors employed to convert penicillins into 6-aminopenicillanic acid, a precursor of semisynthetic penicillins. In these systems, base is added for pH control, which results in local alkaline conditions that promote enzyme inactivation. Hydrolysis and synthesis reactions are also pH dependent. Here, we report work in which the gene coding for Escherichia coli penicillin acylase was subjected to oligonucleotide-directed random mutagenesis at regions coding for amino acids predicted to be at the surface of the enzyme. The resulting mutant library, cloned in E. coli, was screened by a filter paper assay of the colonies for the presence of penicillin acylase activity with enhanced stability at alkaline pH. Characterization of one of the selected clones revealed the presence of a mutation, Trp431-Arg, which would presumably alter the surface charge of the protein. In vitro experiments demonstrated a near twofold increase in the half-life of the mutant enzyme when stored at pH 8.5 as compared with the wild-type enzyme, with a comparable specific activity at several pH values. In general, the mutant displayed increased stability toward the basic side in the pH-stability profile. © 1995 John Wiley & Sons, Inc.
    Additional Material: 8 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/bit.260480208
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  • 4
    Electronic Resource
    Electronic Resource
    An indirect optimization method for biochemical systems: Description of method and application to the maximization of the rate of ethanol, glycerol, and carbohydrate production in Saccharomyces cerevisiae (1997)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 55 (1997), S. 758-772 
    Details
    ISSN: 0006-3592
    Keywords: Optimization ; metabolic systems ; linear programming ; S-system representation ; ethanol ; glycerol ; carbohydrates ; Saccharomyces cerevisiae ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Three metabolic models for the production of ethanol, glycerol, and carbohydrates in yeast are optimized with respect to different production rates. While originally nonlinear, all three optimization problems are reduced in such a way that methods of linear programming can be used. The optimizations lead to profiles of enzyme activities that are compatible with the physiology of the cells, which guarantees their viability and fitness, and yield higher rates of the desired final end products than the original systems. In order to increase ethanol rate production at least three times, six enzymes must be modulated. By contrast, when the production of glycerol or carbohydrates is optimized, modulation of just one enzyme (in the case of glycerol) or two enzymes (in the case of carbohydrates) is necessary to yield significant increases in product flux rate. Comparisons of our results with those obtained from other methods show great similarities and demonstrate that both are valid methods. The choice of one or the other method depends on the question of interest. © 1997 John Wiley & Sons, Inc. Biotechnol Bioeng 55: 758-772, 1997.
    Additional Material: 2 Ill.
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  • 5
    Electronic Resource
    Electronic Resource
    Recombinant microorganisms for industrial production of antibiotics (1997)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 55 (1997), S. 216-226 
    Details
    ISSN: 0006-3592
    Keywords: strain improvement ; cephalosporin ; 7-ACA ; penicillin ; β-lactam ; Streptomyces ; clavulanic acid ; tylosin ; hybrid antibiotic ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: The enhancement of industrial antibiotic yield has been achieved through technological innovations and traditional strain improvement programs based on random mutation and screening. The development of recombinant DNA techniques and their application to antibiotic producing microorganisms has allowed yield increments and the design of biosynthetic pathways giving rise to new antibiotics. Genetic manipulations of the cephalosporin producing fungus Cephalosporium acremonium have included yield improvements, accomplished increasing biosynthetic gene dosage or enhancing oxygen uptake, and new biosynthetic capacities as 7-aminocephalosporanic acid (7-ACA) or penicillin G production. Similarly, in Penicillium chrysogenum, the industrial penicillin producing fungus, heterologous expression of cephalosporin biosynthetic genes has led to the biosynthesis of adipyl-7-aminodeacetoxycephalosporanic acid (adipyl-7-ADCA) and adipyl-7-ACA, compounds that can be transformed into the economically relevant 7-ADCA and 7-ACA intermediates. Escherichia coli expression of the genes encoding D-amino acid oxidase and cephalosporin acylase activities has simplified the bioconversion of cephalosporin C into 7-ACA, eliminating the use of organic solvents. The genetic manipulation of antibiotic producing actinomycetes has allowed productivity increments and the development of new hybrid antibiotics. A legal framework has been developed for the confined manipulation of genetically modified organisms. © 1997 John Wiley & Sons, Inc. Biotechnol Bioeng 55: 216-226, 1997.
    Additional Material: 3 Ill.
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  • 6
    Electronic Resource
    Electronic Resource
    Cell-substratum adhesion strength as a determinant of hepatocyte aggregate morphology (1997)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 53 (1997), S. 415-426 
    Details
    ISSN: 0006-3592
    Keywords: hepatocytes ; cell adhesion ; spheroids ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Cultured hepatocytes typically form multicellular aggregates which are either monolayered or spheroidal in morphology. We propose that the aggregate morphology resulting from a particular cell-substratum interaction has a biophysical basis: when cell contractile forces are greater than cell-substratum adhesion forces, spheroidal aggregates form; when cell contractile forces are weaker than cell-substratum adhesion forces, cells remain essentially spread and form monolayered aggregates. We tested this hypothesis by systematically varying the morphology of hepatocellular aggregates formed on substrata coated with a series of different concentrations of Matrigel, and correlating aggregate morphology with the cell-substratum adhesion strength measured in a shear flow detachment assay. Aggregate morphology was binary - spheroidal aggregates formed at low Matrigel concentrations and monolayered aggregates formed at high Matrigel concentrations. Cell-substratum adhesion strength was similarly binary, with low adhesion strengths correlated with spheroidal aggregates and high adhesion strengths correlated with formation of monolayered aggregates. © 1997 John Wiley & Sons, Inc. Biotechnol Bioeng 53: 415-426, 1997.
    Additional Material: 6 Ill.
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  • 7
    Electronic Resource
    Electronic Resource
    Quantitative aspects of silver deposition in proteins resolved in complex polyacrylamide gels (1982)
    Weinheim : Wiley-Blackwell
    Electrophoresis 3 (1982), S. 197-205 
    Details
    ISSN: 0173-0835
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Deposition of silver or silver compounds in proteins separated in linear gradient polyacrylamide electrophoretic gels is influenced by protein concentration and location, polyacrylamide concentration, extent of washing after incubation in ammoniacal silver hydroxide solution, and development time in a citric acid: formaldehyde solution. Our modification of existing silver staining methods provided routine resolution of polypeptides separated in 1.0 to 1.5-mm-thick gels without high background staining or surface staining and with picogram detection sensitivity. Removing electrophoretic components, such as sodium dodecyl sulfate (SDS), sulfhydryl reagents (2-mercaptoethanol, dithiothreitol and dithioerythreitol), glycerol, tris-glycine, carrier ampholytes, urea, and acetic acid, was essential in order to attain silver staining of proteins with a negligible background. Ammoniacal silver ions or silver ions, associated and unassociated with proteins separated in polyacrylamide gels, were removed during the washing step following incubation in ammoniacal silver hydroxide solution. Loss of stain intensity at discrete protein clusters was shown to correlate with polyacrylamide concentration in gradient gels. Standardizing the washing step enhanced reproducibility of staining. Using this silver staining method and computer-assisted image processing, it was possible to detect quantitatively as little as 27 picograms of protein per mm in 10 %-20 % linear gradient polyacrylamide gels (1.2 mm thick). Based on the lowest detection limits for Coomassie Brilliant Blue R-250 staining of protein (10 nanograms), this method for silver staining is at least 370-fold more sensitive.
    Additional Material: 9 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/elps.1150030404
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  • 8
    Electronic Resource
    Electronic Resource
    On the relationship of amino acid composition to silver staining of proteins in electrophoresis gels: II. Peptide sequence analysis (1991)
    Weinheim : Wiley-Blackwell
    Electrophoresis 12 (1991), S. 409-414 
    Details
    ISSN: 0173-0835
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: The quantification of proteins in silver-stained electrophoresis gels has been limited by the differences in “stainability” of different proteins. Despite efforts by many researchers, the precise basis of the reaction between silver reagents and polypeptides is still unclear, and, depending on the formulation, may even differ. We have tested the hypothesis that differences in stainability among proteins can be attributed to diferences in di- or tripeptide composition. The results indicate that some order of protein structure other than short peptides accounts for the staining differences observed.
    Additional Material: 3 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/elps.1150120605
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  • 9
    Electronic Resource
    Electronic Resource
    Electrophoretic human leukocyte antigen HLA-DQA1 DNA typing after polymerase chain reaction amplification (1991)
    Weinheim : Wiley-Blackwell
    Electrophoresis 12 (1991), S. 1041-1045 
    Details
    ISSN: 0173-0835
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: An electrophoretic method (sodium dodecyl sulfate-polyacrylamide gel electrophoresis) is described which permits the identification of human leukocyte antigen HLA-DQA1 types and subtypes without using allele-specific oligonucleotide prodes and dot-blot methodology. The procedure can be used in miniaturized gels in combination with automated electrophoretic systems. The PhastSystem is particularly recommended since temperature contol is essential. HLA-DQA1*1 and DQA1*0301 can be distinguished in homoduplexes and DQA1*01 subtypes, DQA1*201 and DQA1*0401 in heteroduplexes (in only 5 including DNA extraction and PCR amplification). Additional variations to those recognized using commercially available dot-blot methods can be provided since this this procedure permits the identification of single base-pair substitutions. In addition, this method is faster and less expensive than commercial methods.
    Additional Material: 5 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/elps.1150121208
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  • 10
    Electronic Resource
    Electronic Resource
    The use of radioactive bacteriophage proteins as X-Y markers for silver stained two-dimensional electrophoresis gels and quantification of the patterns (1992)
    Weinheim : Wiley-Blackwell
    Electrophoresis 13 (1992), S. 87-92 
    Details
    ISSN: 0173-0835
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: We have demonstrated the feasibility of using bacteriophage ghost proteins, tritiated by metabolic labeling, as a set of standard markers for two-dimensional gels in which the proteins are to be detected by silver staining. The results indicate that a 2.5 μg load of phage proteins yields a reproducible silver pattern of 48 spots. The spots can also be readily identified by radioautography and radiofluorography, establishing their value as a standard constellation of markers. Quantification of these patterns by computerized densitometry indicates a general agreement between detection by silver staining and detection by radiofluorography.
    Additional Material: 3 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/elps.1150130117
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