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  • 1995-1999  (6)
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Material
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Year
  • 1
    Electronic Resource
    Electronic Resource
    Maximizing the expression of recombinant Kringle 1 (Streptokinase) synthesized in Escherichia coli. Influence of culture and induction conditions (1996)
    Springer
    Biotechnology letters 18 (1996), S. 1261-1266 
    Details
    ISSN: 1573-6776
    Source: Springer Online Journal Archives 1860-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
    Notes: Summary We examined the influence of culture conditions on the production of the recombinant fused protein Kringle 1 (Streptokinase) (K1(SK)) expressed in E. coli under the control of the Ipp-lac promoter. The growth and the protein production were severely inhibited at glucose concentrations higher than 5 g/L. The highest yield of K1(SK) from the soluble extract was obtained using synthetic medium. Concentrations of inducer between 0.1 and 1 mM of IPTG did not significantly affect the production level of K1(SK) or the final cell density. Under optimal conditions the K1(SK) protein was expressed in an intracellular soluble form at about 3–4% of the total cellular protein.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00129951
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    Paper (German National Licenses)
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  • 2
    Electronic Resource
    Electronic Resource
    High volumetric yields of functional dimeric miniantibodies in Escherichia coli, using an optimized expression vector and high-cell-density fermentation under non-limited growth conditions (1996)
    Springer
    Applied microbiology and biotechnology 46 (1996), S. 524-532 
    Details
    ISSN: 1432-0614
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Functional bivalent miniantibodies, directed against the epidermal growth factor receptor, accumulated to more than 3 gl−1 in high-cell-density cultures of Escherichia coli RV308(pHKK) on a pilot scale. The miniantibodies consist of scFv fragments with a C-termi-nal hinge followed by a helix-turn-helix motif, which homodimerizes in vivo. The improved expression vector pHKK is characterized by the hok/sok suicide system, improving plasmid maintenance, and the inducible lac p/o promoter system with the very strong T7g10 Shine-Dalgarno sequence. The expression unit is flanked by terminators. The prototrophic RV308 cells were cultivated in glucose mineral salt medium and reached a cell density of 145 g dry biomass l−1 after 33 h. After induction, growth continued almost unchanged for a further 4 h with concomitant miniantibody formation. In the fed-batch phase, the concentration of glucose was kept almost constant at the physiological level of approximately 1.5 g l−1, using on-line flow injection analysis for control. Surprisingly, E. coli RV308(pHKK) did not accumulate significant amounts of the metabolic by-product acetate under these unlimited aerobic growth conditions.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s002530050855
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    Paper (German National Licenses)
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  • 3
    Electronic Resource
    Electronic Resource
    Influence of stringent and relaxed response on excretion of recombinant proteins and fatty acid composition in Escherichia coli (1995)
    Springer
    Applied microbiology and biotechnology 43 (1995), S. 89-92 
    Details
    ISSN: 1432-0614
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract In contrast to stringent (relA+) cells of Escherichia coli, relaxed (relA) cells excreted recombinant proteins (β-lactamase, interferon α1) into the culture medium during amino acid limitation. Comparative analyses of overall fatty acid composition in relA+ cells and relA cells were performed and revealed that, in wild-type cells, drastic alterations occurred during the stringent response. The portion of saturated fatty acids (C16:0) and the fractions of cyclopropane fatty acids (C17cyc and C19cyc) increased whereas the portions of unsaturated fatty acids (C16:1 and C18:1) decreased. In cells of the relaxed mutant, no significant changes in the overall composition of the fatty acids were observed after the onset amino acid limitation. These results indicate that a change in fatty acid composition of membrane lipids after starvation of cells may be responsible for the prevention of loss of cellular proteins into the culture medium in stringent controlled cells of Escherichia coli.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00170628
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    Paper (German National Licenses)
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  • 4
    Electronic Resource
    Electronic Resource
    Influence of stringent and relaxed response on excretion of recombinant proteins and fatty acid composition in Escherichia coli (1995)
    Springer
    Applied microbiology and biotechnology 43 (1995), S. 89-92 
    Details
    ISSN: 1432-0614
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract  In contrast to stringent (relA+) cells of Escherichia coli, relaxed (relA) cells excreted recombinant proteins (β-lactamase, interferon α1) into the culture medium during amino acid limitation. Comparative analyses of overall fatty acid composition in relA+ cells and relA cells were performed and revealed that, in wild-type cells, drastic alterations occurred during the stringent response. The portion of saturated fatty acids (C16 : 0) and the fractions of cyclopropane fatty acids (C17cyc and C19cyc) increased whereas the portions of unsaturated fatty acids (C16 : 1 and C18 : 1) decreased. In cells of the relaxed mutant, no significant changes in the overall composition of the fatty acids were observed after the onset of amino acid limitation. These results indicate that a change in fatty acid composition of membrane lipids after starvation of cells may be responsible for the prevention of loss of cellular proteins into the culture medium in stringent controlled cells of Escherichia coli.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s002530050375
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    Paper (German National Licenses)
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  • 5
    Electronic Resource
    Electronic Resource
    High-cell-density cultivation of microorganisms (1999)
    Springer
    Applied microbiology and biotechnology 51 (1999), S. 422-430 
    Details
    ISSN: 1432-0614
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract High-cell-density cultivation (HCDC) is required to improve microbial biomass and product formation substantially. An overview of HCDC is given for microorganisms including bacteria, archae and eukarya (yeasts). Problems encountered by HCDC and their possible solutions are discussed. Improvements of strains, different types of bioreactors and cultivation strategies for successful HCDC are described. Stirred-tank reactors with and without cell retention, a dialysis-membrane reactor, a gas-lift reactor and a membrane cyclone reactor used for HCDC are outlined. Recently modified traditional feeding strategies and new ones are included, in particular those for unlimited growth to very dense cultures. Emphasis is placed on robust fermentation control because of the growing industrial interest in this field. Therefore, developments in the application of multivariate statistical control, artificial neural networks, fuzzy control and knowledge-based supervision (expert systems) are summarized. Recent advances using Escherichia coli– the pioneer organism for HCDC – are outlined.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s002530051412
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    Paper (German National Licenses)
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  • 6
    Electronic Resource
    Electronic Resource
    Expression and secretion of functional miniantibodies McPC603scFvDhlx in cell-wall-less L-form strains of Proteus mirabilis and Escherichia coli : A comparison of the synthesis capacities of L-form strains with an E. coli producer strain (1998)
    Springer
    Applied microbiology and biotechnology 49 (1998), S. 51-58 
    Details
    ISSN: 1432-0614
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract The paper describes the synthesis of the phosphorylcholine-binding miniantibody McPC603scFvDhl x in cell-wall-less L-form strains of Escherichia coli and Proteus mirabilis. Cells of these strains were transformed with the plasmid pACK02scKan, carrying the miniantibody (miniAb) coding sequence under the control of the lac promoter. L-form transformants of both species were able to synthesize the functional miniAb as an extracellular soluble product. The highest quantities were obtained by P. mirabilis L-form strains after induction with 5 mM isopropyl β-d-thiogalactopyranoside (IPTG). Yields of 45–75 mg/l total antibody protein and of 10–18 mg/l functional miniAb were estimated in the growth medium of shaking cultures 40–80 h after induction with IPTG. About 10% of the active miniAb remained cell-bound. The yields of functional miniAb could be optimized by lowering the growth temperature from 37 °C to 26–32 °C and by supplementation of the medium with 80 mM sodium fumarate. A comparison of the specific activities revealed that the P. mirabilis L-form strains have a similar synthesis capacity (2–4 mg functional miniAb/g cell dry weight) to that of the producer strain E. coli RV308. The results show that the processes of correct folding and assembling of the miniAb molecules are possible without the periplasmic compartment.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s002530051136
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    Paper (German National Licenses)
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