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Digitale Medien

Production of recombinant herpes simplex virus protease in 10-L stirred vessels using a baculovirus–insect cell expression system (1997)

Schwartz, J L ; Ferrari, E B ; Terracciano, J ; [weitere]

Springer

Journal of industrial microbiology and biotechnology 19 (1997), S. 87-91

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ISSN: 1476-5535

Schlagwort(e): Keywords: baculovirus–insect cell expression vector system (BEVS); Sf-9; HSV protease; glutathione-S-transferase

Quelle: Springer Online Journal Archives 1860-2000

Thema: Biologie , Werkstoffwissenschaften, Fertigungsverfahren, Fertigung

Notizen: A gene expression system using recombinant Autographa californica nuclear polyhedrosis virus (baculovirus) and Sf-9 cells has been scaled up to the 10-L tank level and shown to be capable of producing herpes simplex virus (HSV) protease in serum-free media. High densities of Spodoptera frugiperda (Sf-9) cells were achieved by modifying two 10-L Biolafitte fermenters specifically for insect cell growth. The existing Rushton impellers were replaced by marine impellers to reduce shear and the aeration system was modified to allow external addition of air/O2 mixtures at low flow rates through either the sparge line or into the head space of the fermenter. To inoculate the tanks, Sf-9 cells were adapted to grow to high cell densities (6–10 × 106 cells ml−1) in shake flasks in serum-free media. With these procedures, cell densities of 5 × 106 cells ml−1 were routinely achieved in the 10-L tanks. These cells were readily infected with recombinant baculovirus expressing the 247-amino acid catalytic domain of the HSV-1 strain 17 protease UL26 gene as a glutathione-S-transferase (GST) fusion protein (GST-247). Three days after infection at a multiplicity of infection (MOI) of 3 pfu cell−1, the GST-247 fusion protein was purified from a cytoplasmic lysate by Glutathione Sepharose 4-B affinity chromatography with reproducible yields of 11–38 mg L−1 of recombinant protein and ≥ 90% purity. Maximum production of this protein was observed at a cell density of 5.0 × 106 cells ml−1.

Materialart: Digitale Medien

URL: http://dx.doi.org/10.1038/sj.jim.2900446

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Digitale Medien

Naturally produced isocoumarins: inhibitors of calmodulinsensitive cyclic guanosine 3′,5′-monophosphate phosphodiesterase (1989)

Hegde, V. R. ; Wittreich, H. ; Patel, M. G. ; [weitere]

Springer

Journal of industrial microbiology and biotechnology 4 (1989), S. 209-213

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ISSN: 1476-5535

Schlagwort(e): Isocoumarin ; Phosphodiesterase ; Enzyme inhibitor ; cGMP phosphodiesterase ; Secondary metabolite

Quelle: Springer Online Journal Archives 1860-2000

Thema: Biologie , Werkstoffwissenschaften, Fertigungsverfahren, Fertigung

Notizen: Summary Three isocoumarins have been isolated from a strain ofStreptoverticillium sp. and all inhibit the calmodulin-sensitive cyclic guanosine 3′,5′-monophosphate phosphodiesterase (EC 3.1.4.17, Boehringer Mannheim). Two of the compounds, 6,8-dihydroxy-7-methoxy-3-methyl isocoumarin and 6,7,8-trihydroxy-3-methyl isocoumarin have previously been isolated fromStreptomyces. The third fermentation product, 6,8-dihydroxy-3-methyl isocoumarin, was also found as a metabolite ofCeratocystis minor, a fungal species associated with the blue stain disease of pine [2,3].

Materialart: Digitale Medien

URL: http://dx.doi.org/10.1007/BF01574078

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