ISSN:
1432-0614
Quelle:
Springer Online Journal Archives 1860-2000
Thema:
Biologie
,
Werkstoffwissenschaften, Fertigungsverfahren, Fertigung
Notizen:
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.
Materialart:
Digitale Medien
URL:
http://dx.doi.org/10.1007/s002530050855
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