ISSN:
0006-3592
Keywords:
single-chain Fv antibody fragment
;
recombinant Escherichia coli
;
periplasmic space
;
fed-batch fermentation
;
yeast extract
;
Chemistry
;
Biochemistry and Biotechnology
Source:
Wiley InterScience Backfile Collection 1832-2000
Topics:
Biology
,
Process Engineering, Biotechnology, Nutrition Technology
Notes:
Fed-batch fermentation for production of a single-chain Fv antibody fragment (scFv) expressed as a recombinant periplastic protein from Escherichia coli was investigated. A high cell density of 50 g dry cell weight per liter was routinely achieved in a 14-L vessel by controlled exponential feeding of glucose to impose a constant specific growth rate. Following biomass accumulation, induction of the tac promoter by addition of IPTG was accompaied by a linear feed of yeast extract. The concentration of yeast extract feed was found to be highly influential upon both concentration and location of active product. Although scFv fragments were specifically targeted to the periplasmic space, at yeast extract feed rates of 0.72 g/h the final location was largely extracellular (68% to 79%). Total concentrations (extracellular + periplasmic) were of the order of 5 to 8 mg/L. A ten-fold increase in yeast extract supply increased total scFv concentration to almost 200 mg/L and 78% of this yield was retained in the periplasm. Control of such leakage of the recombinant product is fundamental to process design of downstream operations for product recovery. © 1997 John Wiley & Sons, Inc. Biotechnol Bioeng 53: 611-622, 1997.
Additional Material:
4 Ill.
Type of Medium:
Electronic Resource
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