Cell engineering of Escherichia coli allows high cell density accumulation without fed-batch process control.
A set of mutations in the phosphoenolpyruvate:carbohydrate phosphotransferase system (PTS) was used to create Escherichia coli strains with a reduced uptake rate of glucose. This allows a growth restriction, which is controlled on cellular rather than reactor level, which is typical of the fed-batch cultivation concept. Batch growth of the engineered strains resulted in cell accumulation profiles corresponding to a growth rate of 0.78, 0.38 and 0.25 h(-1), respectively. The performance of the mutants in batch cultivation was compared to fed-batch cultivation of the wild type cell using restricted glucose feed to arrive at the corresponding growth profiles. Results show that the acetate production, oxygen consumption and product formation were similar, when a recombinant product was induced from the lacUV5 promoter. Ten times more cells could be produced in batch cultivation using the mutants without the growth detrimental production of acetic acid. This allows high cell density production without the establishment of elaborate fed-batch control equipment. The technique is suggested as a versatile tool in high throughput multiparallel protein production but also for increasing the number of experiments performed during process development while keeping conditions similar to the large-scale fed-batch performance.
Bäcklund
Emma
E
School of Biotechnology, AlbaNova University Center, KTH, 106 91 Stockholm, Sweden.
Markland
Katrin
K
Larsson
Gen
G
D019342:Acetic Acid / Q000378:metabolism; D003470:Culture Media; D004926:Escherichia coli / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D005818:Genetic Engineering; D005947:Glucose / Q000378:metabolism; D010101:Oxygen Consumption
D003470:Culture Media; D005947:Glucose; D019342:Acetic Acid