Effect of glucose supply strategy on acetate accumulation, growth, and recombinant protein production by Escherichia coli BL21 (λDE3) and Escherichia coli JM109

Shiloach, Joseph; Kaufman, Joyce J.; Guillard, Anne-Sophie; Fass, R.

doi:10.1002/(sici)1097-0290(19960220)49:4<421::aid-bit9>3.0.co;2-r

Cited by 140 publications

(86 citation statements)

References 22 publications

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“…In such cases, TCS083 should perform better than TCS062, since acetateproducing pathways are disrupted. Indeed, shake flask experiments, in which completely aerobic conditions cannot be achieved, have already confirmed that TCS083 does indeed outperform not only TCS062 and the wild-type strain but also the industrial strain BL21 (38) and the reduced-genome strain MDS42 (32) typically used for protein production (Fig. 6).…”

Section: Discussionmentioning

confidence: 79%

Minimal Escherichia coli Cell for the Most Efficient Production of Ethanol from Hexoses and Pentoses

Trinh

Unrean

Srienc

2008

Appl Environ Microbiol

261

259

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To obtain an efficient ethanologenic Escherichia coli strain, we reduced the functional space of the central metabolic network, with eight gene knockout mutations, from over 15,000 pathway possibilities to 6 pathway options that support cell function. The remaining pathways, identified by elementary mode analysis, consist of four pathways with non-growth-associated conversion of pentoses and hexoses into ethanol at theoretical yields and two pathways with tight coupling of anaerobic cell growth with ethanol formation at high yields. Elimination of three additional genes resulted in a strain that selectively grows only on pentoses, even in the presence of glucose, with a high ethanol yield. We showed that the ethanol yields of strains with minimized metabolic functionality closely matched the theoretical predictions. Remarkably, catabolite repression was completely absent during anaerobic growth, resulting in the simultaneous utilization of pentoses and hexoses for ethanol production.

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Section: Discussionmentioning

confidence: 79%

Minimal Escherichia coli Cell for the Most Efficient Production of Ethanol from Hexoses and Pentoses

Trinh

Unrean

Srienc

2008

Appl Environ Microbiol

261

259

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“…Accumulation of acetate depends on the medium composition and on the strain and is connected to the growth and carbon source uptake rates. E. coli BL21 was derived from an E. coli B strain reported to be a low acetate producer compared to the E. coli K12 strain [26,27]. For all fed-batch cultures, acetate concentration in the culture medium was below 2.5 g l )1 .…”

Section: Resultsmentioning

confidence: 99%

Process development for production of recombinant human interferon-? expressed in Escherichia coli

Khalilzadeh

Shojaosadati

Maghsoudi

et al. 2004

Journal of Industrial Microbiology and Biotechnology

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A simple fed-batch process was carried out using constant and variable specific growth rates for high-cell-density cultivation of Escherichia coli BL21 (DE3) expressing human interferon-gamma(hIFN-gamma). The feeding rate was adjusted to achieve an appropriate specific growth rate. The dissolved oxygen level was maintained at 20-30% of air saturation by control of airflow and stirrer speed and, where necessary, by enrichment of inlet air with pure oxygen. Glucose was the sole source of carbon and energy and was provided by following a simple exponential feeding rate. The final cell density in the fed-batch fermentation with constant and variable specific growth rate feeding strategies was ~100 g dry cell wt l(-1) after 36 and 20 h, respectively. The final specific yield and overall productivity of recombinant hIFN-gamma in the variable specific growth rate strategy were 0.35 g rHu-IFN-gamma g(-1) dry cell wt and 0.9 g rHu-IFN-gamma l(-1) h(-1), respectively. A new chromatographic purification procedure involving anion exchange and cation exchange chromatographies was developed for purification of rHu-IFN-gamma from inclusion bodies. The established purification process is reproducible and the total recovery of rHu-IFN-gamma was ~30% (100 mg rHu-IFN-gamma g(-1) dry cell wt). The purity of the rHu-IFN-gamma was determined using HPLC. Sterility, pyrogenicity, and DNA content tests were conducted to assure the absence of toxic materials and other components of E. coli in the final product. The final purified rHu-IFN-gamma has a specific antiviral activity of ~2 x 10(7) IU/mg protein, as determined by viral cytopathic effect assay. These results certify the product for clinical purposes.

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“…Also, this system has been well studied in our lab for expressing b-galactosidase [13]. We decided to use E. coli BL21(DE3) cells, which have been reported to utilize glucose very efficiently, so that very little amount of acetic acid is formed [15]. This strain was transformed with two plasmids, pSSY4, which contained the skc gene under the T7 promoter, and pGP1-2, which contained the T7 RNA polymerase under the heat-inducible k PL promoter and used for studies in continuous culture.…”

Section: Resultsmentioning

confidence: 99%

Continuous-culture studies on the stability and expression of recombinant streptokinase in Escherichia coli

Yazdani

Mukherjee

2002

Bioprocess and Biosystems Engineering

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A series of continuous-culture runs were conducted on a T7-expression-based two-plasmid system for heat induction of recombinant streptokinase. Different dilution rates were used and it was observed that a high dilution rate of 0.3 h -1 , both before and after induction, helped in increasing the product concentration to a fairly high level of 421 plasmin units per ml. Accumulation of active streptokinase, which is toxic to the cells, in the periplasm also resulted in the loss of cell viability. Thus, a slow washout was observed after induction and in just 2.5 h all the growing cells were those which had lost the ability to produce streptokinase. Increasing the selection pressure by using high dosage of antibiotic concentration (5·) in the feed helped in reducing the instability of the culture.

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Effect of glucose supply strategy on acetate accumulation, growth, and recombinant protein production by Escherichia coli BL21 (λDE3) and Escherichia coli JM109

Cited by 140 publications

References 22 publications

Minimal Escherichia coli Cell for the Most Efficient Production of Ethanol from Hexoses and Pentoses

Minimal Escherichia coli Cell for the Most Efficient Production of Ethanol from Hexoses and Pentoses

Process development for production of recombinant human interferon-? expressed in Escherichia coli

Continuous-culture studies on the stability and expression of recombinant streptokinase in Escherichia coli

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