Effects of post-induction feed strategies on secretory production of recombinant streptokinase in Escherichia coli

Ramalingam, Subramanian; Pennathur, Gautam; Mukherjee, Kakali; Jayaraman, Guhan

doi:10.1016/j.bej.2006.09.019

Cited by 40 publications

(23 citation statements)

References 44 publications

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“…On the other hand, there are few reports that demonstrate the absence of any correlation between these two factors [6][7][8]. Some authors have claimed that rather than the preinduction l, it is a high postinduction l, which helps in achieving high-level production of recombinant protein [9], whereas others have shown that the postinduction l is an intrinsic property of the cell that cannot be controlled by bioprocess strategies [10,11]. However, one problem associated with some of these reports is that they have looked at overall expression levels rather than the specific product formation rate (q p ), which is a better measure of product formation kinetics.…”

Section: Introductionmentioning

confidence: 91%

Kinetic studies of recombinant human interferon-gamma expression in continuous cultures of E. coli

Vaiphei

Pandey

Mukherjee

2009

J Ind Microbiol Biotechnol

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A series of continuous cultures was performed to understand the product formation kinetics of recombinant human interferon gamma (rhIFN-gamma) in Escherichia coli at different dilution rates ranging from 0.1 to 0.3 h(-1) in different media. A T7 promoter-based vector was used for expression of IFN-gamma in E. coli BL21 (DE3) cells. The recombinant protein was produced as inclusion bodies, thus allowing a rapid buildup of rhIFN-gamma inside the cell, with the specific product yield (Y(p/X)) reaching a maximum value of 182 mg g(-1) dry cell weight (DCW). In all the media tested, the specific product formation rate (q(p)) was found to be strongly correlated with the specific growth rate (mu), demonstrating the growth-associated nature of product formation. The q(p) values show no significant decline with time postinduction, even though the recombinant protein has been over produced inside the cell. The maximum q(p) level of 75.5 mg g(-1) h(-1) was achieved at the first hour of induction at the dilution rate of 0.3 h(-1). Also, this correlation between q(p) and mu was not critically dependent on media composition, which would made it possible to grow cells in defined media in the growth phase and then push up the specific growth rate just before induction by pulse addition of glucose and yeast extract. This would ensure the twin objectives of high biomass and high specific productivities, leading to high volumetric product concentration.

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Section: Introductionmentioning

confidence: 91%

Kinetic studies of recombinant human interferon-gamma expression in continuous cultures of E. coli

Vaiphei

Pandey

Mukherjee

2009

J Ind Microbiol Biotechnol

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“…Extensive work has been carried out on recombinant streptokinase expression in the E. coli host with upto 25,000 IU/mL of the recombinant protein. 38 The secretion of glycosylated streptokinase has been reported in P. pastoris (3200 IU/mL) with an improved proteolytic resistance and stability. 35 Recently, 4.25 g/L of bioactive streptokinase was produced in P. pastoris with a volumetric productivity of 3727 IU/mL/h.…”

Section: First Generation Plasminogen Activatorsmentioning

confidence: 99%

The evolution of recombinant thrombolytics: Current status and future directions

Adivitiya

Khasa

2016

Bioengineered

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Cardiovascular disorders are on the rise worldwide due to alcohol abuse, obesity, hypertension, raised blood lipids, diabetes and age-related risks. The use of classical antiplatelet and anticoagulant therapies combined with surgical intervention helped to clear blood clots during the inceptive years. However, the discovery of streptokinase and urokinase ushered the way of using these enzymes as thrombolytic agents to degrade the fibrin network with an issue of systemic hemorrhage. The development of second generation plasminogen activators like anistreplase and tissue plasminogen activator partially controlled this problem. The third generation molecules, majorly t-PA variants, showed desirable properties of improved stability, safety and efficacy with enhanced fibrin specificity. Plasmin variants are produced as direct fibrinolytic agents as a futuristic approach with targeted delivery of these drugs using liposome technlogy. The novel molecules from microbial, plant and animal origin present the future of direct thrombolytics due to their safety and ease of administration.

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“…After the exhaustion of glucose, fed-batch experiments were performed by feeding the solutions containing glucose (100 g/l), yeast extract (100 g/l) and ampicillin 100 mg/l. Concentration of glucose in the medium was monitored at regular interval of time and fed by intermittent pulse feeding in proportional to the cell mass [17]- [19]. Feeding of glucose with specific feed rate of 0.5 g glucose per g DCW/h and 0.25 g glucose per g DCW/h was carried out in the growth phase and production phase, respectively and the residual glucose was maintained below 1.0 g/l [19] [20].…”

Section: Fed-batch Cultivationmentioning

confidence: 99%

“…Concentration of glucose in the medium was monitored at regular interval of time and fed by intermittent pulse feeding in proportional to the cell mass [17]- [19]. Feeding of glucose with specific feed rate of 0.5 g glucose per g DCW/h and 0.25 g glucose per g DCW/h was carried out in the growth phase and production phase, respectively and the residual glucose was maintained below 1.0 g/l [19] [20]. The biomass concentration in the bioreactor was calculated throughout the fermentation, by measuring the optical density (OD) as well as the dry cell weight.…”

Section: Fed-batch Cultivationmentioning

confidence: 99%

Batch, Fed Batch Production and Characterization of Glutaminase Free L-Asparaginase II of <i>Pectobacterium carotovorum</i> MTCC 1428 in <i>Escherichia coli</i>

Goswami¹,

Hegde²,

Dasu³

2014

AiM

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The present study describes the production optimization of recombinant L-asparaginase II of Pectobacterium carotovorum MTCC 1428 in Escherichia coli BL21 (DE3) at batch and fed batch bioreactor level. Production of recombinant L-asparaginase II in batch and fed batch mode was found to be 1.34 and 5.38 folds higher, respectively as compared to shake flask culture. SDS-PAGE and native PAGE of the purified enzyme revealed that molecular mass of the subunits and native enzyme are ~37.5 kDa and ~150 kDa, respectively. Optimum range of pH and temperature for hydrolysis of L-asparagine were found to be 7.5 -8.5 and 47˚C -52˚C, respectively. The recombinant enzyme is very specific for its natural substrate, L-asparagine. The activity of recombinant L-asparaginase II is improved by mono cations and diverse effectors including Na + , K + , L-cystine, L-histidine, glutathione and 2-mercaptoethanol whereas, it is moderately inhibited by different divalent cations and thiol group blocking reagent. The kinetic parameters Km, Vmax, kcat and Km/Kcat of purified recombinant L-asparaginase II were determined. The purified L-asparaginase II possesses no partial glutaminase activity, which is prerequisite to reduce the possibility of side effects during the course of anti-cancer therapy.

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Effects of post-induction feed strategies on secretory production of recombinant streptokinase in Escherichia coli

Cited by 40 publications

References 44 publications

Kinetic studies of recombinant human interferon-gamma expression in continuous cultures of E. coli

Kinetic studies of recombinant human interferon-gamma expression in continuous cultures of E. coli

The evolution of recombinant thrombolytics: Current status and future directions

Batch, Fed Batch Production and Characterization of Glutaminase Free L-Asparaginase II of <i>Pectobacterium carotovorum</i> MTCC 1428 in <i>Escherichia coli</i>

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Effects of post-induction feed strategies on secretory production of recombinant streptokinase in Escherichia coli

Cited by 40 publications

References 44 publications

Kinetic studies of recombinant human interferon-gamma expression in continuous cultures of E. coli

Kinetic studies of recombinant human interferon-gamma expression in continuous cultures of E. coli

The evolution of recombinant thrombolytics: Current status and future directions

Batch, Fed Batch Production and Characterization of Glutaminase Free L-Asparaginase II of &lt;i&gt;Pectobacterium carotovorum&lt;/i&gt; MTCC 1428 in &lt;i&gt;Escherichia coli&lt;/i&gt;

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Batch, Fed Batch Production and Characterization of Glutaminase Free L-Asparaginase II of <i>Pectobacterium carotovorum</i> MTCC 1428 in <i>Escherichia coli</i>