Improving the cost effectiveness of enhanced green fluorescent protein production using recombinant Escherichia coli BL21 (DE3): Decreasing the expression inducer concentration

Abstract

Green fluorescent protein (GFP) is a globular protein used as biosensor and biomarker in medical and industrial fields. However, due to the expensive production costs of expressing proteins using high-cost inducers like isopropyl-β-D-1thiogalactopyranoside (IPTG), the number of GFP applications are still scarce. This work studied the production of enhanced GFP (EGFP) using Escherichia coli BL21 (DE3) [pLysS; pET28(a)], aiming to increase its yield and reduce costs. First, the influence of agitation rate, induction time, and concentration of IPTG in the production of EGFP was evaluated, but only the first two parameters were significant. Subsequently, aiming to reduce costs related to the use of inducer, the IPTG concentration (0.005, 0.010, and 0.025 mM) was decreased and, interestingly, the production levels were maintained or increased. These results show that a proper choice of production conditions, particularly through the decrease of inducer concentration, is effective to reduce the upstream production costs and guarantee high EGFP expression.

Economic analysis of the production and recovery of green fluorescent protein using ATPS-based bioprocesses

“…In the last years, some studies are aiming to decrease GFP price by developing new ways to produce it successfully [15,16]. Mainly two strategies are in place, with the first one focusing on improvements in upstream processes by obtaining high titres using recombinant Escherichia coli, the current available commercial product, [17] or tobacco plants [18] to express the protein. Alternatively, there are also studies aiming to reduce costs by improving the recovery and purification techniques in the downstream process as they can represent a large fraction of the production costs [19,20].…”

Galactitol Transport Factor GatA Relieves ATP Supply Restriction to Enhance Acid Tolerance of Escherichia coli in the Two-Stage Fermentation Production of D-Lactate

“…When IPTG is added at low cell densities, the metabolism of the target protein is assumed to harm normal bacterial growth under normal pH condition value 7.0, leading to a decrease in OD 600 [40,41]. Similarly, in this study, when 200 mM IPTG was added during LLP, the recombinant plasmid was activated prematurely and cell growth was retarded thereby negatively affecting D-LA biosynthesis, and the concentration of D-LA was reduced to 36.4 g•L −1 (Figure 4A).…”

Recombinant protein linker production as a basis for non-invasive determination of single-cell yeast age in heterogeneous yeast populations

“…The total protein content of the fermentation was calculated as 75.3 mg L À1 in E. coli BL21 (DE3) culture broth, an amount which was consistent with previously reported yields. 63,64 According to the uorescence and peak area of the HPLC analysis, the protein purity was determined >95%.…”