Department of Bioprocesses and Biotechnology, School of Pharmaceutical Sciences, São Paulo State University (UNESP), Araraquara, Brazil.
Faculty of Bioscience Engineering, Gent University, Gent, Belgium.
Biotechnol Appl Biochem. 2019 Jul;66(4):527-536. doi: 10.1002/bab.1749. Epub 2019 Apr 16.
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-1-thiogalactopyranoside (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.
绿色荧光蛋白(GFP)是一种球形蛋白,在医学和工业领域被用作生物传感器和生物标志物。然而,由于使用异丙基-β-D-1-硫代半乳糖苷(IPTG)等成本高昂的诱导剂来表达蛋白质的生产成本高昂,GFP 的应用数量仍然很少。本研究使用大肠杆菌 BL21(DE3)[pLysS;pET28(a)]来生产增强型 GFP(EGFP),旨在提高其产量并降低成本。首先,评估了搅拌速度、诱导时间和 IPTG 浓度对 EGFP 生产的影响,但只有前两个参数是显著的。随后,为了降低与诱导剂使用相关的成本,降低了 IPTG 浓度(0.005、0.010 和 0.025mM),有趣的是,生产水平得到了维持或提高。这些结果表明,适当选择生产条件,特别是通过降低诱导剂浓度,可以有效降低上游生产成本,并保证高 EGFP 表达。