Abstract The high-yield production of vascular endothelial growth factor (VEGF), as a major therapeutic target in pathological angiogenesis and diabetic wound healing, provides critical advantages for in vitro studies. In the present study, to improve the soluble production of human VEGF8–109 (receptor-binding domain (RBD) of VEGF or VEGF RBD), at first VEGF 8-109 encoded gene was expressed in SHuffle T7 E. coli. Moreover, in two steps, the protein production was optimized based on Taguchi design, by evaluating optimal levels of various induction parameters, such as cell density in induction time, temperature, inducer concentration, and media components. The results indicated that the highest amount of the protein was achieved in TB medium containing glycerol 6 g L−1, peptone to yeast extract ratio 1:1, ethanol 3% and MgSO4 4 g L−1, under inducing with 0.05 mM IPTG in OD600 of 0.7 at 24 °C for 22 h. The bioactivity of the purified protein was confirmed by cell proliferation assay. Finally, bench-scale production of VEGF8–109 was performed under the optimum conditions and resulted in 182 mg of soluble VEGF8–109 expressed per liter. Totally, our results can be considered as a basis for economical production of the recombinant VEGF in future.

中文翻译：

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大肠杆菌中人VEGF8-109受体结合域的可溶性过表达、高水平生产和纯化

摘要 作为病理性血管生成和糖尿病伤口愈合的主要治疗靶点，血管内皮生长因子 (VEGF) 的高产量生产为体外研究提供了关键优势。在本研究中，为了提高人 VEGF8-109（VEGF 或 VEGF RBD 的受体结合域（RBD））的可溶性生产，首先在 SHuffle T7 大肠杆菌中表达 VEGF 8-109 编码基因。此外，在两个步骤中，通过评估各种诱导参数的最佳水平，如诱导时间、温度、诱导剂浓度和培养基成分中的细胞密度，优化了基于田口设计的蛋白质生产。结果表明，在含有甘油 6 g L-1、蛋白胨与酵母提取物比例 1:1、乙醇 3% 和 MgSO4 4 g L-1 的 TB 培养基中获得最高量的蛋白质，在 OD600 为 0.7 的情况下，在 24 °C 下用 0.05 mM IPTG 诱导 22 小时。纯化蛋白的生物活性通过细胞增殖试验得到证实。最后，在最佳条件下进行 VEGF8-109 的小规模生产，每升表达 182 mg 可溶性 VEGF8-109。总的来说，我们的结果可以被认为是未来经济生产重组VEGF的基础。