Wan Aini, Xu Dongsheng, Liu Kedong, Peng Lin, Cai Yanfei, Chen Yun, He Yang, Yang Jianfeng, Jin Jian, Li Huazhong
a The Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology , Jiangnan University , Wuxi , China.
b Laboratory of Molecular Pharmacology, School of Pharmaceutical Sciences , Jiangnan University , Wuxi , China.
Prep Biochem Biotechnol. 2017 Aug 9;47(7):678-686. doi: 10.1080/10826068.2017.1303612. Epub 2017 Mar 10.
Insulin-like growth factor-1 (IGF-1) plays a crucial role in cell development, differentiation, and metabolism, and has been a potential therapeutic agent for many diseases. Chinese hamster ovary (CHO) cells are widely used for production of recombinant therapeutic proteins, but the expression level of IGF-1 in CHO cells is very low (1,500 µg/L) and the half-life of IGF-1 in blood circulation is only 4.5 min according to previous studies. Therefore, IGF-1 was fused to long-circulating serum protein human serum albumin (HSA) and expressed in CHO cells. After 8-day fed-batch culture, the expression level of HSA-IGF-1 reached 100 mg/L. The fusion protein HSA-IGF-1 was purified with a recovery of 35% using a two-step chromatographic procedure. According to bioactivity assay, the purified HSA-IGF-1 could stimulate the proliferation of NIH3T3 cells in a dose-dependent fashion and promote the cell-cycle progression. Besides this, HSA-IGF-1 could bind to IGF-1 receptor on cell membrane and activate the intracellular PI3K/AKT signaling pathway. Our study suggested that HSA fusion technology carried out in CHO cells not only provided bioactivity in HSA-IGF-1 for further research but also offered a beneficial strategy to produce other similar cytokines in CHO cells.
胰岛素样生长因子-1（IGF-1）在细胞发育、分化和代谢中发挥着关键作用，并且一直是许多疾病的潜在治疗药物。中国仓鼠卵巢（CHO）细胞被广泛用于生产重组治疗性蛋白质，但根据以往研究，IGF-1在CHO细胞中的表达水平非常低（1500μg/L），且其在血液循环中的半衰期仅为4.5分钟。因此，将IGF-1与人血清白蛋白（HSA）这种长循环血清蛋白融合，并在CHO细胞中表达。经过8天的补料分批培养，HSA-IGF-1的表达水平达到了100mg/L。采用两步色谱法纯化融合蛋白HSA-IGF-1，回收率为35%。根据生物活性测定，纯化后的HSA-IGF-1能够以剂量依赖的方式刺激NIH3T3细胞的增殖，并促进细胞周期进程。除此之外，HSA-IGF-1能够与细胞膜上的IGF-1受体结合，并激活细胞内的PI3K/AKT信号通路。我们的研究表明，在CHO细胞中进行的HSA融合技术不仅为进一步研究提供了具有生物活性的HSA-IGF-1，还为在CHO细胞中生产其他类似细胞因子提供了一种有益的策略。
