Chen Chuan, Wang Jie, Zhao Yun, Chen Shaopeng, Hu Zhishang, Chen Long, Hang Haiying
Key Laboratory for Protein and Peptide Pharmaceuticals, National Laboratory of Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China.
University of Chinese Academy of Sciences, No.19(A) Yuquan Road, Shijingshan District, Beijing 100049, China.
Antibodies (Basel). 2018 Dec 13;7(4):42. doi: 10.3390/antib7040042.
The induction of somatic hypermutation (SHM) in various cell lines by activation-induced cytidine deaminase (AID) has been used in protein-directed selection, especially in antibody affinity maturation. Several antibody affinity maturation systems based on mammalian cells have been developed in recent years, i.e., 293T, H1299, Raji and CHO cells. However, the efficiency of in vitro AID-induced hypermutation is low, restricting the application of such systems. In this study, we examined the role of Ig and Ek enhancers in enhancing SHM in the episomal vector pCEP4 that expresses an anti-high mobility group box 1 (HMGB1) full-length antibody. The plasmid containing the two enhancers exhibited two-fold improvement of mutation rate over pCEP4 in an AID expression H1299 cell line (H1299-AID). With the engineered episomal vector, we improved the affinity of this antibody in H1299-AID cells by 20-fold.
通过激活诱导胞苷脱氨酶（AID）在各种细胞系中诱导体细胞高频突变（SHM）已用于蛋白质定向筛选，尤其是在抗体亲和力成熟方面。近年来，已经开发了几种基于哺乳动物细胞的抗体亲和力成熟系统，即293T、H1299、Raji和CHO细胞。然而，体外AID诱导的高频突变效率较低，限制了此类系统的应用。在本研究中，我们检测了Ig和Ek增强子在表达抗高迁移率族蛋白盒1（HMGB1）全长抗体的游离载体pCEP4中增强SHM的作用。在AID表达的H1299细胞系（H1299-AID）中，含有这两种增强子的质粒与pCEP4相比，突变率提高了两倍。利用工程化的游离载体，我们将该抗体在H1299-AID细胞中的亲和力提高了20倍。
