Stach Christopher S, McCann Meghan G, O'Brien Conor M, Le Tung S, Somia Nikunj, Chen Xinning, Lee Kyoungho, Fu Hsu-Yuan, Daoutidis Prodromos, Zhao Liang, Hu Wei-Shou, Smanski Michael
State Key Laboratory of Bioreactor Engineering , East China University of Science and Technology , 130 Meilong Road , Shanghai 200237 , P.R. China.
ACS Synth Biol. 2019 Nov 15;8(11):2524-2535. doi: 10.1021/acssynbio.9b00215. Epub 2019 Oct 18.
Chinese hamster ovary (CHO) cells are used for industrial production of protein-based therapeutics (i.e., "biologics"). Here we describe a method for combining systems-level kinetic models with a synthetic biology platform for multigene overexpression to rationally perturb N-linked glycosylation. Specifically, we sought to increase galactose incorporation on a secreted Immunoglobulin G (IgG) protein. We rationally design, build, and test a total of 23 transgenic cell pools that express single or three-gene glycoengineering cassettes comprising a total of 100 kilobases of engineered DNA sequence. Through iterative engineering and model refinement, we rationally increase the fraction of bigalactosylated glycans five-fold from 11.9% to 61.9% and simultaneously decrease the glycan heterogeneity on the secreted IgG. Our approach allows for rapid hypothesis testing and identification of synergistic behavior from genetic perturbations by bridging systems and synthetic biology.
中国仓鼠卵巢（CHO）细胞用于基于蛋白质的治疗药物（即“生物制剂”）的工业化生产。在此，我们描述了一种将系统水平动力学模型与用于多基因过表达的合成生物学平台相结合的方法，以合理扰动N-连接糖基化。具体而言，我们试图增加分泌型免疫球蛋白G（IgG）蛋白上半乳糖的掺入。我们合理设计、构建并测试了总共23个转基因细胞库，这些细胞库表达单基因或三基因糖工程盒，总共包含100千碱基的工程化DNA序列。通过迭代工程和模型优化，我们合理地将双半乳糖基化聚糖的比例从11.9%提高到61.9%，同时降低分泌型IgG上聚糖的异质性。我们的方法通过桥接系统生物学和合成生物学，实现了快速的假设检验和从基因扰动中识别协同行为。
