Co-overexpression of Mgat1 and Mgat4 in CHO cells for production of highly sialylated albumin-erythropoietin

Publication date: Available online 6 May 2017 Source:Enzyme and Microbial Technology Author(s): Hyun-Myoung Cha, Jin-Hyuk Lim, Jung-Heum Yeon, Jeong-Min Hwang, Dong-Il Kim Terminal sialic acids on N-glycan of recombinant human erythropoietin are very important for in vivo half-life, as this glycoprotein has three N-glycosylation sites. N-acetylglucosaminyltransferases I, II, IV, and V (i.e. Mgat1, Mgat2, Mgat4, and Mgat5) catalyze the formation of a glycan antennary structure. These enzymes display different reaction kinetics for a common substrate and generally show low expression in Chinese hamster ovary (CHO) cells. Therefore, genetic control of Mgat expression is an effective method to increase sialic acid contents by enhancing glycan antennarity. To produce highly sialylated albumin-erythropoietin (Alb-EPO), we co-overexpressed the Mgat1 and Mgat4 genes in CHO cells and determined the optimal ratio of Mgat1:Mgat4 gene expression. All transfected cell lines showed increased gene expression of Mgat4, including Mgat1 overexpressing cell line. Sialic acid content of Alb-EPO was highest in co-transfected cells with excess Mgat4 gene, and these cells showed a higher tri- and tetra-antennary structure than control cells. Based on these results, we suggest that co-transfection of the Mgat1 and Mgat4 genes at a ratio of 2:8 is optimal for extension of antennary structures. Also, regulation of Mgat gene expression in the glycan biosynthesis pathway can be a novel approach t...
Source: Enzyme and Microbial Technology - Category: Biotechnology Source Type: research