Abstract
In production of recombinant proteins for biopharmaceuticals, N-glycosylation is often important for protein efficacy and patient safety. IgG with agalactosylated (G0)-N-glycans can improve the activation of the lectin-binding complement system and be advantageous in the therapy of lupus and virus diseases. In this study, the authors aimed to engineer CHO-S cells for the production of proteins with G0-N-glycans by targeting B4Gal-T isoform genes with CRISPR/Cas9. Indel mutations in genes encoding B4Gal-T1, -T2, and -T3 with and without a disrupted B4Gal-T4 sequence resulted in only ≈1% galactosylated N-glycans on total secreted proteins of 3-4 clones per genotype. The authors revealed that B4Gal-T4 is not active in N-glycan galactosylation in CHO-S cells. In the triple-KO clones, transiently expressed erythropoietin (EPO) and rituximab harbored only ≈6% and ≈3% galactosylated N-glycans, respectively. However, simultaneous disruption of B4Gal-T1 and -T3 may decrease cell growth. Altogether, the authors present the advantage of analyzing total secreted protein N-glycans after disrupting galactosyltransferases, followed by expressing recombinant proteins in selected clones with desired N-glycan profiles at a later stage. Furthermore, the authors provide a cell platform that prevalently glycosylates proteins with G0-N-glycans to further study the impact of agalactosylation on different in vitro and in vivo functions of recombinant proteins.
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Disclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.