The tobacco phosphatidylethanolamine‐binding protein NtFT4 simultaneously improves vitality, growth, and protein yield in human cells

• Published in: Biotechnology and bioengineering 2021-10, Vol.118 (10), p.3770-3786
• Main Authors: Kronenberg, Julia, Schrödter, Katrin, Noll, Gundula A., Twyman, Richard M., Prüfer, Dirk, Känel, Philip
• Format: Article
• Language: English
• Subjects: Antibodies; Biopharmaceuticals; Cell culture; Cell Culture Techniques; Cell density; Cell growth; cell line engineering; Cell lines; Cell proliferation; Cell Survival; Culture media; Density; Fluorescent indicators; Genetic transformation; HEK293 Cells; Humans; insulin signaling; Mammalian cells; MCF-7 Cells; Monolayers; Nicotiana - genetics; Phosphatidylethanolamine; Phosphatidylethanolamine Binding Protein - biosynthesis; Phosphatidylethanolamine Binding Protein - genetics; Plant Proteins - biosynthesis; Plant Proteins - genetics; Productivity; proliferation; Protein expression; Proteins; recombinant protein expression; Recombinant Proteins - biosynthesis; Recombinant Proteins - genetics; Tobacco; Transfection
• ISSN: 0006-3592; 1097-0290
• DOI: 10.1002/bit.27853

The production of biopharmaceutical proteins in mammalian cells by transient expression or stable transformation requires robust and viable cells. Cell line engineering must therefore balance improved cell growth and viability with high productivity. We tested the ability of nonmammalian phosphatidylethanolamine‐binding proteins to enhance cell proliferation in monolayers and suspension cultures. The tobacco protein NtFT4 improved the proliferation of multiple human cell lines. Viable cell density is usually impaired by efficient transfection, but we found that the number of HEK‐293TNtFT4 cells at the peak of protein expression was twice that of standard HEK‐293T cells, and the antibody yield increased by approximately one‐third. Improved growth and viability were observed in different cell lines, in different culture media, and also after transient transfection, suggesting the beneficial trait is consistent and transferable. Additional modifications could boost the productivity of high‐density HEK‐293TNtFT4 cells even further as we showed for a fluorescent marker protein and recombinant antibody expressed in monolayer cultures. The HEK‐293TNtFT4 cell line provides a new human model platform that increases cell proliferation, also achieving a fundamental improvement in recombinant protein expression. The authors describe the ability of a non‐mammalian phosphatidylethanolamine‐binding protein to improve recombinant protein production by human cells. Stable transfection with tobacco NtFT4 increased the proliferation rate of human cell lines, and the transfection efficiency of HEK‐293TNtFT4 cells remained equivalent to the control HEK‐293 line. High viable cell densities combined with high transfection efficiencies resulted in 30% higher IgG1 yields during transient expression, and additional process optimization could improve these yields even further.