Selecting cells expressing high levels of recombinant proteins using the GPI-anchored protein with selenocysteine system.

Yi-Shi Liu, Emmanuel Matabaro, Xiao-Dong Gao, Morihisa Fujita

Journal of bioscience and bioengineering 2021 Mar

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Most biopharmaceutical proteins are produced in mammalian cells because they have the advantageous capacity for protein folding, assembly, and posttranslational modifications. To satisfy the increasing demand for these proteins for clinical purposes and studies, traditional methods to improve protein productivity have included gene amplification, host cell engineering, medium optimization, and screening methods. However, screening and selection of high-producing cell lines remain complex and time consuming. In this study, we established a glycosylphosphatidylinositol (GPI)-anchored protein with a selenocysteine (GPS) system to select cells producing high levels of target secretory proteins. Recombinant lysosomal acid lipase (LIPA) and α-galactosidase A (GALA) were fused with a GPI attachment signal sequence and a selenocysteine insertion sequence after an in-frame UGA codon. Under these conditions, most of the recombinant proteins were secreted into the culture medium, but some were found to be GPI-anchored proteins on the cell surface. When sodium selenite was supplied into the culture medium, the amount of GPI-anchored LIPA and GALA was increased. High-expressing cells were selected by detecting surface GPI-anchored LIPA. The GPI-anchored protein was then eliminated by knocking out the GPI biosynthesis gene PIGK, in these cells, all LIPA was in secreted form. Our system provides a promising method of isolating cells that highly express recombinant proteins from large cell populations. Copyright © 2020 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

Citation

Yi-Shi Liu, Emmanuel Matabaro, Xiao-Dong Gao, Morihisa Fujita. Selecting cells expressing high levels of recombinant proteins using the GPI-anchored protein with selenocysteine system. Journal of bioscience and bioengineering. 2021 Mar;131(3):225-233