Expression system for purification of secreted proteins

Lentiviral expression system for the purification of secreted proteins from human cell cultures

Recombinant proteins of therapeutic use are ideally produced in human cells to ensure appropriate co- and post-translational modifications. CHO cells are mostly used but recent studies highlight that proteins produced in this cell line have a different glycosylation pattern than the same proteins produced in HEK cells, which may affect their function. While the exact implications of differences in glycosylation are not yet fully understood, a strong argument for the production of therapeutic proteins in HEK cells is made by the finding that CHO cells can add terminal galactose-α-1,3-galactose (α-Gal) to proteins. α-Gal is a non-human antigenic glycan and its reaction with circulating antibodies present in most individuals can lead to anaphylaxis. This glycan is absent in proteins produced in HEK cells. In this related study, scientists describe a simple and effective method for the purification of a His-tagged human protein from the culture media of lentiviral vector-transduced HEK 293 T cells. All reagents are readily available at a relatively low cost and no specialized equipment is needed to complete the procedure. Using soluble CD4 (sCD4), a truncated version of the CD4 receptor that is secreted, they showed that lentiviral vector mediated over 10-fold higher secretion of the protein in comparison to 293 T cells. Furthermore, small-scale purification from 50 ml of culture media with reduced serum content yielded up to 1 mg of pure sCD4.

The procedure outlined in this study can be performed without the need for specialized reagents or equipment and could easily be adapted by any laboratory. This strategy can be adapted to a large-scale format without significantly increasing the cost to further improve the yield.


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