Characterization of hGalT-CTLA4IgP Expressed in Transgenic Rice Suspension Cell Cultures

초록

Transgenic plants are one of the promising protein production systems because they have several advantages including the capacity of post-translational modifications (PTMs). N-glycosylation is one of the most important PTMs related with the protein stability, serum half-life, and immune responses. Even though there are similarity in N-glycosylation mechanism between plant and mammalian cells, a little difference makes it difficult to develop therapeutic proteins in transgenic plant cells. Mammalian-derived glycoproteins have penultimate β1,4-galactose and terminal sialic acid residues. On the other hand, plant-derived glycoproteins contain plant specific terminal β1,3-galactose, β1,2-xylose and α1,3-fucose residues.1) In this study, transgenic rice cells were transformed with human β1,4-galactosyltransferase (hGalT) gene for the production of more humanized galactose-extended human cytotoxic T-lymphocyte antigen 4-immunoglobulin (hCTLA4IgP).2) To investigate the effect of hGalT expression on the glycan structure of the recombinant protein, purified hGalT-CTLA4IgP was analyzed by Western blot and lectin affinoblot method using antibodies and glycan-specific lectins.3) We confirmed that hGalT in transgenic rice cells is not only functionally very active but also efficiently able to change the glycan structure of the recombinant protein into a more human-compatible type. Moreover, it was possible to interact with sialic acid residues and terminal galactose residues from hGalT-CTLA4IgP by in vitro sialylation. From these results, it is expected that enhanced in vivo half-life and protein stability of hCTLA4IgP with humanized glycan structures as well as reduced potential of immune response from much lower contents of xylose and fucose.