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[Construction of GPI-CTLA4Ig chimeric molecule and its expression on CHO-dhfr(-) cells]
Ji-Yun Yu1, Bei-Fen Shen, Yan Li
1Institute of Basic Medical Sciences, Academy of Military Medical Sciences, Beijing 100850,China.
Aim:
To develop a novel immunosuppressant GPI-CTLAIg modified by glycosyl-phosphatidylinositol(GPI).
Methods:
GPI-modified CTLAIg was produced by linking-up of CTLA4Ig with GPI-modification signal sequences from decay-accelerating factor (DAF). Chimeric molecule GPI-CTLA4Ig gene was cloned into eukaryotic expression vector pCI-dhfr. Using lipofectine-mediated gene transfer technique, pCI-GPI-CTLA4Ig was transfected into CHO-dhfr(-) cells, and the transfectants were screened by methotrexate (MTX). Expression of the recombinant protein was assessed by RT-PCR, ELISA, cell immunofluorescence staining and Western blot, and the purification of expressed protein was performed by protein A affinity chromatography.
Results:
The chimeric molecule GPI-CTLA4Ig has been constructed and stablely expressed on CHO-dhfr(-) cells.
Conclusion:
GPI-modified CTLAIg will may be used as novel immunosuppressant for suppressing reaction in graft rejection.
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