A novel simple method to purify recombinant soluble human complement receptor type 1 (sCR1) from CHO cell culture

The human complement receptor type 1 (CR1, C3 b/C4b receptor) is a polymorphic membrane glycoprotein expressed on human erythrocytes, peripheral leukocytes, plasma and renal glomerular podocytes, which consists of transmembrane and cytoplasmic domains with 30 repeating homologous protein domains knows as short consensus repeats (SCR). CR1 has been used as an inhibitor for inflammatory and immune system for the past several years. Recently, it is reported that CR1 was found to suppress the hyper-acute rejection in xeno-transplantation and can be used to cure autoimmune diseases. A soluble form of CR1, called sCR1, is a recombinant CR1 by cleaving the transmembrane domain at C-terminus and has been expressed in Chinese Hamster Ovary (CHO) cells. Several purification methods for sCR1 from CHO cells have been reported, but most of them require complicated steps at high cost. Moreover, such methods are mostly performed under the pH condition apt to denaturing sCR1 and causes sCR1 losing its activity. We here report a rapid and efficient method to purify sCR1 from CHO cell. The new method consists of a two-stage of cell culture by cultivating cells in serum medium followed by serum-free medium, and a two-stage of column purification by means of heparin and gel filtration column chromatography. By using this novel method, sCR1 can be purified in a simple and effective way with high yield and purity. Furthermore, the purified sCR1 was confirmed to retain its activity to suppress the complement activationin vivo andex vivo.[PUBLICATION ABSTRACT]