EXPRESSION OF HUMAN PROTEINASE 3 IN CHINESE HAMSTER OVARY CELLS (CHO-CELLS)

Proteinase 3(PR3) is a human polymorphonuclear leukocyte serine proteinase and is the main target antigen for antineutrophil cytoplasmic antibodies (ANCA) found in Wegener's granulomatosis (WG). We developed a stable expression system for conformationally intact recombinant PR3 (rPR3) in Chinese hamster ovary cells (CHO-cells). The part of PR3 cDNA that encoded the active form of PR3 was selected by using appropriate primers, and a signal sequence was also added in front ofPR3 eDNA. The signal sequencePR3 (S-PR3) was cloned into the pME 18 expression vector and the result product was electroporated into E. coli (DH5 a strain). After isolation and purification, the presence of pMEI8-S-PR3 was confirmed by using appropriate restriction endonuclease and agarose gel electrophoresis. The pMEI8- S-PR3 was electroporated with CHO-cells and the presence of rPR3 was tested in culture medium after 10 days. There was 12 ng/mL rPR3 in culture medium that had activity and was recognized by ANCA in ELISA.