RECOMBINANT PROTEINASE-3 (WREGENERS ANTIGEN) EXPRESSED IN PICHIA-PASTORIS IS FUNCTIONALLY ACTIVE AND IS RECOGNIZED BY PATIENT SERA

The open reading frame of human proteinase 3 (PR3) without the prepro- peptide was cloned and expressed in Escherichia coli (rcPR3) and in Pi chia pastoris (rpPR3). The 6-histidine tagged rpPR3 was efficiently se creted into culture supernatant from which it could be purified by imm obilized metal chelate chromatography. Purified rpPR3 migrated as a si ngle 32-kD band on SDS-PAGE and harboured protease activity that could be inhibited with inhibitors specific br serine-proteases. By indirec t antigen-capture ELISA using rpPR3, 60% of sera from patients with We gener's granulomatosis bound to the recombinant product, although it w as not recognized in ELISA with directly coated rpPR3.