PURIFICATION AND CHARACTERIZATION OF ALPHA(1)-ANTITRYPSIN SECRETED BYRECOMBINANT YEAST SACCHAROMYCES-DIASTATICUS

The secreted human alpha(1)-antitrypsin (alpha(1)AT) produced by yeast was purified from the culture medium by ultrafiltration, ammonium sul fate fractionation (60-75% saturation), protamine sulfate treatment, a nd ion-exchange chromatography. Molecular mass of the purified alpha(1 )AT was 52 kDa, which is similar to that of human plasma alpha(1)AT. Y east-produced alpha(1)AT was fully functional as an inhibitor compared with the plasma form. Unlike plasma alpha(1)AT, however, treatment of the yeast-produced alpha(1)AT with endoglycosidase H decreased the mo lecular mass to that of recombinant alpha(1)AT produced in Escherichia coli, indicating the high-mannose type N-linked glycosylation of the secreted alpha(1)AT. Glycosylation in yeast cells enhanced kinetic sta bility of alpha(1)AT towards heat deactivation.