Purification and characterization of α₁-antitrypsin secreted by recombinant yeast Saccharomyces diastaticus

Abstract

The secreted human α1-antitrypsin (α1AT) produced by yeast was purified from the culture medium by ultrafiltration, ammonium sulfate fractionation (60-75% saturation), protamine sulfate treatment, and ion-exchange chromatography. Molecular mass of the purified α1AT was 52 kDa, which is similar to that of human plasma α1AT. Yeast-produced α1AT was fully functional as an inhibitor compared with the plasma form. Unlike plasma (α1AT, however, treatment of the yeast-produced α1AT with endoglycosidase H decreased the molecular mass to that of recombinant α1AT produced in Escherichia coli, indicating the high-mannose type N-linked glycosylation of the secreted α1AT. Glycosylation in yeast cells enhanced kinetic stability of α1AT towards heat deactivation.