Purification and characterization of alpha 1-antitrypsin secreted by recombinant yeast Saccharomyces diastaticus.
The secreted human alpha 1-antitrypsin (alpha 1AT) 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 alpha 1AT was 52 kDa, which is similar to that of human plasma alpha 1AT. Yeast-produced alpha 1AT was fully functional as an inhibitor compared with the plasma form. Unlike plasma alpha 1AT, however, treatment of the yeast-produced alpha 1AT with endoglycosidase H decreased the molecular mass to that of recombinant alpha 1AT produced in Escherichia coli, indicating the high-mannose type N-linked glycosylation of the secreted alpha 1AT. Glycosylation in yeast cells enhanced kinetic stability of alpha 1AT towards heat deactivation.
Kwon
K S
KS
Korea Research Institute of Bioscience and Biotechnology, Korea Institute of Science and Technology, Taejon, South Korea.
Song
M
M
Yu
M H
MH
D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D006031:Glycosylation; D006358:Hot Temperature; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D011489:Protein Denaturation; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D012440:Saccharomyces / Q000235:genetics*; D000515:alpha 1-Antitrypsin / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*
D011994:Recombinant Proteins; D000515:alpha 1-Antitrypsin