PURIFICATION AND CHARACTERIZATION OF ALPHA(1)-ANTITRYPSIN SECRETED BY RECOMBINANT YEAST SACCHAROMYCES-DIASTATICUS
PURIFICATION AND CHARACTERIZATION OF ALPHA(1)-ANTITRYPSIN SECRETED BY RECOMBINANT YEAST SACCHAROMYCES-DIASTATICUS
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DOI：
10.1016/0168-1656(95)00079-6
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发表时间：
1995-10-16
影响因子：
4.1
通讯作者：
YU, MH
中科院分区：
文献类型：
--
作者：
KWON, KS;SONG, MY;YU, MH
通过超滤、硫酸铵分级分离（60-75%饱和度）、硫酸鱼精蛋白处理和离子交换色谱法从培养基中纯化酵母产生的分泌型人α（1）-抗胰蛋白酶（α（1）AT）。纯化的α（1）AT的分子量为52 kDa，这与人血浆α（1）AT的分子量相似。与血浆形式相比，酵母产生的α（1）AT作为抑制剂具有完全功能。然而，与血浆α 1 AT不同，用内切糖苷酶H处理酵母产生的α 1 AT，使其分子量降低至大肠杆菌产生的重组α 1 AT的分子量，表明分泌的α 1 AT的高甘露糖型N-连接糖基化。酵母细胞中的糖基化增强了α（1）AT对热失活的动力学稳定性。
The secreted human alpha(1)-antitrypsin (alpha(1)AT) 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(1)AT was 52 kDa, which is similar to that of human plasma alpha(1)AT. Yeast-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 molecular 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 stability of alpha(1)AT towards heat deactivation.