Heterologous expression and purification of protopectinase-N from Bacillus subtilis in Pichia pastoris

被引:9
作者
Liu, ZM [1 ]
Lu, ZX [1 ]
Lv, FX [1 ]
Bie, XM [1 ]
Zhao, HZ [1 ]
机构
[1] Nanjing Agr Univ, Coll Food Sci & Technol, Nanjing 210095, Jiangsu, Peoples R China
关键词
Bacillus subtilis; heterologous expression; protopectinase-N; purification; Pichia pastoris;
D O I
10.1016/j.procbio.2005.10.027
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
A eukaryotic expression system for secretory production of the protopectinase from Bacillus subtilis XZ2 in Pichia Pastoris was constructed in this paper. The protopectinase gene without native signal sequence was amplified by PCR and fused to the pPICZ alpha A plasmid containing alpha-factor sequence encoding secretion signal from Saccharomyces cerevisiae. The heterologous gene was expressed under the AOXI promoter in a culture of P pastoris X-33 in flasks. The recombinant protein was further purified by ammonium sulfate precipitate, Sephadex G75 and ion exchange chromatography and resulted in 18.91-fold purification; the purified protein was a molecular weight about 43 kDa and had an activity of 1948.64 U/Mg. (c) 2005 Elsevier Ltd. All rights reserved.
引用
收藏
页码:975 / 979
页数:5
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