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KMID : 1094720100150030467
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Biotechnology and Bioprocess Engineering
2010 Volume.15 No. 3 p.467 ~ p.475
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Overexpression of Acid Protease of Saccharomycopsis fibuligera in Yarrowia lipolytica and Characterization of the Recombinant Acid Protease for Skimmed Milk Clotting
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Yu Xin-Jun
Chi Zhen-Ming
Li Jing
Li Hui-Juan
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Abstract
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The gene encoding an acid protease natively produced by Saccharomycopsis fibuligera was cloned and overexpressed in Yarrowia lipolytica and the resultant recombinant acid protease was purified and characterized. The molecular mass of the purified enzyme was estimated as 94.8 kDa by gel filtration chromatography. The optimal pH and temperature of the purified acid protease were 3.5 and 33oC, respectively, and the enzyme was very stable over a pH range of 1.0 ~ 3.0. The recombinant acid protease was activated by Zn2+, but was inhibited by Hg2+, Fe2+, Fe3+, and Mg2+, EDTA, EGTA, iodoacetic acid, and pepstatin. The purified recombinant acid protease from the positive transformant 71 had high milk clotting activity, suggesting that it may be used as a rennet substitute in the cheese industry.
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KEYWORD
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Saccharomycopsis fibuligera, Yarrowia lipolytica, acid protease, milk clotting activity
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